Wild Crop Relatives:

Genomic and Breeding Resources

.
Chittaranjan Kole
Editor

Wild Crop Relatives:

Genomic and Breeding
Resources
Millets and Grasses
Editor
Prof. Chittaranjan Kole
Director of Research
Institute of Nutraceutical Research
Clemson University
109 Jordan Hall
Clemson, SC 29634
[email protected]

ISBN 978-3-642-14254-3 e-ISBN 978-3-642-14255-0

DOI 10.1007/978-3-642-14255-0
Springer Heidelberg Dordrecht London New York

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 Dedication

Dr. Norman Ernest Borlaug,1

the Father of Green Revolution,
is well respected for his contri-
butions to science and society.
There was or is not and never
will be a single person on this
Earth whose single-handed ser-
vice to science could save
millions of people from death
due to starvation over a period
of over four decades like Dr.
Borlaug’s. Even the Nobel Peace
Prize he received in 1970 does
not do such a great and noble
person as Dr. Borlaug justice.
His life and contributions are
well known and will remain in
the pages of history of science.
I wish here only to share some
facets of this elegant and ideal
personality I had been blessed to
observe during my personal inter-
actions with him.
It was early 2007 while I was
at the Clemson University as a
visiting scientist one of my lab
colleagues told me that “some-
body wants to talk to you; he
appears to be an old man”. I took the telephone receiver casually and said hello.
The response from the other side was – “I am Norman Borlaug; am I talking to
Chitta?” Even a million words would be insufficient to define and depict the exact
feelings and thrills I experienced at that moment!

1
The photo of Dr. Borlaug was kindly provided by Julie Borlaug (Norman Borlaug Institute for International Agriculture, Texas
A&M Agriculture) the granddaughter of Dr. Borlaug.

v
vi Dedication

I had seen Dr. Borlaug only once, way back in 1983, when he came to New Delhi,
India to deliver the Coromandal Lecture organized by Prof. M.S. Swaminathan on the
occasion of the 15th International Genetic Congress. However, my real interaction
with him began in 2004 when I had been formulating a 7-volume book series entitled
Genome Mapping and Molecular Breeding in Plants. Initially, I was neither confident
of my ability as a series/book editor nor of the quality of the contents of the book
volumes. I sent an email to Dr. Borlaug attaching the table of contents and the
tentative outline of the chapters along with manuscripts of only a few sample chapters,
including one authored by me and others, to learn about his views as a source of
inspiration (or caution!) I was almost sure that a person of his stature would have no
time and purpose to get back to a small science worker like me. To my utter (and
pleasant) surprise I received an email from him that read: “May all Ph.D.’s, future
scientists, and students that are devoted to agriculture get an inspiration as it refers to
your work or future work from the pages of this important book. My wholehearted
wishes for a success on your important job”. I got a shot in my arm (and in mind for
sure)! Rest is a pleasant experience – the seven volumes were published by Springer in
2006 and 2007, and were welcome and liked by students, scientists and their societies,
libraries, and industries. As a token of my humble regards and gratitude, I sent
Dr. Borlaug the Volume I on Cereals and Millets that was published in 2006. And
here started my discovery of the simplest person on Earth who solved the most
complex and critical problem of people on it – hunger and death.
Just one month after receiving the volume, Dr. Borlaug called me one day and
said, “Chitta, you know I cannot read a lot now-a-days, but I have gone through only
on the chapters on wheat, maize and rice. Please excuse me. Other chapters of this
and other volumes of the series will be equally excellent, I believe”. He was highly
excited to know that many other Nobel Laureates including Profs. Arthur Kornberg,
Werner Arber, Phillip Sharp, G€ unter Blobel, and Lee Hartwell also expressed
generous comments regarding the utility and impact of the book series on science
and the academic society. While we were discussing many other textbooks and
review book series that I was editing at that time, again in my night hours for the
benefit of students, scientists, and industries, he became emotional and said to me,
“Chitta, forget about your original contributions to basic and applied sciences, you
deserved Nobel Prize for Peace like me for providing academic foods to millions of
starving students and scientists over the world particularly in the developing
countries. I will recommend your name for the World Food Prize, but it will not do
enough justice to the sacrifice you are doing for science and society in your sleepless
nights over so many years. Take some rest Chitta and give time to Phullara, Sourav
and Devleena” (he was so particular to ask about my wife and our kids during most of
our conversations). I felt honored but really very ashamed as I am aware of my
almost insignificant contribution in comparison to his monumental contribution and
thousands of scientists over the world are doing at least hundred-times better jobs
than me as scientist or author/editor of books! So, I was unable to utter any words for
a couple of minutes but realized later that he must been too affectionate to me and his
huge affection is the best award for a small science worker as me!
In another occasion he wanted some documents from me. I told him that I will
send them as attachments in emails. Immediately he shouted and told me:
“You know, Julie (his granddaughter) is not at home now and I cannot check email
myself. Julie does this for me. I can type myself in type writer but I am not good in
computer. You know what, I have a xerox machine and it receives fax also. Send me
Dedication vii

the documents by fax”. Here was the ever-present child in him. Julie emailed me later
to send the documents as attachment to her as the ‘xerox machine’ of Dr. Borlaug ran
out of ink!
Another occasion is when I was talking with him in a low voice, and he immedi-
ately chided me: “You know that I cannot hear well now-a-days; I don’t know where
Julie has kept the hearing apparatus, can’t you speak louder?” Here was the fatherly
figure who was eager to hear each of my words!
I still shed tears when I remember during one of our telephone conversations he
asked: “You know I have never seen you, can you come to Dallas in the near future by
chance?” I remember we were going through a financial paucity at that time and I could
not make a visit to Dallas (Texas) to see him, though it would have been a great honor.
In late 2007, whenever I tried to talk to Dr. Borlaug, he used to beckon Julie to
bring the telephone to him, and in course of time Julie used to keep alive all the
communications between us when he slowly succumbed to his health problems.
The remaining volumes of the Genome Mapping and Molecular Breeding in
Plants series were published in 2007, and I sent him all the seven volumes. I wished
to learn about his views. During this period he could not speak and write well. Julie
prepared a letter based on his words to her that read: “Dear Chitta, I have reviewed
the seven volumes of the series on Genome Mapping and Molecular Breeding in
Plants, which you have authored. You have brought together genetic linkage maps
based on molecular markers for the most important crop species that will be a
valuable guide and tool to further molecular crop improvements. Congratulations
for a job well done”.
During one of our conversations in mid-2007, he asked me what other book
projects I was planning for Ph.D. students and scientists (who had always been his
all-time beloved folks). I told him that the wealth of wild species already utilized and
to be utilized for genetic analysis and improvement of domesticated crop species
have not been deliberated in any book project. He was very excited and told me to
take up the book project as soon as possible. But during that period I had a huge
commitment to editing a number of book volumes and could not start the series he
was so interested about.
His sudden demise in September 2009 kept me so morose for a number of months
that I could not even communicate my personal loss to Julie. But in the meantime, I
formulated a 10-volume series on Wild Crop Relatives: Genomic and Breeding
Resources for Springer. And whom else to dedicate this series to other than Dr. Borlaug!
I wrote to Julie for her formal permission and she immediately wrote me: “Chitta,
Thank you for contacting me and yes I think my grandfather would be honored with
the dedication of the series. I remember him talking of you and this undertaking quite
often. Congratulations on all that you have accomplished!” This helped me a lot as
I could at least feel consoled that I could do a job he wanted me to do and I will
always remain grateful to Julie for this help and also for taking care of Dr. Borlaug,
not only as his granddaughter but also as the representative of millions of poor people
from around the world and hundreds of plant and agricultural scientists who try to
follow his philosophy and worship him as a father figure.
It is another sad experience of growing older in life that we walk alone and miss
the affectionate shadows, inspirations, encouragements, and blessings from the
fatherly figures in our professional and personal lives. How I wish I could treat my
next generations in the same way as personalities like Mother Teresa and Dr. Norman
Borlaug and many other great people from around the world treated me!
viii Dedication

During most of our conversations he used to emphasize on the immediate impact

of research on the society and its people. A couple of times he even told me that my
works on molecular genetics and biotechnology, particularly of 1980s and 1990s,
have high fundamental importance, but I should also do some works that will benefit
people immediately. This advice elicited a change in my thoughts and workplans and
since then I have been devotedly endeavoring to develop crop varieties enriched with
phytomedicines and nutraceuticals. Borlaug influenced both my personal and pro-
fessional life, particularly my approach to science, and I dedicate this series to him in
remembrance of his great contribution to science and society and for all his personal
affection, love and blessings for me.
I emailed the above draft of the dedication page to Julie for her views and I wish to
complete my humble dedication with great satisfaction with the words of Julie who
served as the living ladder for me to reach and stay closer to such as great human
being as Dr. Borlaug and express my deep regards and gratitude to her. Julie’s email
read: “Chitta, Thank you for sending me the draft dedication page. I really enjoyed
reading it and I think you captured my grandfather’s spirit wonderfully. . .. So thank
you very much for your beautiful words. I know he would be and is honored.”

Clemson, USA Chittaranjan Kole

Preface

Wild crop relatives have been playing enormously important roles both in the
depiction of plant genomes and the genetic improvement of their cultivated counter-
parts. They have contributed immensely to resolving several fundamental questions,
particularly those related to the origin, evolution, phylogenetic relationship, cytolog-
ical status and inheritance of genes of an array of crop plants; provided several
desirable donor genes for the genetic improvement of their domesticated counter-
parts; and facilitated the innovation of many novel concepts and technologies while
working on them directly or while using their resources. More recently, they have
even been used for the verification of their potential threats of gene flow from
genetically modified plants and invasive habits. Above all, some of them are con-
tributing enormously as model plant species to the elucidation and amelioration of
the genomes of crop plant species.
As a matter of fact, as a student, a teacher, and a humble science worker I was, still
am and surely will remain fascinated by the wild allies of crop plants for their
invaluable wealth for genetics, genomics and breeding in crop plants and as such
share a deep concern for their conservation and comprehensive characterization for
future utilization. It is by now a well established fact that wild crop relatives deserve
serious attention for domestication, especially for the utilization of their phytomedi-
cines and nutraceuticals, bioenergy production, soil reclamation, and the phytoreme-
diation of ecology and environment. While these vastly positive impacts of wild crop
relatives on the development and deployment of new varieties for various purposes in
the major crop plants of the world agriculture, along with a few negative potential
concerns, are envisaged the need for reference books with comprehensive delibera-
tions on the wild relatives of all the major field and plantation crops and fruit and
forest trees is indeed imperative. This was the driving force behind the inception and
publication of this series.
Unlike the previous six book projects I have edited alone or with co-editors, this
time it was very difficult to formulate uniform outlines for the chapters of this book
series for several obvious reasons. Firstly, the status of the crop relatives is highly
diverse. Some of them are completely wild, some are sporadically cultivated and
some are at the initial stage of domestication for specific breeding objectives recently
deemed essential. Secondly, the status of their conservation varies widely: some have
been conserved, characterized and utilized; some have been eroded completely
except for their presence in their center(s) of origin; some are at-risk or endangered
due to genetic erosion, and some of them have yet to be explored. The third constraint
is the variation in their relative worth, e.g. as academic model, breeding resource,
and/or potential as “new crops.”

ix
x Preface

The most perplexing problem for me was to assign the chapters each on a
particular genus to different volumes dedicated to crop relatives of diverse crops
grouped based on their utility. This can be exemplified with Arabidopsis, which has
primarily benefited the Brassicaceae crops but also facilitated genetic analyses and
improvement in crop plants in other distant families; or with many wild relatives of
forage crops that paved the way for the genetic analyses and breeding of some major
cereal and millet crops. The same is true for wild crop relatives such as Medicago
truncatula, which has paved the way for in-depth research on two crop groups of
diverse use: oilseed and pulse crops belonging to the Fabaceae family. The list is too
long to enumerate. I had no other choice but to compromise and assign the genera of
crop relatives in a volume on the crop group to which they are taxonomically the
closest and to which they have relatively greater contributions. For example, I placed
the chapter on genus Arabidopsis in the volume on oilseeds, which deals with the
wild relatives of Brassicaceae crops amongst others.
However, we have tried to include deliberations pertinent to the individual genera
of the wild crop relatives to which the chapters are devoted. Descriptions of the
geographical locations of origin and genetic diversity, geographical distribution,
karyotype and genome size, morphology, etc. have been included for most of
them. Their current utility status – whether recognized as model species, weeds,
invasive species or potentially cultivable taxa – is also delineated. The academic,
agricultural, medicinal, ecological, environmental and industrial potential of both the
cultivated and/or wild allied taxa are discussed.
The conservation of wild crop relatives is a much discussed yet equally neglected
issue albeit the in situ and ex situ conservations of some luckier species were initiated
earlier or are being initiated now. We have included discussions on what has
happened and what is happening with regard to the conservation of the crop relatives,
thanks to the national and international endeavors, in most of the chapters and also
included what should happen for the wild relatives of the so-called new, minor,
orphan or future crops.
The botanical origin, evolutionary pathway and phylogenetic relationship of crop
plants have always attracted the attention of plant scientists. For these studies
morphological attributes, cytological features and biochemical parameters were
used individually or in combinations at different periods based on the availability
of the required tools and techniques. Access to different molecular markers based
on nuclear and especially cytoplasmic DNAs that emerged after 1980 refined the
strategies required for precise and unequivocal conclusions regarding these aspects.
Illustrations of these classical and recent tools have been included in the chapters.
Positioning genes and defining gene functions required in many cases different
cytogenetic stocks, including substitution lines, addition lines, haploids, monoploids
and aneuploids, particularly in polyploid crops. These aspects have been dealt in the
relevant chapters. Employment of colchiploidy, fluorescent or genomic in situ
hybridization and Southern hybridization have reinforced the theoretical and applied
studies on these stocks. Chapters on relevant genera/species include details on these
cytogenetic stocks.
Wild crop relatives, particularly wild allied species and subspecies, have been
used since the birth of genetics in the twentieth century in several instances such as
studies of inheritance, linkage, function, transmission and evolution of genes. They
have been frequently used in genetic studies since the advent of molecular markers.
Their involvement in molecular mapping has facilitated the development of mapping
Preface xi

populations with optimum polymorphism to construct saturated maps and also

illuminating the organization, reorganization and functional aspects of genes and
genomes. Many phenomena such as genomic duplication, genome reorganization,
self-incompatibility, segregation distortion, transgressive segregation and defining
genes and their phenotypes have in many cases been made possible due to the
utilization of wild species or subspecies. Most of the chapters contain detailed
elucidations on these aspects.
The richness of crop relatives with biotic and abiotic stress resistance genes was
well recognized and documented with the transfer of several alien genes into their
cultivated counterparts through wide or distant hybridization with or without
employing embryo-rescue and mutagenesis. However, the amazing revelation that
the wild relatives are also a source of yield-related genes is a development of the
molecular era. Apomictic genes are another asset of many crop relatives that deserve
mention. All of these past and the present factors have led to the realization that the
so-called inferior species are highly superior in conserving desirable genes and can
serve as a goldmine for breeding elite plant varieties. This is particularly true at a
point when natural genetic variability has been depleted or exhausted in most of the
major crop species, particularly due to growing and promoting only a handful of
so-called high-yielding varieties while disregarding the traditional cultivars and
landraces. In the era of molecular breeding, we can map desirable genes and poly-
genes, identify their donors and utilize tightly linked markers for gene introgression,
mitigating the constraint of linkage drag, and even pyramid genes from multiple
sources, cultivated or wild taxa. The evaluation of primary, secondary and tertiary
gene pools and utilization of their novel genes is one of the leading strategies in
present-day plant breeding. It is obvious that many wide hybridizations will never be
easy and involve near-impossible constraints such as complete or partial sterility. In
such cases gene cloning and gene discovery, complemented by intransgenic breed-
ing, will hopefully pave the way for success. The utilization of wild relatives through
traditional and molecular breeding has been thoroughly enumerated over the chapters
throughout this series.
Enormous genomic resources have been developed in the model crop relatives, for
example Arabidopsis thaliana and Medicago truncatula. BAC, cDNA and EST
libraries have also been developed in some other crop relatives. Transcriptomes
and metabolomes have also been dissected in some of them. However, similar
genomic resources are yet to be constructed in many crop relatives. Hence this
section has been included only in chapters on the relevant genera.
In this book series, we have included a section on recommendations for future
steps to create awareness about the wealth of wild crop relatives in society at large
and also for concerns for their alarmingly rapid decrease due to genetic erosion. The
authors of the chapters have also emphasized on the imperative requirement of their
conservation, envisaging the importance of biodiversity. The importance of intellec-
tual property rights and also farmers’ rights as owners of local landraces, botanical
varieties, wild species and subspecies has also been dealt in many of the chapters.
I feel satisfied that the authors of the chapters in this series have deliberated on all
the crucial aspects relevant to a particular genus in their chapters.
I am also very pleased to present many chapters in this series authored by a
large number of globally reputed leading scientists, many of whom have contributed
to the development of novel concepts, strategies and tools of genetics, genomics
and breeding and/or pioneered the elucidation and improvement of particular plant
xii Preface

genomes using both traditional and molecular tools. Many of them have already
retired or will be retiring soon, leaving behind their legacies and philosophies for us
to follow and practice. I am saddened that a few of them have passed away during
preparation of the manuscripts for this series. At the same time, I feel blessed that all
of these stalwarts shared equally with me the wealth of crop relatives and contributed
to their recognition and promotion through this endeavor.
I would also like to be candid with regard to my own limitations. Initially I
planned for about 150 chapters devoted to the essential genera of wild crop relatives.
However, I had to exclude some of them either due to insignificant progress made on
them during the preparation of this series, my failure to identify interested authors
willing to produce acceptable manuscripts in time or authors’ backing out in the last
minute, leaving no time to find replacements. I console myself for this lapse with the
rationale that it is simply too large a series to achieve complete satisfaction on the
contents. Still I was able to arrange about 125 chapters in the ten volumes, con-
tributed by nearly 400 authors from over 40 countries of the world. I extend my
heartfelt thanks to all these scientists, who have cooperated with me since the
inception of this series not only with their contributions, but also in some cases by
suggesting suitable authors for chapters on other genera. As happens with a mega-
series, a few authors had delays for personal or professional reasons, and in a few
cases, for no reason at all. This caused delays in the publication of some of the
volumes and forced the remaining authors to update their manuscripts and wait too
long to see their manuscripts in published form. I do shoulder all the responsibilities
for this myself and tender my sincere apologies.
Another unique feature of this series is that the authors of chapters dedicated to
some genera have dedicated their chapters to scientists who pioneered the explora-
tion, description and utilization of the wild species of those genera. We have duly
honored their sincere decision with equal respect for the scientists they rightly
reminded us to commemorate.
Editing this series was, to be honest, very taxing and painstaking, as my own
expertise is limited to a few cereal, oilseed, pulse, vegetable, and fruit crops, and
some medicinal and aromatic plants. I spent innumerable nights studying to attain the
minimum eligibility to edit the manuscripts authored by experts with even life-time
contributions on the concerned genera or species. However, this indirectly awakened
the “student-for-life” within me and enriched my arsenal with so many new concepts,
strategies, tools, techniques and even new terminologies! Above all, this helped me
to realize that individually we know almost nothing about the plants on this planet!
And this realization strikingly reminded me of the affectionate and sincere advice of
Dr. Norman Borlaug to keep abreast with what is happening in the crop sciences,
which he used to do himself even when he had been advised to strictly limit himself
to bed rest. He was always enthusiastic about this series and inspired me to take up
this huge task. This is one of the personal and professional reasons I dedicated this
book series to him with a hope that the present and future generations of plant
scientists will share the similar feelings of love and respect for all plants around us
for the sake of meeting our never-ending needs for food, shelter, clothing, medicines,
and all other items used for our basic requirements and comfort. I am also grateful to
his granddaughter, Julie Borlaug, for kindly extending her permission to dedicate this
series to him.
I started editing books with the 7-volume series on Genome Mapping
and Molecular Breeding in Plants with Springer way back in 2005, and I have since
Preface xiii

edited many other book series with Springer. I always feel proud and satisfied to be a
member of the Springer family, particularly because of my warm and enriching
working relationship with Dr. Sabine Schwarz and Dr. Jutta Lindenborn, with whom
I have been working all along. My special thanks go out to them for publishing this
“dream series” in an elegant form and also for appreciating my difficulties and
accommodating many of my last-minute changes and updates.
I would be remiss in my duties if I failed to mention the contributions of Phullara –
my wife, friend, philosopher and guide – who has always shared with me a love of the
collection, conservation, evaluation, and utilization of wild crop relatives and has
enormously supported me in the translation of these priorities in my own research
endeavors – for her assistance in formulating the contents of this series, for monitor-
ing its progress and above all for taking care of all the domestic and personal
responsibilities I am supposed to shoulder. I feel myself alien to the digital world
that is the sine qua non today for maintaining constant communication and ensuring
the preparation of manuscripts in a desirable format. Our son Sourav and daughter
Devleena made my life easier by balancing out my limitations and also by willingly
sacrificing the spare amount of time I ought to spend with them. Editing of this series
would not be possible without their unwavering support.
I take the responsibility for any lapses in content, format and approach of the
series and individual volumes and also for any other errors, either scientific or
linguistic, and will look forward to receiving readers’ corrections or suggestions
for improvement.
As I mentioned earlier this series consists of ten volumes. These volumes are
dedicated to wild relatives of Cereals, Millets and Grasses, Oilseeds, Legume Crops
and Forages, Vegetables, Temperate Fruits, Tropical and Subtropical Fruits, Indus-
trial Crops, Plantation and Ornamental Crops, and Forest Trees.
This volume “Wild Crop Relatives – Genomic and Breeding Resources: Millets
and Grasses” includes 16 chapters dedicated to Agrostis, Bromus, Cenchrus, Cyno-
don, Dactylis, Dichanthium, Eleusine, Eragrostis, Festuca, Lolium, Panicum, Pas-
palum, Pennisetum, Phleum, Setaria and Zoysia. The chapters of this volume were
authored by 48 scientists from 11 countries of the world namely Algerie, Argentina,
Australia, France, India, Japan, New Zealand, Poland, Portugal, Turkey, and the USA.
It is my sincere hope that this volume and the series as a whole will serve the
requirements of students, scientists and industries involved in studies, teaching,
research and the extension of millets and grasses with an intention of serving science
and society.

Clemson, USA Chittaranjan Kole

.
Contents

1 Agrostis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
B.S. Ozdemir and H. Budak
2 Bromus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
W.M. Williams, A.V. Stewart, and M.L. Williamson
3 Cenchrus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31
S. Goel, H.D. Singh, and S.N. Raina
4 Cynodon . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Yanqi Wu
5 Dactylis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Alan V. Stewart and Nicholas W. Ellison
6 Dichanthium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Vishnu Bhat, C. Mahalakshmi, Shashi, Sunil Saran,
and Soom Nath Raina
7 Eleusine . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113
Susana S. Neves
8 Eragrostis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
Mahmoud Zeid, Vivana Echenique, Marina Dı́az, Silvina Pessino,
and Mark E. Sorrells
9 Festuca . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153
Toshihiko Yamada
10 Lolium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 165
Hongwei Cai, Alan Stewart, Maiko Inoue, Nana Yuyama,
and Mariko Hirata
11 Panicum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 175
Hem S. Bhandari, Masumi Ebina, Malay C. Saha, Joseph H. Bouton,
Sairam V. Rudrabhatla, and Stephen L. Goldman
12 Paspalum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
W.M. Williams, M.L. Williamson, and D. Real

xv
xvi Contents

13 Pennisetum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 217
Thierry Robert, Nadra Khalfallah, Evelyne Martel, Françoise Lamy,
Valerie Poncet, Clémentine Allinne, Marie-Stanislas Remigereau,
Samah Rekima, Magalie Leveugle, Ghayas Lakis, Sonja Siljak-Yakovlev,
and Aboubakry Sarr
14 Phleum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 257
Alan V. Stewart, Andrzej J. Joachimiak, and Nicholas W. Ellison
15 Setaria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 275
Henri Darmency and Jack Dekker
16 Zoysia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 297
Shin-ich Tsuruta, Makoto Kobayashi, and Masumi Ebina

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 311
Abbreviations

5S rDNA gene coding for the 5S rRNA

A/L Accumulation/loss mechanism
ABA Abscisic acid
ACCase Acetyl coenzyme A carboxylase
AFLP Amplified fragment length polymorphism
ALS Acetolactate synthase
ANTC Anthocyanin
APO Aposporous locus
ASGR Apospory-specific genomic region
ASGR Apomixis-related gene region
AusPGRIS Australian Plant Genetic Resource Information Service
BAC Bacterial artificial chromosome
BADH Betaine-aldehyde dehydrogenase
BC Backcross
BP Before present
BSA Bulked segregant analysis
CAD Cinnamyl alcohol dehydrogenase
CCoAOMT Caffeoyl-CoA O-methyltransferase
CCR Cinnamoyl CoA reductase
cDNA Complementary DNA
CENARGEN Centro Nacional de Pesquisa em Recursos Genéticos
e Biotecnologia
CGIAR Consultative Group on International Agricultural Research
CHCH Coiled-coil-helix–coiled-coil-helix
CIAT International Centre for Tropical Agriculture
CID Carbon isotope discrimination
CISP Conserved intron scanning primers
cM Centi-Morgan
CMS Cytoplasmic male sterility
COMT Caffeic acid O-methyltransferase
COMT Caffeic acid/5-hydroxyferulic acid O-methyltransferase
cpDNA Chloroplast-DNA
CPPSE Centro de Pesquisa de Pecuária do Sudeste
CP-sHSP Chloroplast-localized small HSP
Crs-1 Creeping bentgrass specific-1 gene
CTAB Cetyl trimethyl ammonium bromide
cTBP Combinatorial tubulin-based polymorphism

xvii
xviii Abbreviations

cv. Cultivar
CWR Crop wild relative
DAF DNA amplification fingerprinting
DAPI 40 ,6-Diamidino-2-phenylindole
DDBJ DNA Data Bank of Japan
DEF Diferentially expressed fragment
DM Dry matter
EMBL European Molecular Biology Laboratory
EMBRAPA Empresa Brasileira de Pesquisa Agropecuária
EST Expressed sequence tag
FAA Formalin: acetic acid: ethyl alcohol
FAME Fatty acid methyl ester
FISH Fluorescence in situ hybridization
GAI Gene of agronomic interest
GBSSI Granule-bound starch synthase I
GC Gas chromatography
GFP Green fluorescent protein
GI Genes index
GIS Genomic in situ hybridization
GMM Genotype matrix mapping
GR Glyphosate resistant
GRIN Germplasm Resource Information Network (USDA-ARS)
GSS Genome survey sequence
hph Hygromycin phosphotransferase (gene)
HSP Heat shock protein
IBC Institute of Biodiversity Conservation
IBONE Instituto de Botánica del Nordeste
ICRISAT International Crops Research Institute for the Semi-Arid Tropics
IGER Institute of Grassland and Environmental Research
IGFRI Indian Grassland and Fodder Research Institute
ILRI International Livestock Research Institute
IMI Imidazolinone
INDEL Insertion/deletion
INTA National Institute of Agriculture of Argentina
ISSR Inter-simple sequence repeat
ITS Internal transcribed spacer
IVDMD In vitro dry matter digestibility
LEA Late embryogenesis abundant
LG Linkage group
LLR Leucine-rich repeat
MAS Marker-assisted selection
MDA Malondialdehyde
mRNA Messenger-RNA
NADP Nicotinamide adenine dinucleotide phosphate
NADP-ME NADP-malic enzyme
NBPGR National Bureau of Plant Genetic Resources
NBS Nucleotide binding site
NCBI National Center for Biotechnology Information
NDF Neutral detergent fiber
Abbreviations xix

ndhF NADH dehydrogenase F

nirK, nirS Nitrite reductase genes
NMDH NADP-dependent malate dehydrogenase
NOR Nucleolar organizer region
nosZ Nitrous oxide reductase gene
NPGS National Plant Germplasm System
NPK Nitrogen phosphorus potassium
OMT O-Methyl transferase
PAC Plasmid artificial chromosome
PACC Panicoid-arundinoid-chloridoid-centothecoid
PAL Phenylalanine ammonialyase
PC Principal component
PCA Principal component analysis
PCR Polymerase chain reaction
PD Power of discrimination
PGR Plant growth regulator
PHYB Phytochrome B
PIC Polymorphic information content
PSII Photosystem II
Q-PCR Quantitative real-time PCR
QTA Quantitative trait allele
QTL Quantitative trait loci
QTN Quantitative trait nucleotide
R gene Resistance gene
RAPD Random(ly) amplified polymorphic DNA
rDNA Ribosomal DNA
RFLP Restriction fragment length polymorphism
RGL R-gene-like sequence
RIL Recombinant Inbred Line
RNAi RNA-interference
RT-PCR Real-time quantitative PCR
RWC Relative water content
s.str. sensu stricto
SC Secondary constriction
SCAR Sequence-characterized amplified region
SDRF Single dose restriction fragment
SINGER System-wide Information Network for Genetic Resources
SNP Single nucleotide polymorphism
SRAP Sequence-related amplified polymorphism
SSR Simple sequence repeat
STS Sequence tagged site
TBP Tubulin-based polymorphism
TDF Transcript derived fragment
TNL TIR-NBS-LRR
tRNA Transfer-RNA
uidA b-Glucuronidase (gene)
UNED United Nations Conference on Environment and Development
UPGMA Unweighted pair group method with arithmetic mean
xx Abbreviations

USDA United States Department of Agriculture

USDA-APHIS USDA-Animal and Plant Health Inspection Service
USDA-SCS USDA Soil Conservation Service
USGA United States Golf Association
WUE Water use efficiency
List of Contributors

Clémentine Allinne Laboratoire Ecologie Systématique et Evolution UMR (8079)

CNRS, Agroparitech, Université Paris XI, B 360, 91405 Orsay, France, Clementine.
[email protected]
Hem Bhandari Forage Improvement Division, The Samuel Roberts Noble
Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK 73401, USA,
[email protected]
Vishnu Bhat Department of Botany, University of Delhi, Delhi 110007, India,
[email protected]
Joseph Bouton Forage Improvement Division, The Samuel Roberts Noble
Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK 73401, USA,
[email protected]
H. Budak Engineering and Natural Sciences, Biological Sciences and
Bioengineering Program, Sabanci University, Orhanli, 34956 Tuzla, Istanbul,
Turkey, [email protected]
Hongwei Cai Forage Crop Research Institute, Japan Grassland Agriculture and
Forage Seed Association, 388-5, Higashiakata, Nasushiobara, Tochigi 329-2742,
Japan, [email protected]
Henri Darmency INRA, Weed Biology and Management, 17 rue Sully, BP 86510,
21065 Dijon, France, [email protected]
Jack Dekker Weed Biology Laboratory, Agronomy Department, Iowa State
University, Ames, IA 50011, USA, [email protected]
Marina Dı́az Departamento de Biologı́a, Bioquı́mica y Farmacia, Universidad
Nacional del Sur, San Juan 670, 8000 Bahia Blanca, Argentina, [email protected]
Masumi Ebina Forage Plant Breeding and Biotechnology, The National Institute
of Livestock and Grassland Science, 768 Sembonmatsu, Nasushiobara, Tochigi
329-2793, Japan, [email protected]
Viviana Echenique Departamento de Agronomı́a, Universidad Nacional del Sur,
CERZOS-CONICET, San Andrés 800, 8000 Bahia Blanca, Argentina,
[email protected]
Nicholas W. Ellison AgResearch, Private Bag 11008, Palmerston North, New
Zealand, [email protected]

xxi
xxii List of Contributors

S. Goel Department of Botany, University of Delhi, Delhi 110007, India,

[email protected]
Stephen L. Goldman Department of Environmental Science, The University of
Toledo, 2801 W. Bancroft, Toledo, OH 43606, USA, [email protected]
Mariko Hirata Forage Crop Research Institute, Japan Grassland Agriculture and
Forage Seed Association, 388-5, Higashiakata, Nasushiobara, Tochigi 329-2742,
Japan, [email protected]
Maiko Inoue Department of Plant, Soil, and Insect Science, University of
Massachusetts, 8 French Hall, 230 Stockbridge Road, Amherst, MA 01003, USA,
[email protected]
Andrzej J. Joachimiak Department of Plant Cytology and Embryology, Institute
of Botany, Jagiellonian University, Grodzka 52, 31-044 Cracow, Poland,
[email protected]
Nadra Khalfallah Laboratoire de Génétique, Biochimie et Biotechnologies
végétales, Université Mentouri Constantine, Constantine, Algérie, khalfallah@
wissal.dz
Makoto Kobayashi National Institute of Livestock and Grassland Science, Forage
Plant Breeding and Biotechnology, 768 Sembonmatsu, Nasushiobara, Tochigi
329-2793, Japan, [email protected]
Ghayas Lakis Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
Françoise Lamy Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
Magalie Leveugle Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
C. Mahalakshmi Department of Botany, University of Delhi, Delhi 110007, India,
[email protected]
Evelyne Martel Université Lyon 1, UMR CNRS 5023 ‘Ecology of Fluvial
Hydrosystems’, 43 Bd du 11 novembre, F-69622 Villeurbanne Cedex, France,
[email protected]
Susana S. Neves Plant Cell Biotechnology Laboratory,ITQB, Instituto de
Tecnologia Quı́mica e Biológica, Universidade Nova de Lisboa, Apartado 127,
2781-901 Oeiras, Portugal, [email protected]
B.S. Ozdemir Engineering and Natural Sciences, Biological Sciences and
Bioengineering Program, Sabanci University, Orhanli, 34956 Tuzla, Istanbul,
Turkey, [email protected]
List of Contributors xxiii

Silvina Pessino Facultad de Ciencias Agrarias, Universidad Nacional de

RosarioArgentina, Parque Villarino, S2125ZAA Zavalla, Santa Fe, Argentina,
[email protected]
Soom Nath Raina Amity Institute of Biotechnology, Amity University, Noida
201303, Uttar Pradesh, India, [email protected]
Daniel Real Department of Agriculture and Food, Western Australia (DAFWA),
South Perth, WA 6151, Australia, [email protected]
Samah Rekima Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
Marie-Stanislas Remigereau Laboratoire Ecologie Systématique et Evolution
UMR (8079) CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay,
France, [email protected]
Thierry Robert Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
Malay Saha Forage Improvement Division, The Samuel Roberts Noble
Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK 73401, USA,
[email protected]
R.V. Sairam Pennsylvania State University-Harrisburg, The School of Science
Engineering and Technology, Middletown, PA 17057, USA, [email protected]
Sunil Saran Amity Institute of Biotechnology, Amity University, Noida 201303,
Uttar Pradesh, India, [email protected]
Aboubakry Sarr Laboratoire Ecologie Systématique et Evolution UMR (8079)
CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
Shashi Department of Botany, University of Delhi, Delhi 110007, India,
[email protected]
Sonja Siljak-Yakovlev Laboratoire Ecologie Systématique et Evolution UMR
(8079) CNRS, Agroparitech, Université Paris XI, B 360, 91 405 Orsay, France,
[email protected]
H.D. Singh Department of Botany, University of Delhi, Delhi 110007, India,
[email protected]
Mark E. Sorrells Department of Plant Breeding and Genetics, Cornell University,
240 Emerson Hall, Ithaca, NY 14853-1902, USA, [email protected]
Alan V. Stewart PGG Wrightson Seeds, PO Box 175Lincoln, Christchurch 7640,
New Zealand, [email protected]
Shin-ich Tsuruta University of Miyazaki, Faculty of Agriculture, 1-1 Gakuen
Kibanadai-nishi, Miyazaki 889-2192, Japan, [email protected]
xxiv List of Contributors

Warren M. Williams AgResearch, Grasslands Research Centre, Private Bag

11008, Palmerston North 4442, New Zealand, [email protected]
Michelle L. Williamson AgResearch, Grasslands Research Centre, Private
Bag 11008, Palmerston North 4442, New Zealand, michelle.williamson@
agresearch.co.nz
Yanqi Wu Department of Plant and Soil Sciences, Oklahoma State University, 368
Ag Hall, Stillwater, OK 74078, USA, [email protected]
Toshihiko Yamada Field Science Center for Northern Biosphere, Hokkaido
University, Kita 11, Nishi 10, Kita-ku, Sapporo 060-0811, Japan, yamada@fsc.
hokudai.ac.jp
Nana Yuyama Forage Crop Research Institute, Japan Grassland Agriculture and
Forage Seed Association, 388-5, Higashiakata, Nasushiobara, Tochigi 329-2742,
Japan, [email protected]
Mahmoud Zeid Department of Plant Breeding and Genetics, Cornell University,
418 Bradfield Hall, Ithaca, NY 14853-1902, USA, [email protected]
.
Chapter 1
Agrostis

B.S. Ozdemir and H. Budak

1.1 Introduction temperate regions due to their dense nature, low mow-
ing heights without damage, and green appearance.
They are also used for parks and forage. They are
The grass family (Poaceae) emerged 60 million years
cross-pollinating, self-incompatible, and pollinated
ago (Kellogg 2001) and it is one of the largest families
by wind. Agrostis can exhibit both clonal growth and
that include various species with high economical
seed reproduction.
importance, especially the essential cereal crops that
It is hard to classify Agrostis genus taxonomically.
are obligate in the daily diet. Grasses are used for
Due to the similar morphological characters among
many purposes such as in food production, industry,
this genus, others features are needed to be included
lawns, and sports fields. Turfgrasses are used in sports
for identification of new germplasms. Laser flow cyto-
fields and recreation areas whereas they prevent soil
metry for the determination of ploidy level was found
erosion in natural habitats. In the United States, turf-
to be effective in differentiating between diploid,
grass constitutes the second place in the seed market
tetraploid, and hexaploid forms by evaluating six
(Lee 1996).
Agrostis species; A. canina L. subsp. canina, A. canina
Agrostis spp., bentgrass, contains more than 200
L. subsp. montana (Hartm.) Hartm., A. stolonifera var.
perennial turfgrass species as a genus in the Poaceae
palustris (Huds.) Farw., A. capillaris L., A. castellana
family (Hitchcock 1971), but five species of this
Boiss. & Reut., and Agrostis alba L. (Bonos et al.
genus are mainly used as turfgrass, which are all out-
2002).
crossing, perennial, and cool-season grasses: colonial
(Agrostis capillaris L.), velvet (Agrostis canina L.), creeping
(Agrostis stolonifera L.), redtop (Agrostis gigantea Roth),
and dryland (Agrostis castellana Boiss. and Reut.). 1.2 Three Major Bentgrass Species
Agrostis spp. is taxonomically classified under Grami-
neae (Poaceae) family, Pooideae subfamily, Aveneae 1.2.1 Creeping Bentgrass
tribe, and Agrostis genus (Warnke 2003). (A. stolonifera L.)
Bentgrasses are cool-season grasses that are widely
used on golf courses (tees, fairways, and greens) at
Creeping bentgrass is a cool-season grass species that
is native to Western Europe. It is not only adapted to
cool and humid areas but also preferred to be used in
warmer places at golf courses due to its fine texture.
Creeping bentgrass has high density and mowing
H. Budak (*) height as low as 3 mm; these features make it suitable
Engineering and Natural Sciences, Biological Sciences and
to be used especially in greens though it is also used in
Bioengineering Program, Sabanci University, Orhanli, 34956
Tuzla, Istanbul, Turkey tees and fairways of the golf courses (Warnke 2003). It
e-mail: [email protected] requires high maintenance, so it is not ideal for home

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 1
DOI 10.1007/978-3-642-14255-0_1, # Springer-Verlag Berlin Heidelberg 2011
2 B.S. Ozdemir and H. Budak

lawns. It has low growth habit and aggressive spread- season grass and also named as brown top. While it
ing (Casler 2006). is used in lawns and sports fields such as tennis courts
Creeping bentgrass is commonly referred to as and golf courses, it is also a choice for erosion control
A. stolonifera L., but other synonyms are also used (Hubbard 1984). It is compatible with other species
such as Agrostis palustris Huds. and A. stolonifera (Casler 2006).
L. var. palustris (Huds.). It is a strict allotetraploid Colonial bentgrass is preferred on fairways and tees
with a genome A2A2A3A3 (2n ¼ 4x ¼ 28) (Warnke at golf courses with a mowing height of 1.0–2.5 cm;
2003). Complete chloroplast genome sequence of hence, it is not much suitable for greens. It has lower
A. stolonifera (136,584 bp) was presented and com- maintenance needs than creeping bentgrass; instead it
pared with other grass species. The chloroplast genome has more limitations (Ruemmele 2003). Colonial bent-
contains 53.6% coding regions (44.7% protein coding grass can adapt to different environments with forming
genes and 8.9% RNA genes), and the rest is noncoding specific ecotypes (Busey 2003).
regions (Saski et al. 2007). The colonial bentgrass (A. capillaris L.) with the
Creeping bentgrass requires vernalization, and genome A1A1A2A2 and creeping bentgrass (A. stolonifera
flowering occurs generally in late spring to early sum- L.) with the genome A2A2A3A3 are both allotetraploid
mer (Warnke 2003). Interspecific hybridization of and have 14 chromosome pairs (2n ¼ 4x ¼ 28)
creeping bentgrass is possible with five different (Ruemmele 2003; Warnke 2003). Since they are sexu-
Agrostis species (A. canina L., A. castellana Boiss. & ally compatible, hybrids can be produced.
Reut, A. gigantea Roth, A. capillaris L., and A. vinealis Genomics studies facilitate analysis of many spe-
Schreb). cies for their origin and evolution with the use of
Due to its stoloniferous growth, thick thatch layers many tools it provides. Expressed sequence tag
can be formed, if it is not managed properly. This then (EST) sequence analysis of colonial and creeping
serves as a good environment for pathogens and bentgrass A2 genomes suggested that these genomes
insects. It is susceptible to many diseases, but dollar diverged from a common ancestor at about 2.2 million
spot (Sclerotinia homeocarpa), brown patch (Rhizoc- years ago and supported the previous studies that A2
tonia solani), Typhula blight (Typhula incarnata or genomes were common for both species. Their findings
Typhula ishikariensis), the so-called gray snow mold, also showed that both creeping and colonial bent-
and Fusarium blight (Fusarium roseum and Fusarium grasses were closer to tribe Poeae than tribe Aveneae
tricinctum), also named as pink snow mold, are the that are actually closely related tribes (Rotter et al.
main concerns. 2007).
A study with rhizobacteria, isolated from roots of
bentgrass and bermudagrass located from USGA golf
putting greens, showed that Pseudomonas was the
dominant genus in the roots of bentgrass for denitrifi- 1.2.3 Velvet Bentgrass (A. canina L.)
cation. For both species, 17% of the isolates were
identified by GC-FAME (gas chromatography – fatty Velvet bentgrass is referred as A. canina L., which is
acid methyl ester) and 16S rDNA analyses. Besides, native to Europe. It is diploid (2n ¼ 2x ¼ 14) with
nitrous oxide reductase (nosZ) and nitrite reductase genome designation of A1A1. It is confused widely
genes (nirK and nirS) were recognized (Wang and with A. vinealis (2n ¼ 4x ¼ 28) which is an autotet-
Skipper 2004). raploid of A. canina.
Velvet bentgrass is a cool-season grass with very
fine texture, maintaining a good putting surface at golf
courses, and it has stoloniferous growth habit. It is the
most shade-tolerant among other Agrostis spp. It has
1.2.2 Colonial Bentgrass (A. capillaris L.) good heat, low temperature, and drought tolerance.
Besides, it is more tolerant to acidic soils then all
Colonial bentgrass is both referred as A. capillaris L. other bentgrasses. However, it needs high mainte-
and Agrostis tenuis Sibth. Origin of colonial bent- nance (Brilman 2003) and it has low growth habit
grass is Europe and temperate Asia. It is a cool- (Casler 2006).
1 Agrostis 3

1.3 Three Minor Bentgrass Species 1.4 Marker Systems

1.3.1 Redtop (A. gigantea Roth) The origin of colonial bentgrass is temperate Asia and
Europe. It is well adapted to low temperatures and
Redtop is a cool-season, perennial grass, which is partial shade and mostly used for tees and fairways
referred as A. gigantea Roth or A. alba. Redtop bent- of the golf courses. In order to study genetic variation
grass (A. gigantea Roth) is allopolyploid with the and relation among colonial bentgrass populations,
genome A1A1A2A2A3A3 (2n ¼ 6x ¼ 42), which are random amplified polymorphic DNA (RAPD) marker
the inclusion of the genomes of both creeping and analysis was applied, revealing high degree of genetic
colonial bentgrasses. It can be crossed with creeping diversity. The accessions introduced from Europe to
bentgrass to produce infertile hybrids. USA had genetic resemblance with the European cul-
Redtop is used as forage or turf or for reclamation tivars; however, accessions from Bulgaria and Turkey
purposes for revegetation of disturbed areas. It has low showed the lowest genetic similarity, though these two
maintenance (Brede and Sellmann 2003). It has rapid regions are not geographically distant from each other
establishment (Casler 2006). It is tolerant to high (Rajasekar et al. 2007).
metal concentrations and is the most salt-tolerant Twenty-two different colonial bentgrasses (A. capil-
among other bentgrasses. It can be used in breeding laris L.) from different countries, including few com-
programs for enhanced tolerance. It has the potential mercial cultivars, were analyzed with 128 amplified
to be improved as forage or turf (Brede and Sellmann fragment length polymorphism (AFLP) markers,
2003). resulting in high level of genetic biodiversity.
Diverse gene pool availability was especially inves-
tigated in the accession rather than the commercial
cultivars (Zhao et al. 2006). It is natural that during
1.3.2 Highland Bentgrass (A. castellana domestication events, through breeding, cultivated
Boiss. and Reuter) species loses its genetic diversity and gains unifor-
mity. So that wild relatives or other species play an
important role in plant improvement programs.
Highland bentgrass is a dense turf that has good color Colonial bentgrass is a good source with its diverse
in winter times. It is tetraploid, but its hexaploid types germplasm for the improvement of other Agrostis
can also be found. It is adapted to warmer and less species or turfgrass types.
humid areas than A. capillaris. A. castellana has Creeping bentgrass (A. stolonifera L.), colonial
mowing heights ranging from 1.0 to 1.9 cm and can bentgrass (A. capillaris L.), and velvet bentgrass
be used for turf purposes. But it is better if used as a (A. canina L.) are the extensively used bentgrasses
low maintenance grass or for reclamation purposes (Agrostis spp.) on putting greens, tees, and fairways
(Brede and Sellmann 2003); it is a drought-tolerant of the golf courses at temperate regions. They are
species (Casler 2006). highly difficult to be identified using their morpholog-
ical features. However, it was investigated that RAPD
markers were effective in differentiation and identi-
fication of bentgrass species at molecular level
1.3.3 Idaho Bentgrass (Agrostis (Hollman et al. 2005). Besides the use of RAPD mar-
idahoensis Nash) kers, sequence characterized amplified region (SCAR)
markers were designed for colonial and creeping bent-
Idaho bentgrass is a cool-season, perennial grass spe- grasses. They were found to be effective to differ-
cies, which is native to North America. Depending on entiate between these two bentgrasses and these from
its mowing height, it shows more tolerance to diseases other Agrostis spp. This technology was also recom-
than any other bentgrass types. It can be used for mended to be used for selecting the progenies
improvement of Agrostis germplasm in pest resistance produced from interspecific hybridization between
(Brede and Sellmann 2003). colonial and creeping bentgrasses (Scheef et al. 2003).
4 B.S. Ozdemir and H. Budak

Few creeping bentgrass cultivars were identified growth (John et al. 1995) and, especially for turfgrass,
using restriction fragment length polymorphism loss of deep green color, which is an important param-
(RFLP) markers (Caceres et al. 2000), which is one eter for its economic value. SAG12-ipt (ligation of
of the first marker types to be used in linkage studies ipt, adenine isopentenyl transferase gene to SAG12,
and an important tool for genetic mapping. Moreover, senescence-activated promoter) gene integration into
molecular marker-based linkage map was developed creeping bentgrass resulted in transgenic lines that
for creeping bentgrass using AFLP, RAPD, and had faster growth under normal temperatures and
cDNA-RFLP markers. Further use of this map would increased ipt gene expression and cytokinin pro-
be useful in detection of quantitative trait loci (QTL) duction suppressed heat-induced leaf senescence (Xu
and marker-assisted breeding of agronomically impor- et al. 2009) resulting in an improvement for cool-
tant traits like disease resistance (Chakraborty et al. season grasses exposed to heat stress.
2005). Transcript polymorphism analysis using SRAP The main problem of growing cool-season grasses
system in three different bentgrasses (colonial, creep- in temperate areas is the increasing temperature in
ing, and velvet bentgrass) was the first study to be summer time causing a decline in plant productivity.
reported on turfgrasses. The ESTs obtained from this Using differential display analysis, upregulated genes
study could be used in turfgrass improvement pro- (18 in A. stolonifera and 22 in A. scabra) in response
grams (Dinler and Budak 2008). to heat stress were identified and most genes were
AFLP analysis for tetraploid creeping bentgrass found in both species. A. scabra is a heat-tolerant
and hexaploid redtop bentgrass was performed to species for which the significant decline in soluble
investigate genetic diversity of the old and new culti- protein content was observed at 40 C, whereas
vars, European plant introductions, and gray snow A. stolonifera is nontolerant under heat stress, and
mold (T. incarnata Lasch) resistant genotypes of protein content conservation was up to 30 C (Xu and
creeping bentgrass and could be used in plant Huang 2008a, b). Only three of these genes were
improvements and gene mapping studies (Vergara expressed in A. scabra, and it was reported that these
and Bughrara 2004). For dollar spot disease, QTL genes might have been serving for the heat tolerance.
analysis was performed to deeply understand the Previous analysis of thermal and nonthermal A. scabra
genetics of the resistance to this disease. It was found with 60 RAPD loci showed that they were not clus-
that dollar spot resistance is a quantitative trait and tered independently but they were distantly related
highly heritable. RAPD markers tightly linked to this although they were morphologically similar (Tercek
QTL could be a useful tool in breeding programs of et al. 2003). If it is the case, this might enlighten the
Agrostis spp. (Chakraborty et al. 2006). mechanism of plants’ tolerance to heat stress and
facilitate development of in heat tolerant cultivars
(Xu et al. 2008).
Heat shock proteins (HSPs) in plants are syn-
1.5 Heat Stress and Drought Tolerance thesized in response to heat stress for protection. The
small HSPs that are localized in chloroplasts (CP-sHSPs)
Plants are affected from temperature changes since all are essential in heat tolerance expressing varying levels
metabolic activities take place at a certain tempera- of its isoforms. Two additional isoforms of CP-sHSPs
ture. Heat stress tolerance level differs according to in heat-tolerant variants of creeping bentgrass (Penn-
the plant species. Some plants can become tolerant to cross cultivar), which were generated under heat stress
high levels of heat stress by time, and that can be and selected for heat tolerance, were accumulated with
exhibited as a genetic character. respect to heat-sensitive variants that were not sub-
In cool-season grass species, heat stress causes leaf jected to heat stress. These isoforms were genetically
senescence that is regulated by cytokinins. The linked to heat tolerance (Park et al. 1996; Luthe et al.
decrease in cytokinin content causing an increase in 2000). The study of CP-sHSP encoding genes isolated
leaf senescence process was reported in Agrostis spe- from heat-tolerant and heat-sensitive variants showed
cies (Xu and Huang 2007). Chlorosis followed by leaf that the differences and variation in the expression of
senescence causes a decline in chlorophyll content and CP-sHSPs were related with the environmental adap-
photosynthetic capacity resulting in reduced plant tation and suggested that the amount of CP-sHSPs
1 Agrostis 5

might have been the source of heat tolerance (Wang and the root respiration rate was low or downregulated
et al. 2003; Wang and Luthe 2003). in A. scabra at high temperatures. Both factors were
During the growth of creeping bentgrass species, suggested to be the reason for root thermotolerance
the temperature was increased gradually. Following adaptation of cool-season grasses (Lyons et al. 2007).
heat acclimation, the creeping bentgrass showed an Both short-term and long-term respiratory acclimation
improved tolerance under heat stress. In consecutive was investigated to be accompanied with root thermo-
studies, it was found that heat acclimation was asso- tolerance A. scabra at increasing temperatures. A. sca-
ciated with suppressed lipid peroxidation (Larkindale bra was found to be giving less response, in terms of root
and Huang 2004), enhanced heat shock protein expres- respiration, than A. stolonifera. The adjustment of root
sion (He et al. 2005), and enhanced photosynthetic carbon utilization was stated to be caused by respiratory
activity by higher pigment content and rubisco activity acclimization that the need for increased respiratory
(Liu and Huang 2008). energy was lowered at high temperatures, resulting in
A. scabra is a thermal species found to be tolerant increased root survival (Rachmilevitch et al. 2008).
to high temperatures like 40–45 C and to be living Thermal A. scabra has higher root viability than
at the Yellowstone Natural Park (Tercek et al. 2003). heat-sensitive A. stolonifera. The protein and phos-
For heat tolerance capacity, cool-season, perennial phoprotein patterns of A. scabra and A. stolonifera
grasses, A. scabra with two genotypes and commer- were mainly different under heat stress. It was pro-
cially important A. stolonifera with ten genotypes posed that sucrose synthase, glutathione S-transferase,
were investigated for the expression of AsEXP1 gene superoxide dismutase, stress-inducible heat shock pro-
under heat stress. Heat tolerance of the treated plants tein, and aldolase phosphorylation might have been
differed according to the Agrostis species and geno- associated with root thermotolerance in cool-season
types and fell into three classes as being most tolerant grasses under heat stress (Xu and Huang 2008a, b).
to most sensitive. Heat tolerance level was positively Since carbon utilization and accumulation are
correlated with the level of AsEXP1 gene expression. thought to have a role in heat tolerance and enhance-
Totally, four genotypes, two from A. scabra and two ment of plant survival in perennial grass species, three
from A. stolonifera, exhibited the highest level of gene species of cool-season turfgrasses, colonial bentgrass
expression with response to heat stress, being the most (A. capillaris L.), creeping bentgrass (A. stolonifera L.),
heat-tolerant ecotypes. This expansin gene, AsEXP1, and velvet bentgrass (A. canina L.), were examined
was found to be highly upregulated in shoots and based on their shoot and root carbon partitioning and
affected the heat tolerance of both species as “whole- carbohydrate accumulation. Measuring the turf quality
plant.” The identification of this gene as response to and relative leaf water content revealed that velvet
heat tolerance in C3 Agrostis species for the first time bentgrass was the most tolerant under drought condi-
was suggested to be important both for the study and tions and colonial bentgrass had the most amendatory
investigation of the heat tolerant germplasm of grasses potential. According to total nonstructural carbohy-
(Xu et al. 2007). In a further study, novel heat respon- drate content measurements, increased carbon accu-
sive genes were identified in A. scabra by subtractive mulation in roots after initial drought application and
suppression hybridization approach. The differentially then its gathering in stems and leaves at increased
expressed genes were classified based on their role in drought durations suggested an adaptive response for
stress and defense mechanism, signaling and transcrip- drought survival (DaCosta and Huang 2006a, b, c).
tion, and protein or carbon metabolism (Tian et al. Other study with creeping bentgrass with different
2009) irrigation applications resulted in enhanced carbohy-
For plants, the soil temperature is more important drate levels in leaves and roots when irrigating at wilt
than temperature of the air. The soil temperature primar- (Fu and Dernoeden 2008).
ily affects the roots. Heat-tolerant A. scabra is adapted to Velvet bentgrass was found to be the most drought-
high soil temperatures at geothermal locations, whereas tolerant bentgrass species among others. In colonial,
heat-sensitive A. stolonifera, adapted to cool climatic creeping, and velvet bentgrasses, prolonged drought
regions, cannot survive at high temperatures. When stress caused a decrease in antioxidant enzyme activ-
compared to A. stolonifera, whole-plant carbon balance ities and an increase in lipid peroxidation. However, in
and root carbon utilization was maintained positively, velvet bentgrass, oxidative damage was prevented for
6 B.S. Ozdemir and H. Budak

a longer period of time under drought stress compared adaptation to biotic and physiological stresses, and
to other two bentgrasses that resulted in maintenance planting in cultural management of weed in turfgrass
of higher turf quality and leaf relative water content is reviewed by Busey (2003).
(DaCosta and Huang 2007a, b). Besides, velvet bent- Bentgrasses are grown as monoculture on golf
grass had higher degree of osmotic adjustment, which courses. Weed control is the main problem of golf
is crucial for cell turgor pressure maintenance, and this course maintenance. Poa annua L. is the major weed
osmotic adjustment was accompanied with increased that shows similar tolerance responses to herbicides
proline content and deposition of water-soluble carbo- like creeping bentgrass. There is no effective control
hydrates both in velvet and creeping bentgrasses of this weed, so it causes severe problems. So, genetic
(DaCosta and Huang 2006a, b, c). manipulation of turfgrass species for glyphosate or
Velvet bentgrass exhibited less injury and more glufosinate type resistance is mainly to fight with
ABA (abscisic acid) accumulation under drought con- P. annua in greens and unwanted grasses in other
ditions with respect to colonial and creeping bentgrass. areas of golf courses (Duncan 2004).
On the other hand, creeping bentgrass exhibited higher A. capillaris grows in patches with its dense tillers.
recovery from drought stress and, at the same time, Leaf growth of A. capillaris is postponed in spring
showed rapid decline in ABA and incline in cytokinin time. It is a rhizomatous species. A. capillaris was
content, where ABA and cytokinin are regulators in found to be vegetatively the most competitive and
decreased water availability for plant consumption invasive temperate grass species when compared
(DaCosta and Huang 2007a, b). with Festuca rubra, Holcus lanatus, Lolium perenne,
and Poa trivialis, though their time of vegetative
spreading differed during the growing season
(Barthram et al. 2006). It was proposed that the varied
1.6 Salt Tolerance invasion properties of different grass species and the
physical resistance was not only related with lamina
density but also other lamina characteristics should
Efficient irrigation management results in higher turf
have been involved (Barthram et al. 2005).
quality, water saving, and less cost on golf courses by
evaluating the water requirements of the species that
were found to be depending on species and time of the
year (DaCosta and Huang 2006a, b, c). However,
water availability and quality is decreasing in today’s 1.8 Diseases in Agrostis Species
world and at the same time, the demand for salt-tolerant
cultivars is increasing. Salt-tolerant and sensitive cul-
Fungal diseases are tried to be prevented with extensive
tivars were determined among creeping, colonial, and
fungicide application, which generates both environ-
velvet bentgrass species (McCarty and Dudeck 1993;
mental problems and economical losses. So, improve-
Marcum 2001), and red top bentgrass (McCarty and
ment of turfgrass species for resistance to fungal
Dudeck 1993).
diseases such as dollar spot or brown patch, which
cause serious problems in turfgrass management, has
crucial importance (Chakraborty et al. 2006).
Dead spot disease is a newly occurring disease
1.7 Invasion Properties and Weed
caused by Ophiosphaerella agrostis and effective in
Control Invasion creeping bentgrass. It is a major problem of greens in
the golf courses. The disease occurs within the first
Cultural management of weed in turfgrass is important and sixth years of the established greens, and in older
in the fight of weed invasion since it is a more com- greens, it might be seen after fumigation with methyl
bined system. Healthy plants are less prone to weed bromide. This disease appears first in 1–2 cm diameter
colonization. Environmental stresses, both abiotic and area with reddish brown spots and later the patches
biotic, cause weed colonization. The use of mowing, become 8–10 cm in diameter (Dernoeden et al. 1999;
cultivation, fertilization, irrigation, turfgrass selection, Câmara et al. 2000; Kaminski and Dernoeden 2006).
1 Agrostis 7

Gray snow mold is another fungal disease to which conferring the presence of these types of genes in
the creeping bentgrass is susceptible and caused by grasses (Budak et al. 2006).
T. incarnata (Wu and Hsiang 1998). It is favored by
cold and humid conditions like under snow cover.
Dollar spot (Sclerotinia homoeocarpa F.T. Bennett) 1.9 Plant Transformation in Agrostis
is one of the major diseases of turfgrasses. Fungal
diseases cause serious problem in all grasses. Disease
Species
control is mostly done through fungicide application
(Dai et al. 2003). Treatment trials of creeping bent- In order to achieve improvements in agricultural traits,
grass with fungicides and fungicides plus plant growth plant transformation technologies are important to
regulators (PGRs) resulted in a better quality of creep- use with classical breeding methods. Today’s molecular
ing bentgrass after treatment with fungicides plus biology and plant biotechnology techniques enable us
PGRs rather than fungicides alone (Fidanza et al. to study the structure and function of desired gene,
2006). Differences in day and night temperatures and isolate it from its wild relatives, other cultivars, and
high humidity fortify the dollar spot disease. Though even from other species, and transfer that gene of inter-
the disease resistance in creeping bentgrass has been est to the existing cultivar that is in the process of
genetically identified, its tolerance varies partially improvement. Application and integration of the mod-
with environmental conditions (Bonos et al. 2003; ern science into conventional breeding methods reduce
Bonos 2005). In creeping bentgrass species, it was the time spent for plant improvement studies and also
proposed that the dollar spot disease was most proba- provide us the ability of transferring defined gene or set
bly a quantitatively inherited trait, and parent selection of genes without contamination of undesired genes. The
was crucial for improvements in disease resistance desired gene(s) to be integrated into the existing cultivar
(Bonos 2006). can be obtained from its domestic cultivars, wild rela-
While creeping bentgrass is susceptible to dollar tives, and related genera (or any other organism) by
spot disease and shows different levels of sensitivity expanding the availability of gene pool.
among its cultivars, colonial bentgrass species has resis- In turfgrass species, although microprojectile bom-
tance to this severe disease. Interspecific hybrids bardment and protoplast transformation are the major
between these two species were performed and in gene transfer systems to achieve transgenic plants,
some of the hybrids produced, high resistance to dollar Agrobacterium-mediated transformation is used to
spot disease and even no disease symptoms were get low copy number of the transgene in the trans-
observed (Belanger et al. 2004). During studies with formed cultivars. Agrobacterium-mediated transfor-
resistance to dollar disease with creeping and colonial mation is efficiently used in dicot plants since they
bentgrass species, it was found that a gene loss was occur as a natural host for Agrobacterium tumefaciens.
occurring in the Agrostis genus. The creeping bent- However, by the improvements and modifications of
grass-specific-1 gene, Crs-1, was examined in creeping the bacterial strains in this system, many monocots,
bentgrass plants conferring the loss in most of the especially the important cereal crops, can be trans-
individuals, which occurs rarely, but the function of formed. Turfgrass was also transformed by using sili-
the protein has not been investigated yet (Li et al. 2005). con carbide fibers or whiskers, electroporation, and
Resistance (R) genes are important for genetic polyethylene glycol-mediated techniques. Herbicide
improvement of plant species. There are more than resistance, disease or insect resistance, and stress tol-
five different classes of R-genes. In a study of Budak erance were the main traits to be improved in plant
and co-workers, constitutively expressed R-gene-like transformation studies. The transgenic Agrostis spe-
sequences (RGLs) from different Agrostis species cies with different transformation methods, marker
(creeping, colonial, and velvet bentgrasses) were systems, and transgene types are outlined in Table 1.1.
isolated and characterized by PCR-based motif-directed By using green fluorescent protein (GFP) as a
RNA fingerprinting. It was found that RGLs from these reporter gene, the first study of creeping bentgrass
Agrostis species were highly conserved and some shared (A. stolonifera L.) by Agrobacterium-mediated trans-
conserved motifs with other disease-resistant genes. formation was done by Yu et al (2000). In a latter
Two TNL (TIR-NBS-LRR)-type RGLs were isolated report, higher efficiency with large number of
8 B.S. Ozdemir and H. Budak

Table 1.1 Genetic transformation of bentgrass species with different transgene expression [modified from Wang and Ge (2006)]
Plant species Transgenes Method Outcome References
Agrostis alba (redtop) npt2 (neomycin phosphotransferase II) Protoplasts Transgenic plants Asano and Ugaki
(1994)
Agrostis stolonifera gusA (b-glucuronidase) Biolistics Transgenic plants Zhong et al. (1993)
(creeping bar (phosphinothricin acetyltransferase) Biolistics Transgenic plants Hartman et al.
bentgrass) (1994)
bar (phosphinothricin acetyltransferase) Protoplasts Transgenic plants Lee et al. (1996)
hph (hygromycin phosphotransferase), Biolistics Transgenic plants Xiao and Ha (1997)
gusA (b-glucuronidase)
bar (phosphinothricin acetyltransferase) Protoplasts Transgenic plants Asano et al. (1998)
hph (hygromycin phosphotransferase), Whiskers Transgenic plants Dalton et al. (1998)
gusA (b-glucuronidase)
gfp (green fluorescent protein) Agrobacterium Transgenic plants Yu et al. (2000)
hs2 (chitinase-like protein from American Biolistics Transgenic plants Chai et al. (2002)
elm), bar (phosphinothricin
acetyltransferase)
PAPII, PAP-Y (pokeweed antiviral proteins), Biolistics Transgenic plantsa Dai et al. (2003)
hph (hygromycin phosphotransferase)
PR5K (receptor protein kinase gene from Biolistics Transgenic plantsa Guo et al. (2003)
Arabidopsis thaliana)
RCH10 (chitinase gene from rice), ALG Biolistics Transgenic plantsa Wang et al. (2003)
(glucanase gene from alfalfa), bar
(phosphinothricin acetyltransferase)
bar (phosphinothricin acetyltransferase) Agrobacterium Transgenic plants Luo et al. (2004)
TLPD34 (thaumatin-like protein from rice), Agrobacterium Transgenic plantsa Fu et al. (2005a, b)
bar(phosphinothricin acetyltransferase)
hph (hygromycin phosphotransferase), Agrobacterium Transgenic plants Han et al. (2005)
gusA (b-glucuronidase)
bar (phosphinothricin acetyltransferase), Agrobacterium Transgenic plants Wang and Ge (2005)
gusA (b-glucuronidase)
Agrostis tenuis hph (hygromycin phosphotransferase), Agrobacterium Transgenic plants Chai et al. (2004)
(colonial bentgrass) gusA (b-glucuronidase)
a
Transgenic plants transformed with agronomically important traits

transgenic herbicide (bar gene integration)-resistant cultures or directly the embryos are used as explant
creeping bentgrass plants was achieved with low copy sources. But either way, calli formation is required
number of transgene integration by using super binary before or after the transformation process, which is
vector system (Luo et al. 2004). Transgenic herbicide- time-consuming. Mature embryo-derived embryo-
resistant creeping bentgrass plants were produced genic calli was used to transfer uidA (b-glucuronidase)
recently using Agrobacterium-mediated transformation and hph (hygromycin phosphotransferase) gene to
(Kim et al. 2007). Transgenic turfgrass was first pro- creeping bentgrass with Agrobacterium-mediated
duced via bombardment of A. palustris Huds. embryo- transformation (Han et al. 2005). For creeping
genic callus tissues (Zhong et al. 1993). On the other bentgrass species, stolon nodes were used with Agro-
hand, A. alba L. (redtop) plants were regenerated from bacterium-mediated transformation. This procedure
protoplasts that were achieved from suspension culture maintained shoot formation directly from infected
of embryogenic calli (Asano and Sugiura 1990). nodes by-passing the callus formation step (Wang
The importance of tissue culture parameters in the and Ge 2005). Also, different tissues of creeping bent-
efficiency of plant transformation system is well grass cultivar Penn A4 were compared for their nucle-
known. In order to obtain higher number of transgenic ase activity and its effect on transient GUS expression
lines, first step is to establish an efficient tissue culture efficiency after bombardment (Basu et al. 2003).
system. In genetic transformation studies, especially in Using the whole vector construct during transforma-
monocots, immature or mature embryo-derived callus tion results in integration of vector backbone sequences
1 Agrostis 9

into the genome. Using a simple cassette rather than the stress-inducible promoters was done, and transgenic
whole plasmid in transformation of creeping bentgrass plants with improved tolerance to drought were developed
by the bombardment of tissues with GFP reporter gene (Fu et al. 2007).
increased the low copy number of transgene integration Creeping bentgrass differs in color from a range of
since the system eliminates the nonessential insertions olive green to pale green. For ornamental purposes,
(Jayaraj et al. 2008) by time eliminating the disadvan- creeping bentgrass was transformed with maize flavo-
tage of biolistic transformation. noid/anthocyanin biosynthetic pathway transcription
Plant biotechnology is also widely used in improve- factor genes, Lc (leaf color) and Pl (purple leaf) via
ment of disease resistance. Creeping bentgrass is sen- Agrobacterium-mediated gene transfer technique to
sitive to many insects, weeds, disease-causing fungi, obtain purple colored creeping bentgrass and the result
and bacteria; hence, this species is used in many was three different phenotypes (Han et al. 2009).
genetic improvement studies. Biolistic transformation The RTS gene expressed in tapetal cells was
of creeping bentgrass expressing three forms of poke- isolated from rice panicles. It was shown that this
weed antiviral proteins (ribosome-inactivating pro- gene had a role in pollen development and its promoter
teins) was established via biolistics (Dai et al. 2003). conferred cell-specific expression. Transformation of
Creeping bentgrass cultivar was transformed with the rice, Arabidopsis, and creeping bentgrass with RTS
CP4 EPSPS gene that confers resistance to glyphosate promoter and cytotoxic barnase gene induced male
herbicide. This Roundup Ready® glyphosate-resistant sterility showing the promoter’s anther-specific
creeping bentgrass is under USDA-APHIS regulated expression, its role in pollen development, and use in
status and for production control studies. It was planted male sterility both in dicot and monocot plants (Luo
for a four-year trial. Since creeping bentgrass is an out- et al. 2006). The use of FLP/FRT site-specific DNA
crossing species with wind dispersal mechanism, at the recombination system was evaluated using transgenic
end of the fourth year its dispersal area was heavily creeping bentgrass for yeast FLP recombinase expres-
widened. Though in situ hybrids were not found, the sion and site-specific recombination was suggested to
potential transgene flow from creeping bentgrass and the be a useful system for genome modification and trans-
possibility of its dispersion via water ways due to its gene manipulation in turfgrass by altering the gene
vegetative growth by stolon formation was expressed. escape due to outcrossing and vegetative growth (Hu
The importance of wind for pollen dispersal was empha- et al. 2006).
sized and reported. Moreover, the study of Watrud et al. Agrobacterium-mediated transformation was done
(2004) emphasized the importance of pollen dispersal with mature embryo-derived calli of A. stolonifera.
by wind and demonstrated the long distance of viable PMI/mannose selection and GFP screenable marker
pollen movement and gene flow using CP4 EPSPS as system was used in this study by transferring GFP
a marker. Also, Reichman et al. (2006) reported the gene and E. coli manA (also termed as pmi and codes
identification of glyphosate-resistant creeping bentgrass for phosphomannose isomerase) gene. This system
plants outside the control area and the transgene escape was suggested to be environment and ecosystem
from cultivated area to the native population area. Not friendly when compared with the use of herbicide or
for all transgenic crop species but especially the out- antibiotic selection systems (Fu et al. 2005a, b).
crossing ones were suggested to be evaluated for risk
assessment more strictly (Zapiola et al. 2008). Factors
affecting gene flow and the status of glyphosate-resistant
1.10 Hybridization Studies and Gene
(GR) crops including the GR creeping bentgrass that
are commercially available or under deregulation in Escape
the USA were all reviewed by Mallory-Smith and
Zapiola (2008). Interspecific and intergeneric hybridization between
Overexpression of late embryogenesis abundant glyphosate-resistant transgenic creeping bentgrass
(LEA) proteins was shown to cause improved toler- and nontransgenic species from both Agrostis and
ance to water deficit in various species. Transforma- Polypogon genus was performed. From A. capillaris
tion of creeping bentgrass with barley hva1 gene, and P. fugax fertile hybrids were achieved, which
member of LEA protein family, using constitutive or makes the transgene movement possible within and
10 B.S. Ozdemir and H. Budak

between species. This increased the risk of contamina- Asano Y, Ito Y, Fukami M, Sugiura K, Fujiie A (1998)
tion of the weeds with herbicide resistance like genes Herbicide-resistant transgenic creeping bentgrass plants
obtained by electroporation using an altered buffer. Plant
and the possibility of new weed occurrence (Zhao Cell Rep 17:963–967
et al. 2007). Barthram GT, Elston DA, Mullins CE (2005) The physical
Interspecific hybridization between transgenic resistance of grass patches to invasion. Plant Ecol 176:79–85
creeping bentgrass (A. stolonifera L.) that conferred Barthram GT, Elston DA, Griffiths JH, Bolton GR, Wright G
(2006) Within-year variation in the vegetative spread of five
resistance to glufosinate herbicide and four other non- temperate grasses. J Veg Sci 17:315–322
transgenic Agrostis species (A. castellana Boiss. and Basu C, Luo H, Kusch A, Chandlee J (2003) Transient reporter
Reut., A. gigantea Roth, A. canina L., and A. capillaris L.) gene (S) in creeping bentgrass (Agrostis palustris) is affected
were examined under field conditions. Interspecific by in vivo nuclease activity. Biotechnol Lett 25:939–944
Belanger FC, Meagher TR, Day PR, Plumley K, Meyer WA
transgenic hybrid frequency was at a lower rate with (2003) Interspecific hybridization between Agrostis stoloni-
respect to intraspecific hybridization (0.631%). While fera (creeping bentgrass) and related Agrostis species under
the interspecific transgenic hybrids were formed between field conditions. Crop Sci 43:240–246
creeping bentgrass and A. capillaris (0.044%) and Belanger FC, Bonos S, Meyer WA (2004) Dollar spot resistant
hybrids between creeping bentgrass and colonial bentgrass.
A. castellana (0.0015%), no transgenic hybrids with Crop Sci 44(2):581–586
A. gigantea or A. canina were observed. The flow- Bonos SA (2005) Creeping bentgrass cultivars with improved
ering time difference between the Agrostis species dollar spot resistance. Golf Course Manag 73:96–100
and the only substantial overlap of flowering between Bonos SA (2006) Heritability of dollar spot resistance in creep-
ing bentgrass. Phytopathology 96(8):808–812
A. stolonifera and A. capillaris was the main concern Bonos SA, Plumley KA, Meyer WA (2002) Ploidy determina-
of these hybridization rates (Belanger et al. 2003). tion in Agrostis using flow cytometry and morphological
Using the data from the previous study, transgene traits. Crop Sci 42(1):192–196
pollen dispersal model was performed. It was shown Bonos SA, Casler MD, Meyer WA (2003) Inheritance of dollar
spot resistance in creeping bentgrass. Crop Sci 43:2189–2196
that pollen dispersal varied from site to site with the Brede DA, Sellmann MJ (2003) Three minor Agrostis species:
environmental conditions. It was stated that there was Redtop, Highland Bentgrass, and Idaho Bentgrass. In: Casler M,
risk of transgene escape, but the persistence of this Duncan RR (eds) Turfgrass biology, genetics and breeding.
trait was possible only with herbicide selection, and Wiley, Hoboken, NJ, USA, pp 207–223
Brilman LA (2003) Velvet bentgrass (Agrostis canina L.). In:
the herbicide application was also causing natural Casler M, Duncan RR (eds) Turfgrass biology, genetics and
resistance in weed populations. They suggested that breeding. Wiley, Hoboken, NJ, pp 201–205
the results of such gene flow could cause minimum Budak H, Su S, Ergen N (2006) Revealing constitutively expressed
problem at ecological aspect, and mismanagement of resistance genes in Agrostis species using PCR-based motif-
directed RNA fingerprinting. Genet Res 88:165–175
herbicide application could cause more serious pro- Busey P (2003) Cultural management of weeds in turfgrass: a
blems (Meagher et al. 2003). review. Crop Sci 43:1899–1911
Herbicide-resistant transgenic A. stolonifera was Caceres ME, Pupilli F, Piano E, Arcioni S (2000) RFLP markers
produced by engineering the CP4 EPSPS gene. It are an effective tool for the identification of creeping bent-
grass (Agrostis stolonifera L.) cultivars. Genet Resour Crop
would have been the first transgenic wind-pollinated Evol 47:455–459
perennial crop grown in open areas, but it was reported Câmara MPS, O’Neill NR, van Berkum P, Dernoeden PH, Palm
that pollen-mediated transgene flow was causing a ME (2000) Ophiosphaerella agrostis sp. nov. and its rela-
serious risk. Cytoplasmic male sterility or plastid engi- tionship to other species of Ophiosphaerella. Mycologia
92:317–325
neering might be the solution in these cases to prevent Casler MD (2006) Centenary review: perennial grasses for turf,
transgene escape through pollen dispersal. sport and amenity uses: evolution of form, function and
fitness for human benefit. J Agric Sci 144:189–203
Chai B, Maqbool SB, Hajela RK, Green D, Vargas JM Jr,
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.
Chapter 2
Bromus

W.M. Williams, A.V. Stewart, and M.L. Williamson

2.1 Introduction (Krecz & Vved.) Tournay. (one species). Section Bois-
siera (Hochst. ex Steudel) P. M. Smith was included in
section Bromus by Smith (1970) and separated as a
Bromus L. is a genus of approximately 150 C3 grass
separate section by Smith (1985). The major agricul-
species (Clayton and Renvoize 1986; Watson and
tural species come from sections Pnigma (B. inermis
Dallwitz 1992) that can be considered to be intermedi-
Leyss.) and Ceratochloa (B. catharticus Vahl,
ate between the Festuceae and the Triticeae. The genus
B. sitchensis Trin. in Bong). Polyploidy and hybridi-
is distributed widely in Asia, Europe, Africa, and the
zation feature strongly in the evolution of many spe-
Americas, and today, introductions are widespread
cies (Stebbins 1981). The sections are distinguished
in the temperate world. There is a high incidence of
morphologically according to numbers of nerves in the
polyploidy, species ranging from 2x to 12x. Many of
glumes, spikelet shape, and lemma and awn morphol-
the high polyploids are of allopolyploid (hybrid) ori-
ogy, as well as karyotypes, genome relationships
gin (Stebbins 1981; Armstrong 1991). There is a
(chromosome pairing), ploidal levels, and serological
diversity of annuals and perennials with a range of
differences.
bunchgrass and rhizomatous morphologies.
Because of its complexity, no worldwide taxo-
nomic treatment exists, but many regional descriptions
and identification keys have been published, including
those of Alaska (Mitchell 1967), Mexico and Central
2.2 Evolution and Systematics America (Soderstrom and Beaman 1968), North
America (Wagnon 1952; Allred 1993; Pavlick 1995),
The genus is taxonomically difficult with several South America (Pinto-Escobar 1981, 1986; Matthei
unresolved species complexes, especially in section 1986; Gutierrez and Pensiero 1998; Planchuelo and
Ceratochloa. For the purposes of this chapter, the Peterson 2000), Malesia (Veldkamp et al. 1991), New
classification of Bromus into seven sections (Smith Zealand (Forde and Edgar 1995), South-East Asia
1970, 1985) will be used (Table 2.1). These sections (Chen and Kuoh 2000), and Europe and North Africa
are: Bromus (30–40 species), Genea Dumort. (seven (Spalton 2002, 2004).
species), Pnigma Dumort. (about 60 species), Cerato- Stebbins (1981) hypothesized that Bromus proba-
chloa (P. Beauv.) Griseb. (10–16 species), Neobromus bly arose in Eurasia, when the Festuceae and Triti-
(Shear.) Hitchcock (two species), and Nevskiella ceae were separating. The original Bromus species
are extinct and were probably wiped out during the
dramatic climatic fluctuations of the Pliocene and
Pleistocene. During the Pliocene, differentiation of
sections Neobromus, Ceratochloa and Pnigma
W.M. Williams (*)
AgResearch, Grasslands Research Centre, Private Bag 11008,
occurred. Sections Neobromus and Ceratochloa
Palmerston North 4442, New Zealand spread to North America and later to South America.
e-mail: [email protected] These sections became extinct in Eurasia, and even

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 15
DOI 10.1007/978-3-642-14255-0_2, # Springer-Verlag Berlin Heidelberg 2011
16 W.M. Williams et al.

Table 2.1 Genus Bromus taxonomic delimitation by different phylogenies require clarification, and much wider sam-
authors [modified from Armstrong (1991)] pling of the species is also needed.
Tzvelev (1976) Smith (1970, 1985) Stebbins (1981)
7 Genera 7 Sections 7 Subgenera
Anisantha Genea Stenobromus
Bromus Bromus Bromus 2.3 Agricultural Status
Bromopsis Pnigma Festucaria
Ceratochloa Ceratochloa Ceratochloa
– Neobromus Neobromus About ten species are used in agriculture and many
Boissiera Boissiera Boissiera more have weed status. Two sections contribute
Nevskiella Nevskiella Nevskiella important agricultural species:
Littledalea Section Pnigma has the winterhardy Bromus inermis
group, B. inermis and B. biebersteinii R. & S. (2n ¼
56), B. riparius Rehm. (2n ¼ 70), and B. erectus Huds.
diploids and tetraploids became extinct in the (2n ¼ 28) with more than 40 cultivars in North Amer-
New World, with the build-up of octoploids in Cer- ica and Europe. Additionally, this section also contains
atochloa. In the Pleistocene, section Pnigma spread B. auleticus Trin. ex Nees (2n ¼ 42) used to a limited
to Africa and North America, while polyploids extent in southern Brazil and Uruguay.
increased in Eurasia. Sections Bromus and Genea Section Ceratochloa has the hexaploid B. catharti-
have evolved more recently. cus complex (2n ¼ 42), including B. stamineus
The basic chromosome number is x ¼ 7, like other E. Desv. and B. valdivianus Phil. with over 30 cultivars
Poaceae. Chromosome and nuclear genome sizes in Argentina, France, Chile, New Zealand, Australia, and
have changed considerably during speciation (Arm- USA. The octoploid B. carinatus Hook. and Arn.
strong 1991). Stebbins (1981) proposed that the trend complex (2n ¼ 56), including B. sitchensis and
was towards larger genomes. On this basis, section B. marginatus Nees ex Steud., has over five cultivars
Ceratochloa would have the most primitive genome in France and USA, while the South American octo-
size. ploid B. coloratus Steud. has one cultivar in Australia.
Comparisons of chloroplast restriction site maps of The duodecaploid B. arizonicus (Shear) Stebbins
B. inermis with the cereals have shown that Bromus is (2n ¼ 84) has one very winter active annual cultivar,
closer to the Triticeae than to the Aveneae and, within Cucamonga used in USA.
the Triticeae, it is closer to barley than to wheat and In addition, some annual species of section Bromus
rye (Pillay 1993, 1995). have had cultivars developed for revegetation uses.
Attempts to intercross species from different sec- For example, B. arvensis Guss. (2n ¼ 14) has a culti-
tions indicate that the reproductive barriers between var Dos in Russia and B. mollis L. (2n ¼ 28) has the
sections are strong. Although some intersection cultivar Blando in USA. From section Genea,
hybrids have been produced, none has shown pairing B. rubens L. has the cultivar Panache (Alderson and
of the chromosomes from the different sections, Sharp 1995).
indicating wide differentiation of the chromosomes B. mango E.Desv. (section Ceratochloa) was used
(Armstrong 1991). as a cereal grain crop in Chile. This plant was
Molecular marker and DNA sequence phylogenies cultivated as a biennial cereal by the Araucana Indians
are revealing that some of the old sections are artificial of Chile until at least the middle of the 1800s. Thought
groupings that require revision. A study of 46 species, to have become extinct (Scholz and Mos 1994), it is
representing a wide sample of the species diversity in claimed to have been rediscovered and has been clas-
Bromus, was conducted by Saarela et al. (2007), using sified as a form of B. catharticus ssp. catharticus
nuclear and chloroplast DNA sequences. This study (Vahl) Herter (Massa et al. 2004).
showed that current systematic classifications do The potential agricultural value of many of the
not fully reflect phylogeny within the genus. Some Bromus species for New Zealand was assessed by
differences between nuclear and chloroplast sequence Rumball (1968) and Rumball and Forde (1976).
2 Bromus 17

2.4 Mediterranean and SW Asian indicated that B. tectorum can outcross, giving popu-
Annual Species of Section Genea lations the capacity to absorb new genetic variation
and potentially to increase their invasive capacity
(Ashley and Longland 2007). The high polyploids,
Section Genea (subgenus Stenobromus; Stebbins 1981) B. rigidus (6x) (ripgut brome) and B. diandrus (8x)
consists of seven annual species that are self-fertiliz- (great brome) are severe weeds of crops in Australia
ing, with ploidies ranging from 2x to 8x. The diploids (Kon and Blacklow 1990) and New Zealand (Dastgheib
are B. sterilis L., B. tectorum L., and B. fasciculatus et al. 2003). These weeds form acceptable early spring
C. Presl, and the tetraploids B. madritensis L. and forage in the early growth stages, but the presence
B. rubens. In addition, there is a hexaploid B. rigidus of awns on mature seeds makes them a danger to the
Roth and an octoploid B. diandrus Roth. health of grazing animals in late spring and summer.
Among the diploid species, B. sterilis and B. tec-
torum are distinct and widespread in Europe and Eur-
asia while B. fasciculatus is restricted to the eastern
Mediterranean region. However, the widespread tetra-
ploids have variously been classified as subspecies 2.5 Section Pnigma
(Sales 1994) or separate species (Oja 2002). The hexa-
ploid and octoploid species occur in the Mediterranean Species of this large section (about 60) are native to
region and southern and western Europe. They have Eurasia, Africa, and the Americas. Two distinct spe-
also been variously classified as separate species, sub- cies groups were proposed by Armstrong (1981,
species, or varieties, and as a polyploid species com- 1983). The first was a group of long-lived perennials
plex. Most of the species of this section have become (rhizomatous or densely tufted) from Eurasia that were
weeds in many countries. mostly polyploids (4x–10x) with large anthers and
An elegant molecular phylogenetic study (Fortune small chromosomes. In this group was B. pumpellia-
et al. 2008) has shown that all of the polyploid species nus Scribn., which also occurs in North America, as
of the section have hybrid origins and, apart from one well as the Eurasian species B. inermis, B. erectus,
unidentified lineage, all three other parental lineages B. variegatus M. Bieb., and B. riparius. The second
were closely related to the three diploid species. group included mainly diploid, relatively short-lived,
Designating the diploid genomes as B. sterilis (SS), loosely tufted, non-rhizomatous species from North
B. fasciculatus (FF), and B. tectorum (TT), the poly- America characterized by small anthers and large
ploids have been proposed (female donor first) as: chromosomes. This group included the B. ramosus
B. madritensis (FFSS), B. rubens (FFTT), B. rigidus Huds. complex from Eurasia (Armstrong 1981, 1983,
(XXFFTT), and B. diandrus (XXFFTTSS), where XX 1991; Stebbins 1981). The Eurasian and North Ameri-
is an unidentified parent (Fortune et al. 2008). The two can species were reported to be difficult to hybridize
4x species were demonstrated to be distinct species, (Armstrong 1983), supporting the existence of the
and the 8x B. diandrus appears to have originated as groups and suggesting that they may have quite differ-
a hybrid between 6x B. rigidus and 2x B. sterilis. This ent evolutionary histories.
last observation has been supported by earlier research However, further analyses of these species, while
(Fortune et al. 2008). confirming the existence of groups with very differ-
Species of this section have apparently donated gen- ent chromosome sizes, have not supported a simple
omes via hybridization with species of section Bromus evolutionary situation. Analysis has not been helped
to form the tetraploid B. pectinatus Thunb. complex by difficult taxonomy in which similar taxa with
(Stebbins 1981; Saarela et al. 2007) (see Sect. 2.6). widely different ploidy levels have been given the
The main characteristic of this group of species is same species designations. Examples include B. var-
their propensity for weediness. B. tectorum is gener- iegatus, which is said to exist in a range of ploidy
ally considered to be cleistogamous and has become levels from diploid to decaploid. However, a deca-
an invasive noxious weed in dry, open areas of North ploid is very unlikely to be an autoploid form derived
America, where it is called cheatgrass or downy chess. from the diploid. It is far more likely that hybridi-
Simple sequence repeat (SSR) marker research has zation has been involved and that these taxa are
18 W.M. Williams et al.

morphologically similar but probably have very dif- Sutkowska and Mitka (2008) used random ampli-
ferent genomic constitutions. fied polymorphic DNA (RAPD) analyses to obtain
The existence of the two species groups with dif- evidence consistent with B. erectus and its close rela-
ferent chromosome sizes was established by hybridi- tives being the oldest group of species in the section.
zation studies and confirmed by nuclear DNA This evidence was also consistent with B. variegatus
analyses. Hybrids were used to show that the chromo- having the B genome.
somes of the Eurasian B. ramosus complex were larger The inconsistencies in chromosome behavior are
than the other Eurasian species B. inermis, B. variega- reflected by the results of molecular phylogenetic ana-
tus, B. erectus, B. pumpellianus, and B. riparius. How- lyses. Nuclear DNA (internal transcribed spacer, ITS)
ever, they were similar in size to B. pacificus Shear sequence analysis has shown that section Pnigma is far
and B. richardsonii Link from North America. The from being monophyletic (Saarela et al. 2007). The
North American species may all have variations of a Eurasian species of section Pnigma did not occur in
single (L) genome. Interspecific hybrids were made one clade and the American species formed several
between several of the diploid North American species well defined clades. The molecular data indicated that
and, despite good chromosome pairing, they were section Pnigma is probably an artificial group and that
sterile. This was consistent with these species all con- its current status as a section may be inappropriate.
taining the same (L) genome and in the process of This was supported by the results of C-banding and
differentiating by cryptic chromosomal rearrange- DNA content analyses (Tuna et al. 2005). However,
ments (Armstrong 1991). care is needed as reticulate evolution may have mixed
The Eurasian species have so far had two geno- the ancestral genomes such that they are apparently
mes (A and B) identified by cytogenetic analyses now distributed between continents (Sutkowska and
(Armstrong 1991) and there may be a third (C) (see Mitka 2008). There are many conflicting observations
Sect. 2.5.1). The A and B genomes are the two gen- and there is a clear need to use further molecular
omes in tetraploid forms of B. inermis and B. pumpel- analyses to resolve the species relationships in this
lianus, one of which (A) is also close to that of artificial group so that more effective use of the poten-
tetraploid B. erectus. The B genome has some homol- tially valuable wild relatives can be achieved.
ogy (homeology) with the first but is of uncertain Analysis of chloroplast DNA using species-specific
origin. Armstrong (1983) suggested that the species restriction fragment length polymorphisms (RFLPs)
of the B. ramosus complex may be the progenitors, or following interspecific hybridization has indicated
be related to the progenitors, of the American species. uniparental maternal inheritance in Bromus species
Sutkowska and Mitka (2008) also hypothesized that so far investigated (Pillay and Armstrong 2001). In
this Eurasian complex is the source of the L genome. crosses between B. arvensis (2n ¼ 14) as female and
These authors suggested that the L genome may have B. inermis (2n ¼ 28, 56), and B. erectus (2n ¼ 42, 56,
originated from the B genome, in line with the sugges- 70), all F1 plants had the female chloroplast restriction
tion (Stebbins 1981; Armstrong 1983) that the large patterns. No paternal or biparental inheritance was
chromosome species were derived from small chro- detected.
mosome species, possibly before the migration to In summary, there are about 35 Eurasian and 25
America, and before the evolution of the polyploid American species in this section. To-date, very few
species currently found in Eurasia. species have been studied, and conclusions about the
In support of the existence of two groups of species, evolution of the groups may be clarified as more species
there appeared to be little or no pairing homology in are given close research attention (Armstrong 1984).
hybrids between the large chromosomes of B. ramosus
and B. pacificus and the small chromosomes of the
Eurasian polyploids (Armstrong 1984). However,
2.5.1 B. inermis (Smooth Bromegrass,
there was also little pairing homology between the
chromosomes of B. ramosus and B. pacificus, possibly
Russian Brome)
emphasizing the difficulties of using pairing affinities
in tetraploids, where genetic control of chromosome One of the most important agricultural species,
pairing can occur (Armstrong 1984). B. inermis, belongs to this section. ITS DNA sequence
2 Bromus 19

analysis (Saarela et al. 2007) placed this species in a Interspecific hybrids indicate that the A genome
small clade with four other Eurasian species, B. erec- may come from B. erectus (2n ¼ 4x ¼ 28) (Armstrong
tus, B. korotkoyi Drob., B. pumpellianus (also native to 1991; Walton 1980). However, a similar genome also
North America), and B. riparius. Somewhat surpris- occurs in a diploid form of B. variegatus Bieb. and a
ingly, these formed a sister clade to a larger group diploid that resembles B. inermis (Armstrong 1991).
comprising all of the monophyletic sections Genea, Hybrids between diploid B. variegatus and tetraploid
Neobromus, and Ceratochloa, as well as B. bra- and octoploid B. inermis had chromosome pairing
chyanthera Doll. from South America. patterns that suggested that the B. variegatus genome
The form commonly grown in North America was not the same as either the A or B genome but was
is autoallooctoploid (2n ¼ 8x ¼ 56). There are also similar to both (Armstrong 1984). Karyotype and
allotetraploid (2n ¼ 4x ¼ 28) and hexaploid (2n ¼ Giemsa C-banding analyses have indicated that the
6x ¼ 42) forms (Tan and Dunn 1977). The >250 chromosomes of both B. erectus and B. variegatus
accessions held by USDA Plant Germplasm System differ from each other and from those of B. inermis.
are predominantly 8x, with a few 4x and no 6x forms Therefore, if either species is a progenitor of B. iner-
(Tuna et al. 2001). The tetraploid has regular meiosis mis, significant chromosomal change should have
(Carnahan and Hill 1960). The octoploid is irregular occurred post-hybridization and polyploidization
at meiosis, forming mostly quadrivalents and bivalents. (Tuna et al. 2006). Natural B. erectus x B. inermis
A chlorophyll mutant has exhibited tetrasomic inheri- hybrids have been reported from Ukraine (Sutkowska
tance and an intermediate chromosome-chromatid type et al. 2002).
of segregation pattern (Ghosh and Knowles 1964). The Tetraploid and octoploid forms of B. inermis have
genomic constitutions of the octoploid and tetraploid 2C DNA contents of 11.74 pg and 22.15 pg (Tuna
have been proposed to be AAAABBBB and AABB, et al. 2001). These values are significantly lower than
respectively, and the A and B genomes appear to be those that would have resulted from multiples of the
closely related (Armstrong 1979). Giemsa C-banding proposed diploid progenitors. Therefore, it is likely
has confirmed the alloploid nature of the tetraploid. that significant DNA loss has occurred during poly-
However, karyotype analysis and Giemsa C-banding ploidization. Such loss of DNA has been recorded for
has indicated that the octoploid cytotype is not the the Triticeae (Vogel et al. 1999).
same as a doubled version of the tetraploid cytotype Hybrids with B. pumpellianus ssp. dicksonii indi-
(Tuna et al. 2004). There is evidence that the genomic cated that the two species have similar chromosomes,
constitution could be AAAABBCC, based on karyo- differing only by inversions and translocations
type analyses showing that B. inermis has two pairs (Armstrong 1982).
of chromosomes with large satellites and only one
pair with small satellites (Ghosh and Knowles 1964; 2.5.1.1 Genetic Diversity in B. inermis
Wilton 1965; Armstrong 1973). However, this charac-
teristic appears to be either inconsistent or polymor- Based on molecular analyses, the present distribution
phic (Rychlewski 1970; Armstrong 1981). Tuna et al. of B. inermis in Poland was attributed to post-glacial
(2004) confirmed the inconsistency of expression of migrations from two separate refuges (Sutkowska
the small satellite chromosomes and generally also et al. 2002). North American breeders of B. inermis
confirmed the karyotypic analysis of Armstrong generally classify the natural variation of the species
(1977b) showing two large satellite pairs and one into two ecotypes – a northern or “meadow” type adap-
small satellite pair. These authors indicated that the ted to valleys and moist regions of eastern Europe and
AAAABBBB hypothesis of Armstrong (1977b) was temperate Asia, and a southern, or “steppe” type
not supported by their chromosome analyses. To-date, adapted to dry steppe regions. A third, intermediate
the genome of B. inermis has remained relatively intrac- group has developed apparently by intermixing of the
table to cytogenetic analyses using karyotype and first two. The ecotypes differ morphologically in
C-banding. There is a need for the application of fluo- root depth and leaf size (Fernandez and Coulman
rescence in situ hybridization (FISH) and genomic in 2004). Amplified fragment length polymorphism
situ hybridization (GISH) techniques. (AFLP) analyses of 14 cultivars revealed that the
20 W.M. Williams et al.

older (pre-1980) varieties based on southern germ- be resolved (Pavlick 1995). However, its description
plasm were distinctive and less diverse than varieties fits that for B. riparius according to Smith (1980). It is
based on northern germplasm and more recent south- a long-lived perennial with chromosome numbers of
ern or mixed varieties. These results suggest that inter- 2n ¼ 2x ¼ 14, 2n ¼ 8x ¼ 56, and 2n ¼ 10x ¼ 70.
mixing of southern and northern ecotypes has occurred In addition to being a useful species itself, it is a
recently in North American breeding programs (Fer- potential source of germplasm for the improvement
nandez and Coulman 2004). A diversity analysis of of B. inermis, it has lower growing points and so
cultivars based on morphological characteristics (Cas- recovers better from defoliation, is less aggressively
ler et al. 2000) indicated that similar morphological rhizomatous, and has a longer growing season into the
types had been developed from very diverse genetic autumn than B. inermis (Armstrong 1991).
backgrounds. The common cultivars have 2n ¼ 10x ¼ 70 and
are believed to have the same chromosome constitu-
tion as B. inermis plus an additional genome of
2.5.1.2 Breeding Progress
unknown origin (Armstrong 1991). Sutkowska and
Mitka (2008) obtained evidence using RAPDs that
In recent years, there has been very little private sector
this additional genome could be closely related to, or
breeding of B. inermis and relatively little cultivar
ancestral to, the L genome. Diploid B. riparius has a
development by public sector plant breeders. Empha-
somatic cell (2C) DNA content of 6.14  0.09 pg
sis on selection for improved quality has significantly
(Tuna et al. 2001) – very similar to B. erectus (Tuna
improved digestibility and reduced fiber concen-
et al. 2006). Common B. riparius has a 2C DNA
trations. Yield improvement has been slow, not only
content of 22.15 pg (Tuna et al. 2001).
because of the low effort but also because of the
The diploid form of B. riparius was found in
complex polyploid genetics and research emphasis
Kazakhstan and is quite similar to the tetraploid
on genetics rather than breeding (Casler et al. 2000).
B. inermis from the same region. This led Armstrong
There has been almost no molecular breeding in
(1987) to suggest that diploid B. riparius might be a
Bromus. One of very few studies investigated the
progenitor of polyploid B. inermis. A C-banding anal-
prospective use of marker-assisted selection for forage
ysis of diploid B. riparius was completed by Tuna
quality using neutral detergent fiber (NDF) concentra-
et al. (2001), and Tuna et al. (2004) analyzed the
tion as a predictor of animal intake (Diaby and Casler
respective karyotypes and showed that B. riparius is
2005; Stendal et al. 2006). Although RAPD markers
an unlikely progenitor of B. inermis. Sterile hybrids
were difficult to use because of low repeatability, the
between B. variegatus and diploid B. riparius have
association of markers with NDF was established.
been produced and were reported to have normal
This has opened the way for marker-assisted selection
chromosome pairing behavior (Armstrong 1991).
for NDF concentration using more repeatable marker
B. riparius is widely grown in North-central USA
systems.
and in Canada. It resembles B. inermis in appearance
B. inermis is one of the most freezing-tolerant
but has shorter rhizomes, awned seeds, and pubescent
perennial grass species. Consequently, it is the subject
leaves (Knowles et al. 1993). The cultivar “Regar”
of considerable research into the genetics of freezing
was released in 1966 by the Colorado Experiment
tolerance, as well as being a potential genetic resource
Station.
for isolation of important freezing tolerance genes (see
Introgression of B. riparius traits (e.g., lower
Sect. 2.10).
growing points and less vigorous rhizomes) into
B. inermis would be useful. To this end, Armstrong
(1990) obtained fertile hybrids between B. inermis
(2n ¼ 8x ¼ 56), as female, and B. riparius (2n ¼
2.5.2 B. riparius Rehm. (Meadow
10x ¼ 70) without difficulty. The F1 plants were
Bromegrass) backcrossed to B. inermis as male and produced
seed when open-pollinated. They had 2n ¼ 63 chro-
This species is often called B. biebersteinii and is mosomes. The F2 population ranged from 2n ¼ 56 to
confused with B. erectus, and its taxonomy is still to 72 and tended to be in the 63–70 range. Chromosome
2 Bromus 21

pairing results were equivocal and there was no clear and other disturbed places, as well as in pastures. It is
evidence for or against recombination between the often confused with B. riparius. Because it probably
genomes of the two species. However, there was a shares genomes with B. inermis, B. erectus is a potential
suggestion that reversion to the parental types source of genetic diversity for B. inermis breeding. It
occurred, as had been observed earlier by Nielsen also shares a genome with B. pumpellianus (Armstrong
et al. (1965). Backcross progenies rapidly reverted 1981).
to the parental chromosome numbers and appeared to
lose B. riparius traits as they did so. Cytoplasmic
effects were also apparent, and no clear strategy
emerged for the use of B. riparius germplasm in the 2.5.4 B. variegatus
improvement of B. inermis. Nevertheless, a hybrid
population was selected and interpollinated for sev-
B. variegatus forms part of the meadow bromegrass
eral generations by Knowles and Baron (1990).
complex of decaploid (2n ¼ 70) species, along with
Selections from this population were analyzed using
B. riparius and B. biebersteinii. A diploid population
RAPD markers and compared with the parental cul-
of this species is one of very few diploids among the
tivars (Fernandez et al. 2001). A hybrid population
Eurasian species of the section. It has a somatic cell
was genetically intermediate between the parents,
DNA content of 6.76  0.05 pg (Tuna et al. 2006).
although closer to B. inermis, possibly because it
As already discussed, the role of the diploid form
had been selected for B. inermis traits. Thus, it is
in the ancestry of the polyploid species remains
possible that recombination had occurred between
unresolved.
B. inermis and B. riparius genomes in that population
and that similar hybrids may be useful for the genetic
improvement of smooth bromegrass.
2.5.5 B. pumpellianus (Arctic
Bromegrass)
2.5.3 B. erectus (Erect Bromegrass)
This octoploid (2n ¼ 8x ¼ 56) species has popula-
This diploid perennial species has a somatic nucl- tions that are native to Asia, as well as the mountains
eus (2C) DNA content of 6.19  0.08 pg (Tuna of western North America, where it provides useful
et al. 2006). Armstrong (1973) used B. erectus (2n ¼ high quality forage in high altitude grasslands up to
4x ¼ 28) to make 6x hybrids with B. inermis (2n ¼ 3,350 m altitude (Casler and Carlson 1995). Hybrids
8x ¼ 56). On the basis of chromosome pairing and between 8x B. inermis and 8x B. pumpellianus are
karyotype analysis, he concluded that the B. erectus fertile, suggesting that they are subspecies (Armstrong
genome was present in tetrasomic condition in B. iner- 1991). However, there is confusion because, at the
mis. Armstrong (1977b) carried out further analysis of tetraploid level, they appear to be different species.
the karyotypes of these hybrids and B. inermis and Armstrong (1985) suggested that these are very het-
favored the view that the B. erectus genome constituted erogeneous taxa that involve allopolyploidy and
the A genome in 8x (AAAABBBB) B. inermis. How- have undergone introgression. A variety, “Polar” was
ever, he was unable to rule out the AAAABBCC developed from such interspecific hybrids (Hodgson
hypothesis of Ghosh and Knowles (1964). et al. 1971).
B. erectus is naturally distributed in Europe, Britain,
Ireland, and North Africa (Meusel et al. 1965) and is
introduced and widespread in the North-Eastern states
2.5.6 Other North American Species
of USA and sporadically elsewhere (Pavlick 1995).
The present distribution of B. erectus in Poland was in Section Pnigma
studied by polymerase chain reaction (PCR) and attrib-
uted to a single post-glacial expansion (Sutkowska et al. Nodding brome (B. anomalus Rupr. ex Fourn.) and
2002). It is a tufted perennial that occurs on roadsides tetraploid (2n ¼ 28) fringed brome (B. ciliatus L.)
22 W.M. Williams et al.

provide native grazing in the western mountain native pastures (Martinello and Schifino-Wittmann
regions of North America (Casler and Carlson 1995). 2003). It has caught the attention of agronomists and
A diploid form of B. ciliatus is considered to be the plant breeders, and some isozyme and RAPD charac-
ancestral American species (Armstrong 1991). A terizations have been done (Yanaka 2002). B. uru-
detailed Giemsa C-banding chromosome analysis of gayensis is also hexaploid, while B. macranthus has
B. ciliatus by Tuna et al. (2005) revealed that the been reported as having 4x, 8x, 10x, and 16x chromo-
genomes in this American species are quite different somes (Stebbins 1981). It is uncertain how these spe-
from those of any European species in the section cies are related to the remainder of section Pnigma.
Pnigma. In addition, the 2C nuclear DNA content of
B. ciliatus was 19.13 pg, contrasting strongly with that
of tetraploid B. inermis (11.74 pg). The chromosomes
of B. ciliatus were almost double the length of those of
2.6 Section Bromus
Eurasian species – consistent with previous observa-
tions that the two groups of species differ in chromo- This section of about 40 species is considered to be the
some size. Additionally, the C-banding indicated quite most advanced. It is native to Europe and Asia and
different patterns of constitutive heterochromatin. consists of annual or biennial diploid and tetraploid
These observations, along with the strong genetic iso- species. The tetraploids are likely to have hybrid ori-
lation between the two groups, support the contention gins (Stebbins 1981). Southwest Asia and the eastern
that they are not closely related. Mediterranean regions are the centers of diversity
B. ciliatus is of interest in North America as a (Stebbins 1981). Several annual or biennial species
potential outcrossing native grass species for restoring have spread widely beyond their native regions and
vegetation to ecologically important sites. Genetic have become significant weeds, e.g., B. briziformis
diversity of Canadian populations was studied by Fu Fisch. and C.A. Mey., B. commutatus Schrad. (meadow
et al. (2005) using AFLPs. This identified regional brome), B. hordeaceus L. (B. mollis), B. japonicus
Canadian forms. However, there was surprisingly lit- Thunb. in Murr., B. racemosus L., B secalinus L., and
tle intrapopulation variation, suggesting that perhaps B. squarrosus L. are now widespread throughout North
more self-fertilization occurred than was expected. America (Pavlick 1995).
This emphasized the need for proper breeding system Only a small number of artificial interspecific hy-
studies to be carried out where genetic diversity is an brids have been made within section Bromus (Armstrong
important element in the end-use of a species. 1991). B. mollis was crossed with B. arenarius (Knowles
Although most North American species are diploid, 1944), leading to an indication that B. mollis contains
B. frondosus (Shear) Woot. and Standl., B. richardso- two distinct genomes and that the same two occur
nii, and B. pacificus all behave like allotetraploids in B. racemosus. One of the genomes in B. mollis was
(Armstrong 1984). B. mucroglumis Wagnon from partly homologous with one of the genomes in B. are-
Mexico and adjacent areas of the USA is similar to narius (4x).
B. richardsonii (Peterson et al. 2001), but the other B. commutatus, B. macrostachys, and B. secalinus
three tetraploid species in North America do not are allotetraploids. Hybrids of B. arvensis (2x) x
appear to have been studied. Hybrids between tetra- B. commutatus (4x) and B. secalinus (4x) showed
ploid B. ciliatus and B. frondosus indicated similar some chromosome pairing, indicating a relationship
genomic structures (Barnett 1957). between the genome of B. arvensis and those in the
tetraploid species (Jahn 1959). There were also pairing
homologies between some chromosomes of B. macro-
stachys and B. mollis.
2.5.7 South American Species in Section
Sections Boissiera (one species, B. pumilio (Trin.)
Pnigma P.M. Smith) and Triniusia (two species, B. danthoniae
Trin. ex C.A. Mey., B. pseudodanthoniae Drobow),
Native to southern Brazil and Uruguay, B. auleticus recognized by some authors (Smith 1985; Scholz
Trin. ex Nees is a perennial allogamous hexaploid 1998; Saarela et al. 2007), were included in Section
(2n ¼ 6x ¼ 42) that provides outstanding forage in Bromus by Smith (1970). They comprised annual
2 Bromus 23

species from Asia and the eastern Mediterranean than expected frequencies, suggesting that outcrossing
region. ITS and chloroplast DNA sequence analysis was a significant evolutionary factor in these predom-
(Saarela et al. 2007) confirmed that the species of inantly self-fertilized diploids.
section Triniusia belong in section Bromus. B. arvensis is native to the Mediterranean and into
The B. pectinatus complex (five 4x species rang- southern and central Europe and is widespread in
ing from Africa to Asia) is intermediate in morphol- Eurasia (Oja and Paal 2007). It has been introduced
ogy between sections Bromus and Genea, leading to Australia and the USA in the 1920s where it became
Stebbins (1981) to suggest that these species arose adapted to the corn-belt region and eastward. It pro-
as hybrids between species of the two sections. The vides excellent winter cover, soil stabilization, and
DNA sequence data of Saarela et al. (2007) support green manure as a result of its extensive root system.
this idea. If this is correct, then section Bromus is B. japonicus and B. squarrosus are common in the
monophyletic. Mediterranean and Southwest Asia and have become
significant weeds in several countries.
B. arvensis (2n ¼ 14) has been hybridized with the
perennial species B. inermis (2n ¼ 56) and B. erectus
2.6.1 B. arvensis (Field Bromegrass)
(2n ¼ 28) of section Pnigma, but the hybrids were
and Its Close Relatives sterile (Armstrong 1977a). This intersectional hybridi-
zation does not appear to be a fruitful source of germ-
This diploid (2n ¼ 14) winter annual species and its plasm for the widening of the B. inermis or B. erectus
close relatives, B. japonicus and B. squarrosus, form a gene pools (Armstrong 1977a). However, Armstrong
complex that is taxonomically difficult and has been (1977b) was able to use 5x hybrids between B. arven-
subject to several different treatments. B. arvensis is sis and B. inermis to analyze the alloploid gametic
allogamous (Oja et al. 2003) unlike the other two, (AABB) chromosome complement of B. inermis
which are almost exclusively self-fertilizing. Serolog- because the B. arvensis chromosomes were much
ical analysis (Smith 1972), isozyme analysis (Oja et al. larger and could be clearly distinguished.
2003), and DNA analyses (Ainouche and Bayer 1997;
Ainouche et al. 1999) indicated that B. japonicus and
B. squarrosus might be sister species or a species
complex distinct from B. arvensis. Oja et al. (2003) 2.6.2 B. hordeaceus
suggested that B. japonicus and B. squarrosus are
possibly self-fertilizing derivatives of B. arvensis. Also known as B. mollis, this is an aggressive species
However, a later morphological separation, using dis- that has a wide climatic range that has enabled it to
criminant analysis to determine the most useful char- spread widely within Eurasia, Africa, the Americas,
acters for distinguishing the taxa (Oja and Paal 2007), and Australia. It is a probable allotetraploid (2n ¼ 28,
indicated that the three species could be fairly reliably Stebbins 1981). It shows wide morphological variation
separated using floral morphological traits, especially indicative of ecotype differentiation and, in the Medi-
anther length and lemma width. terranean region, may hybridize with B. lanceolatus.,
The Mediterranean self-fertilizing diploid B. inter- a closely related tetraploid (Ainouche et al. 1995). On
medius Guss. is also very similar in morphology to the basis of its morphological variation, Pavlick
B. japonicus var. villosus (Oja 2005) and apparently (1995) separated the species into four subspecies. It
also belongs to this species complex. This is supported has been the subject of analyses using isozymes to
by isozyme data (Oja 2005) and ITS sequence analysis relate population genetic diversity to habitat diversity
(Ainouche and Bayer 1997). It may also be a self- in Australia (Brown et al. 1974). This led to the iden-
fertilizing derivative of B. arvensis (Oja 2005). Iso- tification of fitness-related polymorphism at the
zyme analyses (Ainouche et al. 1995) indicated that alcohol dehydrogenase locus (Brown et al. 1976).
North African populations of B. intermedius and its Isozyme analyses have also indicated quite high out-
close relative B. squarrosus were much less variable crossing rates in some populations (Brown et al.
than the tetraploids B. hordeaceus and B. lanceolatus. 1974). However, Ainouche et al. (1995) found high
Nevertheless, heterozygotes were present in higher levels of intragenomic homozygosity in North African
24 W.M. Williams et al.

populations, suggesting high selfing rates in this spe- time, many of these so-called species have become
cies (and B. lanceolatus). High levels of intergenomic amalgamated into a hyper-variable species. In this
heterozygosity in these two species were consistent sense, it may be fairer to say that the hexaploids are
with their allotetraploid genomic structure. just over the “cusp” of speciating. It is unclear how the
African and American species relate.
The commonly named taxa that constitute this
hexaploid group from South America (following
2.7 Section Ceratochloa Planchuelo and Peterson 2000) are B. catharticus,
B. coloratus, B. lithobius, B. stamineus, B. mango,
Section or subgenus Ceratochloa is a small section and B. tunicatus Phil. Using a multivariate analysis of
consisting of up to 16 polyploid perennial and annual this group, including morphological and molecular
species. No diploid or tetraploid species are found in data, Massa et al. (2001, 2004) indicated that the
this section. The section Ceratochloa has the smallest variation was consistent with the existence of a single
genome size in the genus (Armstrong 1991). Hybridi- hexaploid species, B. catharticus, with two nearly
zation is rife in this section, making species bound- continuous subspecies, B. catharticus ssp. catharti-
aries obscure and the taxonomy very difficult. cus (Vahl) Herter and B. catharticus ssp. stamineus
Section Ceratochloa appears to be monophyletic (E. Desv.) Massa. B. catharticus ssp. catharticus
and most species display almost identical chloroplast includes, among others, the following older taxa:
DNA sequences (Pillay and Hilu 1990, 1995), suggest- B. unioloides Kunth, B. tunicatus, and B. burkartii
ing a similar maternal genome. Similarly, the ITS Munoz, as well as B. mango, while B. catharticus
sequence and other nuclear DNA sequence variation ssp. stamineus includes B. stamineus, B. coloratus,
is trivial (Saarela et al. 2007), a situation found in most B. lithobius, B. fonkii Phil., and B. valdivianus Phil.
post glacial grass expansions. This suggests that much These authors identified octoploid forms of B. color-
of Ceratochloa is very recent and post glacial, a con- atus and B. lithobius in South America, which they
clusion drawn by Stebbins (1981). The agricultural reclassified as B. coloratus Steud.
value of many of the species in section Ceratochloa These three newly defined taxa are separable using
was reviewed by Stewart (1996). a simple three-step morphological key. There are also
The species of section Ceratochloa can be divided differences in adaptation, the populations of ssp. sta-
into four genomically distinct classes: mineus being perennial, predominantly cleistogamous
and from humid places, while those of ssp. catharticus
are facultatively cleistogamous and are from high alti-
tudes in open woodlands of the precordillera as well as
2.7.1 The Hexaploid B. catharticus
the lower slopes of the Andes.
Complex (2n ¼ 42) Species from this complex have been introduced to
many countries and there are more than 30 cultivars
This group of species is endemic to South America marketed in Argentina, France, Chile, New Zealand,
(Massa et al. 2004), with members also present in Australia, and USA.
Africa, B. leptocladis in South Africa and B. runssor-
oensis in highland Central Africa. The hexaploid
species of section Ceratochloa are all found to be
strict allopolyploids with genomic formula AABBCC 2.7.2 Two Disjunct Octoploid Groups
(Stebbins and Tobgy 1944). These genomes are
almost indistinguishable and have a nuclear DNA These are the B. carinatus Hook. and Arn. complex
content of 12.7–15.1 pg (mean 1C ¼ 2.32 pg) (Klos (2n ¼ 56), found in higher latitudes of North America,
et al. 2009). and the 8x B. coloratus referred to above, found in the
Morphological variation in these hexaploids is higher latitudes of South America (Massa et al. 2004).
large and it is fair to say this group is the oldest of They have a genomic constitution ABCL where L is a
the four groups in Ceratochloa. Originally, this com- larger genome, probably from section Pnigma. DNA
plex was separated into a number of species but, over contents of B. carinatus (20.9–22.9 pg/cell) were more
2 Bromus 25

than 70% larger than B. catharticus (13.0–14.9 pg/ cultivars have been developed and are used in the
cell) (Joachimiak et al. 2001). Therefore, the octoploid NW USA and Canada, as well as in New Zealand.
species of the B. carinatus complex are considered
to be intersectional amphidiploids between diploids
of section Pnigma and hexaploid species of section 2.7.2.3 B. sitchensis (Sitka or Alaska Brome)
Ceratochloa (Stebbins 1956). This was supported by
Pillay (1996) who found rDNA sequences common Native to mountain meadows of the Pacific coast of
between this group of species and B. inermis. The North America from Washington state northwards to
seven species of the B. carinatus complex are Alaska. It is closely related to B. aleutensis and is used
B. aleutensis Trin. ex Griseb., B. carinatus sens. str., for pastures in northern Europe.
B. marginatus, B. maritimus (Pip.) A.S. Hitch.,
B. polyanthus Scribn., B. sitchensis, and B. subveluti-
nus Shear. Based on intergrading morphologies and 2.7.3 B. arizonicus (2n ¼ 84) (Arizona
partial interfertilities, some taxonomists believe that
all of these taxa belong to a single diverse species,
Brome)
B. carinatus sens. lato (e.g., Soderstrom and Beaman
1968; Stebbins 1981). However, Pavlick (1995) This duodecaploid is native to California and east
argued that the major differences in morphology and to Arizona. Morphological and cytological analyses
barriers of hybrid sterility justify separation into sepa- performed by Stebbins et al. (1944) indicated that
rate species. A revision of this group using molecular B. arizonicus is an allopolyploid derived from B. cath-
phylogenetic methods is required (Saarela et al. 2007). articus and hexaploid B. trinii Desv. in Gay (B. berter-
The main agricultural species of the group are as ianus Colla) or an unknown close relative of this
below. species. B. berterianus is the representative of the sec-
tion Neobromus, native to the Pacific coast of North and
South America. The genomic constitution of B. arizo-
nicus was postulated as ABCC’DE (Stebbins 1947).
2.7.2.1 B. carinatus (California Brome)
Klos et al. (2009) have shown that all twelve genomes
in B. arizonicus are approximately the same average
This is annual, biennial, or a short-lived perennial with
size (2.1–2.3 pg) and the same size as the genomes in
deep roots, strong leafy growth, and good seed pro-
the hexaploid Ceratochloa species. A winter active
duction that is used for grazing animals. It is naturally
annual cultivar, Cucamonga, is used in California.
distributed along the west coast of North America
from the Canadian border region to Baja California.
Two major varieties are often distinguished (e.g. Pavlick
1995) – var. carinatus generally west of the Sierra 2.7.4 Duodecaploid (2n ¼ 84)
Nevada and Cascade mountain ranges and the more Accessions Found in Andean
perennial var. hookerianus (Thurb.) Shear. to the east Regions of South America
in the Columbia River Basin. The two intergrade and
var. hookerianus intergrades with B. marginatus and
Two 12x accessions investigated by Klos et al. (2009)
B. subvelutinus at higher altitudes to the east.
differed markedly from B. arizonicus in nuclear DNA
content and in chromosome size. In particular, these
South American duodecaploids had some large chro-
2.7.2.2 B. marginatus (Mountain Brome) mosomes, making up about 37% of the genome. This
group potentially has the genomic constitution
This perennial occurs generally at high altitudes ABCLLL (Klos et al. 2009) and is clearly different in
throughout the western half of North America from constitution from the North American duodecaploid,
Canada to N. Mexico. It intergrades with B. carinatus B. arizonicus. These accessions may belong to the
and B. subvelutinus to the West, B. aleutensis to the species B. ayacuchensis (Saarela et al. 2006), for
North, and B. polyanthus to the Southeast. Several which a chromosome number has not been determined.
26 W.M. Williams et al.

2.8 Section Neobromus 2.10.3 Protoplast Fusion

This section consists of two annual hexaploids native Fusion of protoplasts from hexaploid (2n ¼ 42) wheat
to the western coasts of North and South America., cv. 99P embryo-derived callus and UV-irradiated pro-
including B. gunckelli Matthei and B. berterianus toplasts from tetraploid (2n ¼ 28) B. inermis resulted
(B. trinii, Pavlick 1995). in asymmetric somatic hybridization (Xiang et al.
1999). Three albino hybrid explants were obtained
with 42–54 chromosomes, including small chromo-
somes and chromosome fragments from B. inermis.
2.9 Section Nevskiella

This section comprises one species, B. gracillimus 2.10.4 Genome Mapping

Bunge from Asia and the eastern Mediterranean.

The B. inermis chloroplast genome was physically

mapped by Pillay (1993) using barley and wheat chlo-
2.10 Role of Bromus Species in Crop roplast DNA probes to map restriction sites. The orga-
Improvement Using Biotechnology nization of the Bromus chloroplast genome was
similar to other grasses, with a pair of inverted repeat
regions flanked by single copy regions. DNA sequence
2.10.1 Tissue Culture colinearity was shared with wheat and other cereal
grasses.
Callus cultures from immature inflorescences of
B. inermis have been grown and regenerated into
green plants (Wattanasiri and Walton 1993). Signifi-
cant somaclonal variation was detected for several
2.11 Bromus Species in Genetic
plant traits, and although none of 10 somaclones was Research
taller than the parent plant in a replicated trial, some
had more tillers and larger leaf-stem ratios. It was 2.11.1 Herbicide Resistance
concluded that this approach should aid the develop-
ment of superior clones of B. inermis.
B. tectorum (downy brome) has developed resistance
to herbicides based on acetolactate synthase (ALS) –
inhibitors and PS II inhibitors and ACCase inhibitors.
This multiple herbicide resistance was traced, in part,
2.10.2 Cell Culture to a single amino acid substitution in protein product
of the psbA gene, the target site of PS II inhibitors
Non-embryogenic suspension cell cultures of B. iner- (Park and Mallory-Smith 2005).
mis have been used for over 20 years and this system
has been thoroughly characterized (Ishikawa et al.
1990; Robertson et al. 1994; Wilen et al. 1996). It
has proved to be useful for analysis of the mechanisms 2.11.2 Genetic Diversity and Weediness
of stress tolerance induced either by drought or cold
treatment or by abscisic acid (ABA) in warm condi- Molecular markers (mainly AFLP and SSR) have been
tions (e.g., Lee and Chen 1993; Wu et al. 2005). used to study the population genetics of the spread
Nakamura and Ishikawa (2006) have successfully of B. tectorum in North America. As noted earlier,
transformed B. inermis cell cultures using Agrobacter- although this species is treated as an obligate self-
ium tumefaciens, opening the way for the analysis of pollinator, molecular evidence indicates some out-
gene functions through reverse genetics. crossing. Nevertheless, strong associations have been
2 Bromus 27

found between marker allele presence and phenotypic species. However, this work should be supported
variation in several adaptive traits (Ramakrishnan by new DNA sequence analyses to resolve species
et al. 2004, 2006), suggesting that marker diversity relationships, especially in the large section Pnigma,
can be used to infer adaptive variation. where hybridization and polyploidy, followed by
genomic changes, have mixed the genomes and
blurred species relationships.
There is an on-going need to collect Bromus spe-
2.12 Endophytic Fungi in Bromus cies from the wild in both Eurasia and in the Americas
in order to realize the potential offered by the genus
Many grasses host choke forming Clavicipitaceae for agricultural purposes.
endophytic fungi of the genus Epichlöe or their asex-
ual derivatives Neotyphodium. The sexual stroma of
Epichlöe causes “choke disease,” which prevents
seedheads emerging. Non-choke inducing asexual References
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fescue (Festuca arundinacea), and they provide the Bromus species (section Bromus, Poaceae): insights from
host with advantages such as improved insect resis- internal transcribed spacer sequences of nuclear ribosomal
tance and grazing deterrence (Schardl and Phillips DNA. Genome 40:730–743
Ainouche ML, Misset MT, Huon A (1995) Genetic diversity in
1997). Mediterranean diploid and tetraploid Bromus L. (section
The Bromus species at present known to host Bromus Sm.) populations. Genome 38:879–888
endophytes appear to be restricted to section Pnigma, Ainouche ML, Bayer RJ, Gourret JP, Defontaine A, Misset MT
with reports for B. auleticus, B. brachyanthera, (1999) The allotetraploid invasive weed Bromus hordeaceus
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B setifolius, B. benekenii, B. erectus, B. ramosus, tion. Folia Geobot 34:405–419
B. tomentellus, and B. anomalus (Groppe et al. 2002; Alderson J, Sharp WC (1995) Grass varieties in the United
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cant ecological effects and may well provide insect Allred KW (1993) Bromus section Pnigma in New Mexico, with
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The full relevance of endophyte fungi in Bromus is (2n ¼ 56). Can J Genet Cytol 15:427–436
not known. However, it will be important that endo- Armstrong KC (1977a) Hybrids of the annual Bromus arvensis
with perennial B. inermis and B. erectus. Z Pflanzenz€ucht
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28 W.M. Williams et al.

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Chapter 3
Cenchrus

S. Goel, H.D. Singh, and S.N. Raina

3.1 Introduction Germplasm Resource Information Network (GRIN)

of USDA recorded 35 different species of Cenchrus.
This genus belongs to the tribe Paniceae of Poaceae
Grasses (Poaceae) feed the world either directly as
family. Majority of the taxa in the tribe Paniceae are
food crops, such as wheat rice, millets, and other
polyploids with basic chromosome no x ¼ 9 or 10;
grains, or indirectly as primary fodder for the live-
however, basic chromosome number determination is
stock. It is estimated that grasslands cover around
almost impossible due to the presence of an aneuploid
20% of the earth’s total land area. Cenchrus, a member
series in this taxa. This aneuploid series results in
of the tribe paniceae of the Poaceae family, is also
abnormal meiotic chromosome behavior during sexual
one of the important components of major grass cover
reproduction, resulting in sterility and subsequently
of the world. It is distributed throughout the arid and
culminating in predominance of aposporic embryo
semiarid tropical regions of the world (Bogdan 1977).
development (Snyder et al. 1955). But both sexual
Considering the diversity and present-day distribution,
and apomictic modes of reproduction have been
Cenchrus probably originated in the eastern tropical
reported in C. ciliaris.
Africa and tropical Asia and is widely naturalized in
new world countries.
Cenchrus ciliaris and two closely related species,
C. setiger and C. pennisetiformis, are three main spe-
cies of Cenchrus having the potential for forage 3.2 Morphology, Taxonomy,
production across the world. They are well adapted and Geographical Distribution
to harsh climatic conditions and flourish vigorously of Genetic Diversity
with the availability of soil moisture. C. ciliaris is
more widespread and more valued as forage grass for
The name Cenchrus comes from the Greek word
dry areas because of its high biomass production and
“Kenchros” meaning millet. The Latin word Cenchrus
tolerance to low rainfall conditions. Some of the spe-
was used by Plinius, for an Arabian diamond or an
cies of Cenchrus are also potential invader weeds.
unknown kind of piece, big as a grain of millet (Quat-
After invasion, they replace the native species by
trocchi 2008). Cenchrus belongs to family Poaceae
changing not only the ecophysiological characteristics
(Gramineae), subfamily Panicoideae, tribe Paniceae.
but also the fire regime.
The subfamily Panicoideae includes approximately
The grass base of Royal Botanical Garden, Kew
208 genera and ~3,300 species. Cenchrus is an
has recorded 25 species of Cenchrus. However,
extremely variable genus having both annual and
perennial species. It is tufted, sometimes shortly rhi-
zomatous grass with branching culms. Culms are
S.N. Raina (*)
ascending, erect, or decumbent and stoloniferous,
Amity Institute of Biotechnology, Amity University, Sector 125,
Noida, Uttar Pradesh, India ranging from 68 to 300 cm in length (Clayton et al.
e-mail: [email protected] 2006 onwards). Some species reach up to 9 feet in

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 31
DOI 10.1007/978-3-642-14255-0_3, # Springer-Verlag Berlin Heidelberg 2011
32 S. Goel et al.

height. Leaves are with ligules, either with a ciliate All species of Cenchrus are very much similar in
membrane or fringe of hairs. Leaf blades are flat, their morphology. The genus Cenchrus has been sepa-
linear, or lanceolate. It is a bisexual plant with an rated further to different species level mainly based
erect or nodding cylindrical spike-like panicle inflo- on the nature of bristles and their degree of fusion.
rescence with angular rachis. Panicle has an axis bear-
ing deciduous spikelet clusters. Each cluster has one to 1. Cenchrus argimonioides Trin., commonly known
eight spikelets subtended by a deciduous involucre of as agrimony sandbur in US and as kamanomano in
flattened and often spiny bristles, which are connate at Hawaii, is native to North Pacific regions of the
the base. Spikelets are sessile, lanceolate or ovate, world, i.e., western coast of the US and Hawaii
closely compressed, and unawned. Each spikelet has Islands (GRIN Taxonomy USDA) and is endemic
one basal sterile or male floret and one fertile (herma- to this region (Quattrocchi 2008). This is a peren-
phodite) floret. Each floret has one or two glumes, nial species with robust culms, which are
which are unequal, shorter, or longer than the adjacent 25–200 cm long. Leaf blades are 16–41 cm long,
lemmas. Lemmas are as long as the spikelet, membra- 6–16 mm wide, green to bluish green in color.
nous to coriaceous, the flat margins of which cover Panicle spiciform, loose, 7–28 cm long. Decidu-
much of palea. Lower glume is minute or is sometimes ous spikelets in clusters of 1–2 spikelets sub-
suppressed. Lodicules are absent in some species and tended by an involucre of numerous deciduous
are with two lodicules in some other species. Flowers bristles, which connate 2–3 mm forming a cup at
are with three anthers and two stigmas. Fruit is small, the base. Bristles are rigid, retrorsely scaberulous,
typical of grass, i.e., caryopsis with adherent pericarp, ciliate and spiny (Clayton et al. 2006 onwards).
dorsally compressed. The shape of the fruit varies Fertile spikelets with 1 basal sterile and 1 fertile
from ellipsoidal, oblong, or ovoid to obovoid. Endo- floret. Basal sterile floret barren and is with sig-
sperm is hard, is without lipids, and contains only nificant palea. Lemma of lower sterile floret with
simple starch grains. 5 veins.
Morphologically, Cenchrus is very similar to 2. Cenchrus biflorus Roxb., also known as Indian
Pennisetum with few species of Setaria namely Sandbur, is native to Indian subcontinent and
Setaria grisebachii and Setaria macrostachya, which tropical African countries (GRIN Taxonomy
often lead to confusion for identification but can be USDA; Clayton et al. 2006 onwards; Quattrocchi
differentiated from each other by the morphology of 2008). It is an annual species with tufted habit.
the involucral bristles. In Cenchrus, bristles are fused The culms are 5–90 cm long, ascending to erect
with each other at the base, forming a spiny cup, but and are unequal in length. The upper internodes
they are free in Pennisetum (Stieber and Wipff 2000; are longer than the lower (Clayton et al. 2006
Tu et al. 2002). In C. ciliaris, the extent of fusion is onwards). Leaf blades 2–25 cm long, 2–7 cm
minimal, which led to its placement in the genus wide. Panicle 2–15 cm long with axis bearing
Pennisutum as Pennisetum ciliare (Fisher et al. 1954; deciduous spikelet in clusters of 1–3 spikelets
Taliaferro and Bashaw 1966; Vielle-Calzada et al. subtended by a rigid involucre of two whorls of
1996). The membership of the C. ciliaris in genus bristles, which connate at the base for 0.5–1 mm,
Cenchrus is justified by its close relationship with forming a diamond-shaped shallow disc at the
C. setiger. In Setaria, the spikelets are borne above base. Involucral bristles ovoid in shape, 4–11 mm
the bristles, and the bristles remain attached to the long retrorsely barbellate and tenaciously prickly.
rachis of the inflorescence even after the spikelets Inner spines flattened and their abaxial surface
fall off, whereas in Cenchrus, the bristles fall off with 1–3 grooves. Outer bristles numerous and
with the spikelet as a unit at maturity. This species spiny, shorter than the inner, rarely suppressed.
comprises many ecotypes differing greatly in their Fertile spikelets with 1 basal sterile and 1 fertile
morphogenetic characters across accessions collected floret. Basal sterile floret barren and is with sig-
from the same locality or natural distribution (Arshad nificant palea. Lemma of lower sterile floret with
et al. 2007). Even differing number of chromosomes 5 veins. This species is well adapted to hot and dry
has been discovered between the ecotypes (Hignight tropical areas and it might be a paleotropical inva-
et al. 1991). sive weed. Impressions of its seeds were found in
3 Cenchrus 33

the archeological site of Tichit Chekka III (Maur- 5. Cenchrus ciliaris (L) Link, commonly known as
itania, west Africa) (Quattrocchi 2008). Buffelgrass, is native to tropical Africa and Asia
3. Cenchrus brownii Roem. and Schult is native to and is naturalized widely in subhumid and semi-
coastal plains of southeastern US, Central Amer- arid tropics and subtropics (GRIN Taxonomy
ica, and northern coast of southern America USDA; Clayton et al. 2006 onwards; Quattrocchi
(GRIN Taxonomy USDA; Quattrocchi 2008). 2008). It is also known with other synonyms
This species is distributed in tropical and temper- “Pennisetum ciliare” and “Pennisetum cen-
ate regions of Asia as well as Australia (Clayton chroids” (GRIN Taxonomy USDA). C. ciliaris is
et al. 2006 onwards). It is also commonly called as perennial species with tufted habit. Culms are
Green Sandbur or Slimbristle Sandbur. This is an 10–50 cm long at maturity and are ascending to
annual species with tufted herbaceous habit. erect, profusely branched, shrub-like in growth
Culms 25–100 cm long, ascending or erect. Root- habit. Leaf blades 3–25 cm long, 4–10 mm wide
ing at the decumbent base, culm nodes are some- and green or bluish green in color. Panicle spici-
times found in this species (Quattrocchi 2008). form, 2–14 cm long, bears spikelets in clusters of
Leaf blades 8–30 cm long, 4–11 mm wide. Pani- 1–4 spikelets. Involucral bristles are numerous
cle linear or oblong, 3–12 cm long. Deciduous and united at the base to form a shallow disc up
spikelets in clusters of 2–3 spikelets subtended to 0.5 mm above the rim. The bristles in C. ciliaris
by involucral bristles connate at the base up to are antrorsely scaberulous and not prickly. The
2.5–4 mm above to form a cup. Inner bristles inner bristles are flexuous, filiform in shape, and
subequal to outer bristles with the longest slightly exceed much in length than spikelets, one of them
emergent, flattened, rigid retrorsely scaberulous longer and stouter than the rest (DeLisle 1963).
and spinose. Fertile spikelets with one basal ster- Basal sterile florets are barren and are without
ile and one fertile floret. Basal sterile floret barren significant palea. Lemma of the sterile floret with
and is with significant palea. Lemma of lower 5 veins. This species grows mainly on sandy soils
sterile floret with 5 veins. This species grows in and alluvial plains, on shallow marginally fertile
sandy waste places, forest borders, savannah, on soils, along roadsides, rocky hillsides, hot and
beaches, and near the ocean (Quattrocchi 2008). dry areas, and denuded arid lands (Quattrocchi
4. Cenchrus calicalatus Cav., commonly known as 2008).
Hillside burrgrass or large burrgrass, is native 6. Cenchrus distichophyllus Griseb. is distributed in
to tropical Southeast Asia, Australia, New Zeal- Caribbean Islands (Clayton et al. 2006 onwards).
and, and Pacific Islands (GRIN Taxonomy USDA; It is a perennial species with distinct rootstock
Quattrocchi 2008). This perennial species grows habit. Culms are erect, 15–40 cm long. Leave
in clumps forming dense mats with its short sheaths are longer than adjacent internodes. Leaf
woody rhizome trailing or scrambling and rooting blade 2.6–3 cm long, 1.5 mm wide. Panicle
at the lower nodes. Culms erect or ascending, 2.5–4 cm long with axis bearing spikelet clusters
30–200 cm long, sometimes reaching up to of one spikelet. Involucral bristles connate at the
300 cm. Leaf blade flat and narrowly lanceolate, base to form a cup. Basal sterile florets barren and
5–56 cm long and 1–19 mm wide. Panicle spici- are with palea. Lemma of lower sterile floret ovate
form, loose and 4–23.5 cm long with axis bearing and is with 3 veins (Clayton et al. 2006 onwards).
deciduous spikelet clusters of 1–3 spikelets. Invo- 7. Cenchrus echinatus L., commonly known as
lucral bristles connate at the base to form a disc, Sandbur or hedhog sandbur, is found in tropical
which extends above for 0.5 mm above. Bristles and subtropical regions of the world. This is an
numerous, 8–12 in principle whorl. The inner annual species with herbaceous to subshrubby
bristles terete, rigid and spinose and are without habit. Culms are erect and 15–90 cm long. Leaf
grooves. Basal sterile florets male or barren, with blades 4–25 cm long, 3–10 mm wide. Panicle
or without significant palea. Lodicules are absent linear and interrupted. Axis bears spikelet clusters
in this species. This species is found growing on of 2–3 spikelets. Involucral bristles connate for
poor soils, on rocky coasts and on open sunny 2–5 mm at the base into a cup. The inner bristles
areas. flattened and outer whorl thinner than the inner
34 S. Goel et al.

and are tereted. Basal serile florets barren and are Panicle linear, 2–8.5 cm long, and often partially
with palea. Lemmas of the lower sterile floret are enclosed by the upper leaf sheath (Quattrocchi
with 5 veins. Seed heads of this species are com- 2008). Spikelets in clusters of 2–4 spikelets. Invo-
posed of spiny burrs. These spines cause problems lucral bristles connate for 2–7 mm into a cup
in cultivated lands and are very irritating to humans. below with clefts on two sides. Bristles rigid,
This species commonly grows on waste ground, flattened, and retrorsely scaberulous, with the lon-
coastal dunes near the ocean, open areas, roadsides, gest bristle scarcely emergent. Basal florets barren
and sandy or limestone soils (Quattrocchi 2008). and are with palea. Lemma of the lower sterile
8. Cenchrus elymoides is found distributed in Aus- floret is with 5–7 veins. This species grows mostly
tralia (Clayton et al. 2006 onwards). It is a peren- in disturbed areas, dry sites, dry sand in saline
nial species with robust habit. Culms are meadows, sand hills, road verges, and paddocks.
60–150 cm long. Leaf blades 14–34 cm long and This species often acts as indicator for the poor
3.3–10.2 mm wide. Panicle spiciform, dense and fertility of the soil (Quattrocchi 2008).
10.5–16 cm long. Spikelet clusters with 1–3 spi- 11. Cenchrus longispinus (Hackel) Fernald., com-
kelets in a cluster. Involucral bristles connate for monly known as long spine sandbur or spiny
0.5 mm below to form a disc at the base. The inner burrgrass, is native to Argentina, Mexico, and
bristles are longer than the outer bristles with one US, and is naturalized widely in Australia
conspicuously longer than the others, which (GRIN Taxonomy USDA). This is an annual or
expand at the base. Bristles are flexible, antrorsely short-lived perennial spiny species with coarse
scaberulous, and attenuate. Basal florets barren and tufted habit. Culms are ascending or some-
and are with palea. Lemma of the lower sterile times decumbent and ranges from 10 to 90 cm
floret is with 3–5 veins. in length. Leaf blades 6.3–18.7 cm long and
9. Cenchrus gracillimus Thunb. is commonly called 3–7.2 mm wide. Panicle spiciform and dense,
as Slender sandbur and is a native to southeastern 4.1–10.2 cm in length. Spikelets in clusters of
US and Caribbean Islands. It is a perennial spe- 2–3 spikelets. Involucral bristles connate to form
cies (Stieber and Wipff 2000; Quattrocchi 2008) a cup below. Bristles flattened, without grooves,
with dense tufted growth habit. Culms are rigid, retrorsely scaberulous, pubescent, and spi-
15–60 cm in length. Leaf blades 4.5–17 cm nose. Basal sterile florets barren and are with palea.
long, 1.1–3.3 wide. Panicle spiciform, loose and Lemma of the lower sterile floret is with 3–7 veins.
2–6.8 cm long. Axis bears spikelet clusters of This is an invasive weed species with low forage
1–3 spikelets. Involucral bristles connate at the value. It is a restricted grass species and is
base to form a cup. Bristles rigid, flattened, retro- included in aquatic or terrestrial noxious weed
rsely scaberulous, and emerged irregularly from and/or noxious-weed seed in U.S. This species
the body of the burr. Basal florets barren and grows in disturb soils, dry road side gravel, rail-
are with palea. Lemma of the lower sterile floret roads, and in waste grounds. This species is very
is with 3–5 veins. This weed species grows in similar in morphology and habit with Cenchrus
sandy soils of pinelands, wet prairies, and river echinatus, hence it is difficult to distinguish from
flats, behind the sandy dunes. each other (Quattrocchi 2008).
10. Cenchrus incertus M.A. Curtis., commonly 12. Cenchrus melanostachyus A. Camus. is found
known as Coastal Sandbur grass or Coastal San- distributed in eastern Africa at western coastline
bur, is native to northern and southern America of Indian ocean and Madagascar (Clayton et al.
(GRIN Taxonomy USDA). It is found distributed 2006 onwards). This is a perennial species with
to other parts of the world – in southwestern, short rhizomes. Culms are ascending, 40–60 cm
southeastern, and eastern Europe, in tropical in length, and swollen at the base. Leaf blades
Africa, and temperate Asian countries. It is an 25–35 cm long and 4–5 cm wide. Panicle spici-
annual or short lived perennial grass species with form and 3–8 cm long. Spikelets in clusters of
tufted, erect with shallow root system. The lower 3 spikelets. Involucral bristles connate for 1 mm
nodes of the prostrate stem give rise to secondary into a disc below. Bristles are flattened, rigid,
roots. Leaf blade 2–18 cm long and 2–6 mm wide. and antrorsely scaberulous and spiny. Basal sterile
3 Cenchrus 35

floret male and are with palea. Lemma with 5 and is with palea. Lemma with 3–7 veins. This
veins. species is very palatable and grows mostly along
13. Cenchrus mitis Anderss. is native to Africa, Tan- roadsides and in other waste places (Quattrocchi
zania, Mozambique, and Kenya (GRIN Taxon- 2008).
omy USDA; Clayton et al. 2006 onwards; 16. Cenchrus palmeri Vasey. is an annual species
Quattrocchi 2008). It is an annual species. Culms found growing in Mexico (Clayton et al. 2006
are 20–100 cm long and geniculately ascending or onwards). Culms are solitary, 9–35 cm long.
sometimes decumbent. Leaves flat 5–30 cm long Leaf blades 4.4–10 cm long and 3.8–6.8 mm
and 2–10 mm wide. Panicle spiciform, linear and wide. Panicle 2–4.3 cm long. Spikelets in clusters
4–18 cm long. Spikelets in clusters of 2 sometimes of 4–8 spikelets. Involucral bristles connate to form
3 spikelets. Involucres globose to ovoid, body of a cup below. Bristles numerous, terete, rigid, retro-
involucre pubescent but not grooved. Bristles con- rsely scaberulous, and spiny. Basal sterile floret
nate for 2–6 mm or 1/3–2/3 of their length to form male and is with palea. Lemma with 5–7 veins
a cup at the base. Bristles rigid, antrorsely scaber- (Clayton et al. 2006 onwards).
ulous, and spiny. Basal sterile floret male and is 17. Cenchrus pennisetiformis (Hochst. and Steud),
with palea. Lemma with 5–7 veins. This species commonly known as Cloncurry buffelgrass,
grows in bushland, coastal bushland, sandy soils, White or slender buffelgrass, is native to African
and abandoned cultivated lands (Quattrocchi countries including Kenya, Ethiopia, Somalia,
2008). Sudan, and Asian countries including India,
14. Cenchrus multiflorus J. Presl. is native to Pakistan, Yemen, and southern Iran (Quattrocchi
Mexico and Mesoamericana (Clayton et al. 2006 2008). It is naturalized in the Mediterranean reg-
onwards). It is an annual species with tufted habit. ion, Southeast Asia and northern Australia (GRIN
Culms 60–140 cm long. Leaf blades 16–25 cm Taxonomy USDA; Clayton et al. 2006 onwards).
long and 8–20 mm wide. Panicle linear, dense, It is a perennial species behaving occasionally as
and 7–18 cm long. Axis bears spikelets in clusters an annual. Culms are 10–40 cm long and are
of 2–4 in spikelets. Involucral bristles connate at ascending. Leaf blades linear, 2–20 cm long and
the base for 3 mm to form a cup below. Inner 2–5 mm wide. Panicle 2–7 cm long. Axis bears
bristles longer than outer. Bristles flattened and spikelets in clusters of 1–3. Involucral bristles
without grooves, rigid, antrorsely scaberulous, connate at the base for 1–2.5 mm into a cup
and attenuate. Basal sterile floret male and is below. Bristles are numerous, flexible, antrorsely
with palea. Lemma with 5–7 veins. One distinct scaberulous, and ciliate. Basal sterile florets are
character of this species is a reduced or scarcely with anthers and palea. Lemma with 5 veins.
developed leaf in the early development stage Morphologically it is similar to small-medium
of the plant life (cataphyll) (Clayton et al. 2006 type C. ciliaris. It is differentiated from C. ciliaris
onwards). by its extent of fusion of the involucral bristles. In
15. Cenchrus myosuroides Kunth. is commonly called C. pennisetiformis, the fusion of inner bristles
as Big Sandbur and is native to South-central US extends more above the rim of the basal disc.
and northern South American countries and Apart from it, plants of C. pennisetiformis are of
Caribbean Islands (GRIN Taxonomy USDA; smaller stature, usually annual, and favor subde-
Clayton et al. 2006 onwards; Quattrocchi 2008). sert habitats (Quattrocchi 2008).
This is a perennial tall bunchgrass species. Culms 18. Cenchrus pilosus Kunth. It is found distributed in
are 50–200 cm long, tough, stout, and erect with Mexico, northern South America, and southwest-
decumbent base. Leaf blades 12–38 cm long and ern America (Clayton et al. 2006 onwards). This is
4–13 mm wide. Panicle linear 6.5–23 cm long and an annual species with dense and tufted habit.
dense. Spikelets in clusters of 1(-3) spikelets. Bur Culms 30–60 cm in length. Leaf blades 6–30 cm
consists of single whorls of bristles. Bristles long and 4–11 mm wide. Panicle spiciform, dense,
united at the base to form a disc below. Bristles and 2–13 cm long. Spikelets in clusters of 2–3
as long as the spikelets, flattened, rigid, retrorsely spikelets. Involucral bristles connate for 2–4 mm
scaberulous, and spiny. Basal sterile floret male at the base to form a cup. Bristles flattened,
36 S. Goel et al.

flexible, smooth, and attenuate. Basal sterile flor- terete, retrorsely scaberulous, and spiny. This spe-
ets barren and with palea. Lemma with 3–7 veins. cies can be distinguished from others by its stiff
19. Cenchrus platyacanthus. Annual species dis- inner spiny bristles and vestigial outer bristles.
tributed in western South America (Clayton 23. Cenchrus somalensis Clayton. is a perennial spe-
et al. 2006 onwards). Clumps weak, either erect cies native to Somalia (Clayton et al. 2006
or ascending, 20–90 cm in length. Leaf blades onwards). Plants densely tufted in habit. Culms
elongate, 4–24 cm long and 2–8 mm wide. Panicle erect and 25–45 cm long. Leaf blades narrow and
spiciform, dense, and 3.8–8.6 cm long. Spikelet convolute, 5–15 cm long and 1 mm wide. Panicle
clusters with single spikelet. Involucral bristles linear to oblong, 2.5–6 cm long. Axis bears spike-
connate at the base for 2.8–6 mm into a cup lets in clusters of 1–2. Involucral bristles connate
below. Bristles terete, rigid, retrorsely scaberu- for 2–4 mm into a cup below. Bristles flattened
lous, and spiny. Basal sterile florets barren and and flexible, antrorsely scaberulous, and ciliate.
with palea. Lemma with 3 veins. Basal sterile florets with anthers and palea.
20. Cenchrus prieurii (Kunth) Maire. This species is Lemma with 5 veins. This species grows mainly
distributed in India and tropical African region under shade of trees and among the bushes.
(Clayton et al. 2006 onwards). It is an annual or 24. Cenchrus spinifex Cav. is commonly known as
short species like perennial tufted species, with Coastal sandbur. This species is found distributed
erect to ascending habit. Culms 30–75 cm long. in US and South America (Clayton et al. 2006
Leaf blades 8–30 cm long and 5–10 mm wide. onwards; Quattrocchi 2008). Plants are annual
Panicle 6–14 long. Spikelets in clusters of 2. or short-lived perennial, culms 30–100 cm long,
Involucral bristles connate at the base for 1 mm geniculately ascending, or decumbent. Panicle
into a disc below. Bristles numerous and are 3–5 cm long. Spikelets in clusters of 2–4 and are
flattened, grooved on the face, rigid, antrorsely with sharp spines. Bristles connate at the base for
scaberulous, and ciliate. Basal sterile florets bar- 1.5–2.5 mm into a cup. Bristles numerous, rigid,
ren and without significant palea. Lemma with 3 flattened, retrorsely scaberulous, and spiny. Basal
veins. sterile florets are with anthers or barren. Florets
21. Cenchrus robustus R.D. Webster is found are with palea or sometimes absent. Lemma with
distributed in Australia (Clayton et al. 2006 5–7 veins. It is a noxious weed growing on sandy
onwards). This is a perennial species with short soil in both open areas and thin wood, along road-
rhizome. Distinct cataphyll. Culms are 40–100 cm sides, and waste fields and places. It is very simi-
long and erect. Leaf blades 6–22 cm long and lar in morphology with C. longispinus. But it has
3–7 mm wide. Panicle 4–8 cm long. Axis bears shorter spikelets, fewer bristles, wider inner bris-
spikelets in clusters of 1–2 spikelets. Bristles con- tles, and flattened outer bristles (Stieber and Wipff
nate for 0.5 mm below to form a disc. Bristles 2000).
numerous, rigid, terete, retrorsely scaberulous, 25. Cenchrus tribuloides L., commonly known as
and spiny. Basal sterile florets barren and are American burrgrass, is native to northeastern and
with palea. Lemma with 5 veins. southeastern US, Caribbean Islands, and Brazil
22. Cenchrus setiger Vahl. is commonly known as (Clayton et al. 2006 onwards; Quattrocchi 2008).
Birdwood grass and is native to East and North- This is an annual species with sharp spines, which
east Africa, Northwest India, Arabia, Yemen, are painful to touch. Culms are 10–70 cm long,
Australia, and Brazil (GRIN Taxonomy USDA; erect or ascending, branching and rooting at the
Quattrocchi 2008). It is a perennial species, which lower nodes. Leaf blades 2–14 cm long and
grows vigorously and rapidly into bunch, more or 3–14 mm wide and gray green in color. Spikelets
less erect, forming clumps from a bulbous base. hard and prickly, infrequent, not in clusters, articu-
Culms are 40–100 cm long. Leaf blades 6–22 cm lated, solitarily on the panicle axis, and densely
long and 3–7 mm wide. Panicle spiciform, 4–8 cm pubescent. Bristles connate into a cup below. Spi-
long. Axis bears spikelets in clusters of 1–3 spike- kelets numerous, flattened, rigid, retrorsely scaber-
lets. Involucral bristles connate below for 1–3 mm ulous, ciliate, and spinose. This is a weed species,
into a cup at the base. Bristles numerous, rigid, which is a useful stabilizer of sand dunes and grows
3 Cenchrus 37

in moist and sandy dunes, on coastal sands, open quadrivalents, but there is no significant difference in
dunes, tropical salt marshes, and open grasslands to the size of the paired chromosomes (Read and Bashaw
dense shrub-cultivated fields. The young plants are 1969). This relationship between these two species is
used as fodder in dry areas. well supported by isozyme and DNA-based marker
analyses (Chandra and Dubey 2007, 2008; Kellogg
These species of Cenchrus are mainly distributed in
et al. 2009).
the sub-Saharan central and eastern Africa, arid and
Cenchrus is a much variable genus. It has base
semi-arid south-central to southwestern US to north-
chromosome numbers x ¼ 9, 10, 15, 16, 17, 18 (Fisher
ern Mexico on the west, temperate Asia (Middle East
et al. 1954; DeLisle 1963; Vij and Chaudhary 1981;
countries), tropical Asia (India, Pakistan, Indonesia)
Morrone et al. 2006). Majority of the species possess
on the east and semi-arid regions of Australia
chromosome numbers, which are multiple of the orig-
(Queensland, northern South Wales). Eastern Africa
inal basic number, i.e., are mainly polyploids and also
is considered as the center of diversity for tropical
show high degree of aneuploidy. Somatic (2n) chro-
grasses with over 90% of the cultivated forage grasses
mosome numbers of different species ranges from 18
including Cenchrus originated in this region (Suttie
to 70 (Table 3.1). The available cytological data sug-
et al. 2005). Out of the 25 species, two species are
gested that majority of the species are stable cytologi-
reported in Europe, 11 in Africa, eight in temperate
cally with the exception of a few species. Greater
Asia, eight in tropical Asia, 11 in Australia, seven
instability in chromosome numbers is observed in
species in North America, and 13 species in South
C. ciliaris, C. Browni, C. biflorus, Cenchrus mysur-
America (Clayton et al. 2006 onwards). Out of the 25
oides, and Cenchrus setiger (Vij and Chaudhary
species, 13 species are perennial, 10 are annual, and
1981). Lagging of chromosomes during first meiotic
two behave as both annual and perennial in habit.
anaphase due to precocious meiotic division is a com-
Cenchrus grows mainly on sandy soils and alluvial
mon phenomenon in cytotypes with multiple dip-
plains, on shallow marginally fertile soils, along road-
loid chromosome numbers (Fisher et al. 1954). Such
sides, rocky hillsides, hot and dry areas, and denuded
arid lands, growing when soil moisture is available.

Table 3.1 Chromosome numbers of different species of the

genus Cenchrus
Species Chromosome numbers
3.3 Cytology and Cytogenetics C. biflorus 30, 32, 34, 36
C. brownie 34, 36, 66ca
The most widespread species of Cenchrus, C. ciliaris, C. cathcarticus 34
shares the tertiary gene pool of pearl millet (Pennise- C. ciliaris 18, 32, 34, 36, 36+0-2Bs,
40, 44, 50, 52, 54, 56
tum glaucum) along with other apomictic species of C. echinatus 34, 68
Pennisetum (Jessup et al. 2000; Martel et al. 2004). C. glaucus 36, 38
This grass species has a small genome size of ~ C. gracillimus 34
3,200 MB with 1.33 pg of DNA per haploid genome, C. incertus 32, 34
which is low for a polyploid grass species (Jessup et al. C. inflexus 34
2000; Roche et al. 2002). However, limited studies C. longispinus 34
have been done so far to identify other species belong- C. mitis 34
C. multiflorum 34
ing to the primary, secondary, and tertiary gene pool of
C. mysuroides 54, 68 ca 70
C. ciliaris. C. setiger may be in the secondary gene C. palmeri 34
pool of this grass species. Cytological examination of C. parviceps 34
meiotic chromosome behavior in tetraploid (4x ¼ 36) C. pauciflorus 34, 36
accessions of both species show similar pairing pat- C. pennisetiformis 35, 42, 54
terns (normal disjunction) during metaphase I (Fisher C. pilosus 34
et al. 1954). These two species can cross with each C. prieuri 34
C. setiger 34, 36, 37, 54
other, producing fertile hybrids. Meiotic chromosome
C. tribuloides 34
spreads of the hybrids show more bivalents and fewer
38 S. Goel et al.

variation in chromosome number in different species

of Cenchrus may have aroused due to the natural
interspecific or intergenic hybridizations, which
occurs frequently in grasses. Majority of these spe-
cies showing cytological instability are facultative
apomicts with the exception of C. ciliaris that repro-
duces through sexual fertilization also. Therefore,

Strange
ASGR
there is more chance of preserving the new chromo- 1 2 3 4 5 6 7 8 9
some combinations. On the other hand, an increase in
chromosome number may induce meiotic abnormal-
ities resulting in sterility and subsequently may have Fig. 3.1 Quantitative ideogram of Mitotic Cenchrus ciliaris
chromosome. Red, green, and yellow circles indicate C101,
evolved apomictic mode of reproduction to escape
18S rDNA, and 5S rDNA probes, respectively. Source:
the sterility. These variations in chromosome number Akiyama et al. 2005
or genome size might have contributed to the varia-
tions in certain morphological features, develop-
mental variations, and productive capacity (Mnif Many authors had reported Cenchrus setiger with 2n
et al. 2005). chromosome number of 34, 36, 37, and 54 (Fisher et al.
C. ciliaris alone shows variable ploidy levels with 1954; Vij and Chaudhary 1981; Ahsan et al. 1994).
2n ¼ 18, 27, 32, 36, 43, 45, 48, 54 (Fisher et al. 1954; However, 2n ¼ 34 seems to be most widespread. The
Snyder et al. 1955; Bashaw and Hignight 1990; Hig- chromosomes are of metacentric and submetacentric
night et al. 1991; Mnif et al. 2005), but tetraploid with types with length ranging from 3.61 to 1.00 m, and pollen
chromosome no. 36 (2n ¼ 4x ¼ 36) is presumably the mother cell meiosis shows variable number of quadriva-
most widespread ecotype (Hignight et al. 1991). It lents (1–5) and bivalents (Vij and Chaudhary 1981).
behaves like a segmental allotetraploid, showing at The somatic chromosome complement of Cenchrus
the average of two to four quadrivalents and 10–14 biflorous ranges from 2n ¼ 30, 32, 34, 36 with the base
bivalents during metaphase I (Bashaw 1962; Hignight chromosome numbers x ¼ 16, 17, 18 (Vij and Chaudh-
et al. 1991). This segmental behavior of the buffel- ary 1981; Ahsan et al. 1994). The chromosomes are
grass genome is verified genetically by Jessup et al. predominated by submetacentric types with length
(2003). Analysis of the polymorphic probes in genetic ranging from 2.73–1.07 m. The chromosomes can be
linkage maps reveal repulsion phase associations of 26 grouped into two types with relatively larger group of
linkage groups indicating preferential pairing of link- size range of 2.72–1.74 m and smaller group of
age groups. The paternal and maternal chromosomes 1.50–1.07 m. The meiotic behavior of the chromosomes
behave differentially from each other, with the mater- differs with different cytotypes showing difference in
nal parent having higher frequency of crossing over the number of bivalents of the smaller group of chro-
against the paternal parent (Jessup et al. 2003). The mosomes. However, cytotype with n ¼ 17, which
chromosomes are very similar in morphology as well shows more stability during pollen meiosis, seems to
as in size and difficult to distinguish. Akiyama et al. be predominant (Vij and Chaudhary 1981).
(2005) constructed a quantitative ideogram of tetra- Majority of the species of Cenchrus reproduce apo-
ploid Buffelgrass (C. ciliaris) using fluorescent in situ mictically, but low level of sexuality has been reported
hybridization (FISH) with 5S rDNA, 18s rDNA, and in some species. The reduced female gametophyte is
bacterial artificial chromosome (BAC) clones contain- of polygonum type with two synergids, an egg cell,
ing apomixis-linked markers (C101) as probes to two polar nuclei in the central cell, and three antipo-
DAPI stained spreads (Fig. 3.1). Thirty-six chromo- dals toward the chalazal end (Bashaw 1962). Apospor-
somes were classified into nine morphologically ous unreduced female gametophytes can be identified
similar chromosome types with two additional chro- cytologically by the absence of antipodals in mature
mosomes with no similarity with other chromosomes. embryo sac or presence of multiple embryo sacs in
All chromosomes are of metacentric and submetacen- cleared ovules. Ovules can be effectively cleared in
tric types with arm ratio ranging from 1.1 to 1.7 methyl salicylate after fixing for 24 h in FAA (forma-
(Akiyama et al. 2005). lin: acetic acid: ethyl alcohol) and storing in 70% ethyl
3 Cenchrus 39

alcohol (Young et al. 1979). In both sexual and apo- differ in the base chromosome number x ¼ 9 and
mictic plants, the differentiation of the archesporial x ¼ 10. Cenchrus belongs to the x ¼ 9 clade along
cell and subsequent reductional division to form a with the type genera of the tribe, Panicum, and other
linear tetrad is similar, but in apomictic ovules, all species-rich genus Pennisetum and Setaria. Most of
members of linear tetrad degenerate, although rarely, the relative genera are important forage grasses except
and the chalazal megaspore develops into 2-nucleate one species P. glaucum (Pearl Millet), which is
stage (Bashaw 1962). In the meantime, the unreduced regarded as a major cereal crop of the semiarid tropics.
nucellar cells undergo two successive equational divi- This clade is known informally as “Bristle clade”
sion and enlarge rapidly to form multiple 4-nucleate (Doust and Kellogg 2002) because of the presence of
unreduced embryo sacs; one egg, two synergids, and sterile inflorescence bristle, which is unique among
one diploid polar nuclei and occupy a large portion of grasses. Based on morphological similarity, Cenchrus
the ovule. Despite its unreduced egg cell reproduction, was earlier placed under genus Pennisetum (Jauhar
fertilization of the central cell through cross pollina- 1981). Different species have been described in both
tion is needed to develop endosperm to nourish the genera at different times, and several authors also
developing embryo and hence to ensure seed set (Hig- indicated that the two genera may be merged into a
night et al. 1991). Presence of such dual mode of single genus (DeLisle 1963; Correlld and Johnston
reproduction may be reason for the high levels 1970). Presently, two genera are still maintained as
of polyploidy and aneuploidy in different collections separate genus (Doust and Kellogg 2002; Kellogg
of Cenchrus (Fisher et al. 1954). An increase in chro- et al. 2009). Both of them show very similar pattern
mosome number in aneuploid may induce meiotic of early inflorescence development, but the difference
abnormalities resulting in sterility and subsequently appears late in development. In many species of Cen-
may have evolved apomictic mode of reproduction to chrus, except C. ciliaris and C. myosuroides, the lat-
escape the sterility. These variations in chromosome eral branches or inflorescence bristles become
number or genome size might have contributed to the broadened laterally, while those of Pennisetum remain
variations in certain morphological features, develop- more or less terete (Doust and Kellogg 2002). Their
mental variations, and productive capacity (Mnif et al. separation into two separate genera is also supported
2005). by the extent of fusion of the inflorescence bristles. In
Cenchrus, the lateral edges fuse to variable extent
while they are free in Pennisetum and elongate more
just before anthesis (Doust and Kellogg 2002). Phylo-
3.4 Phylogenetic Relationship genetic analysis shows that, within the paraphyletic
Pennisetum, Cenchrus forms a derived monophyletic
The subfamily Panicoideae of the large family Poa- subclade (Giussani et al. 2001; Doust and Kellogg
ceae includes approximately 208 genera and ~3,300 2002; Martel et al. 2004; Doust et al. 2007; Kellogg
species. This large subfamily shows monophyly, et al. 2009; Fig. 3.2).
which is well supported morphologically by the pres- The genus Cenchrus consists of both perennial and
ence of bifloral spikelets (lower sterile or staminate annual species. The knotted 1 marker gene analysis of
flower and upper fertile spikelets) and anatomically by the members of panicoideae including six species of
the presence of simple starch grains in caryopsis (Alis- the Cenchrus reveals that three species of Cenchrus,
cioni et al. 2003). The molecular level analysis of C. ciliaris, C. setiger, and C. echinatus, are very
nuclear and chloroplast genes also supported the closely related, forming a close monophyletic clade,
monophyly of this large subfamily (Giussani et al. and other three species C. calycalatus, C. myosuroids,
2001; Doust and Kellogg 2002; Doust et al. 2007; and C. pilosus form a separate group (Doust et al.
Kellogg et al. 2009). Panicoideae comprises two 2007). Another study with chloroplast ndhF marker
tribes: Paniceae and Andropogoneae. The tribe Pani- shows closer relationship of C. echinatus with C. caly-
ceae is a highly diverse and species-rich assemblage of calatus, C. myosuroids, and C. pilosus group and also
>200 species comprising one-fifth of the total species reconfirms the close relationship between C. ciliaris
of grass (Duvall et al. 2001) and about 60% of pani- and C. setiger (Kellogg et al. 2009). The close
coideae. This tribe is paraphyletic with two clades that relationship among these three species, C. ciliaris,
40 S. Goel et al.

Fig. 3.2 ML phylogram for full kn1 dataset. Support values above branches are ML bootstrap/MP bootstrap; below branches are
Bayesian posterior probabilities

C. setiger and C. pennisetiformis, may be because of species, C. ciliaris and C. setiger are morphologically
chance hybridization among the three species (Doust very similar and a few workers have placed C. setiger
et al. 2007). Although separated into two different as a variety of C. ciliaris. Both apomictic and sexual
3 Cenchrus 41

forms are reported in C. ciliaris, but sexual reproduc- competition with other grasses, thus reducing its pro-
tion has not been reported in C. setiger (Read and ductivity (Cox et al. 1988). Growth initiated in the
Bashaw 1969). These two species can also cross with spring season continues to grow during the hot sum-
each other, producing fertile hybrids. The growth habit mer temperature and produces new growth with sea-
of the hybrid is intermediate of the two parents: rang- sonal rain.
ing from upright as C. ciliaris and procumbent as Many species of Cenchrus are also potential inva-
C. setiger. The seed set under open-pollination reach sive weeds, which displace native species. They mod-
up to 99% and up to 84% under self-pollination, which ified to a good extent natural plant communities in
are comparable to the progenies of sexual buffelgrass. California, Texas, and Hawaii in US, Sonora in Mex-
The closer relationship between C. ciliaris and ico, and in Australia (Martin et al. 1995; Tix 2000;
C. setiger is also supported by isozyme, sequence Tunyalee 2000; Chen and Kuoh 2004; Franklin et al.
tagged site (STS) and random amplified polymorphic 2006). Spines on the surface and the seeds of the
DNA (RAPD) markers (Chandra and Dubey 2007, C. echinatus, C. tribuloides, and C. biflorus are very
2008). It seems that the variation between species is irritating, and one cannot walk bare footed in the areas
affected by the type of habit of the species. The RAPD, naturalized by these plant (Quattrocchi 2008). Like
isozyme and STS markers clearly separated the annual other invasive grasses, Cenchrus is very efficient in
forms from perennial species with the exception of water usage. Its rhizomatous root draws soil moisture
C. mysuroides, which is a perennial species grouping rapidly as compared to other plants. C. ciliaris also
with annual species. Two perennial species C. ciliaris exudes some allelopathic chemicals, which inhibit the
and C. setiger are shown to be most closely related and germination of other plants (Tunyalee 2000), and as a
they are most distant from the strictly annual species result, C. ciliaris once established in an area rapidly
C. prieurri. This indicates that C. mysuroides may be modifies the physiochemical properties of natural
acting as a bridging species between annual and peren- habitat and makes the soil unfit for others. Following
nial forms (Chandra and Dubey 2008). invasion, this weed species forms dense thicklets.
The dense growth habit of this rhizomatous grass
has the potential to carry the fire rapidly across
broad areas and change the fire regime of the eco-
3.5 Ecological Behavior system, where fire does not normally play important
role. Like most others, this grass penetrates the soil,
The available distribution data suggested that many of and hence, fire has minimal negative effect on this
the species of Cenchrus originated from tropical and grass and it recovers very quickly. Recurrent fire
subtropical India and Africa, where humid warm sea- in the area maintains C. ciliaris populations and the
sonal bimodial (distributed in two summer growing ecological result is the conversion of native scrub
season) rainfall predominates. Mean minimum and communities to African type Savannah with reduced
maximum temperatures of the region vary from 21 to native biological diversity (Tu et al. 2002). It also
24 C and 31 to 36 C, respectively (Cox et al. 1988). It imposes economic cost through need to manage fire
is a xerophytic C4 grass, which grows in bunches and and to protect biodiversity and also the infrastructural
more often with creeping rhizomes. It spreads predom- establishments.
inantly in areas where soil is of coarse texture sandy
loam type with marginal productivity on clay soils.
Poor soil aeration and limited availability of phospho-
3.6 Traits of Agronomic Importance
rous limits the survival and productivity of this grass
in clay soils (Ibarra-F et al. 1995). These grass species and Scope for Domestication
can be established under minimal level of precipitation
ranging from as low as 100 mm to 1,000 mm with a Since new to cultivation, C. ciliaris has not yet gone
few exceptions to more than 2,500 mm annual aver- through bottleneck of domestication, and extreme
age. However, seed germination and establishment of variability exists in wild at intraspecific level also.
the plants during extended wet periods are affected by During 1880–1930, frequent droughts in Africa,
allelopathy, infection by insects and pathogens, and North America, and Australia resulted in reduction of
42 S. Goel et al.

livestock numbers. Many botanists, even the military production of forage and in some cases destroys stands
personnel, were sent to many parts of the world in (Cook et al. 2005).
search for grass species, which produced abundant In addition to C. ciliaris, other two closely related
forage in the minimum level of precipitation. Many species, C. pennisetiformis and C. setiger (birdwood
seeds from harsh climate were collected that under- grass), are also potential grass for pasture plantation,
gone many trials. From these thousands of trials, a few but they are less common than C. ciliaris (Weed
grass species were collected for their ease of establish- Management 2008). C. pennisetiformis is adapted to
ment and forage production in harsh environments arid conditions and slightly alkaline soils. This grass
(Cox et al. 1988) and C. ciliaris was one of them. species shows more tolerance to drought and dry con-
This grass species occupies the major component ditions and survives seasonal flooding. It grows com-
of Dichanthium–Cenchrus–Lasiurus grass cover of monly on deep sandy soils of semi-desert habitats
India. It was one of the most commercially significant and is very useful for the stabilization of sand dunes.
sown pasture grasses in Australia during 1960s and C. setiger can adapt to more wide range of soil types as
also planted as a valued pasture grass in Texas. Alto- compared to C. ciliaris; however, it prefers light tex-
gether, 34 different cultivars with different eco-physi- tured sandy soils. C. Setiger shows more resistance to
ological characteristics such as high forage quality and heat and frost compared to C. ciliaris; however, it has
high yield have been released for pasture improvement lower yield as compared to C. ciliaris and C. penni-
in US, Australia, Brazil, South Africa, Mexico, India, setiformis. This species is well adapted to arid and
and tropical African countries (Cook et al. 2005). This semiarid regions with a long dry season and is very
grass can grow in minimum amount of water as com- effective in stabilizing the moving sand. These agro-
pared to other grass species (Osman et al. 2008), nomic traits if incorporated in C. ciliaris can yield rich
suggesting that the potential of buffelgrass could be dividends. Importance of wild species in improvement
harnessed in dry Arabian Peninsula to develop grass- of C. ciliaris can be realized with these two species
lands and reclamation of degraded rangelands with (C. pennisetiformis and C. setiger), although more
minimum irrigation. studies are warranted in wild species of the genus
Various traits are known to exist in wild species of Cenchrus.
genus Cenchrus, although due to lack of enough stud-
ies, full potential of these species in improvement of
cultivated C. ciliaris has not been realized. C. ciliaris
3.8 Application of Molecular Techniques
although adapted to arid zone conditions, is suscepti-
ble to frost and various biotic stresses. More resistance in Crop Improvement
to frost has been reported in C. setiger, but studies are
required to establish the source of resistance to biotic Compared to other members of the family Poaceae,
stresses at intra and interspecific level. limited studies have been done so far to assess the
diversity at genetic level in tropical grasses such as
Cenchrus. RAPD, AFLP (amplified fragment length
polymorphism), ISSR (inter-simple sequence repeat),
3.7 Breeding and Crop Improvement and STS markers have been used to reveal genetic
distance at interspecific and intraspecific level (Gustine
C. ciliaris is a perennial warm season grass species et al. 1996, Griffa et al. 2006; Chandra and Dubey
quite tolerant to drought conditions and with good 2007, 2008; Gutierrez-Ozuna et al. 2009). RAPD anal-
forage characteristics. On the contrary, buffelgrass ysis with 106 primers in 10 populations of C. ciliaris
establishment in other regions is limited due to lack consisting of a sexual line, an apomictic line, and eight
of winter hardiness and salt tolerance in this plant. It progenies of the cross between two lines generated
can adapt to only a narrow range of soil types and is 569 distinct bands, out of which 495 bands showed
susceptible to several diseases (Jessup et al. 2000), the differential expression among the lines (Gustine et al.
most destructive being a rampant leaf spot disease 1996). In another RAPD study (Chandra and Dubey
(Rust) caused by the fungus, Magnaporte grisea (ana- 2008), a higher degree of polymorphism was recorded
morph Pyricularia grisea), which reduces quality and among eight species of Cenchrus collected from
3 Cenchrus 43

different regions of the world. One hundred and Among the 17 Stylosanthes STS primers tested, 14
eighty-seven decamer primers generated a total of STS primers amplified 195 scorable bands, out of
1,296 scorable bands, out of which 1,204 (92.9%) which 162 showed polymorphism between five spe-
showed polymorphism among the eight species cies, indicating high amplification success of 82%.
(Chandra and Dubey 2008). This result is the indica- These bands show reasonable genetic distance ranging
tion of a broad genetic base in the genus. from 0.51 to 0.79 between species. This result pro-
Gutierrez-Ozuna et al. (2009) investigated the vides an opportunity to use the primer sets in decipher-
genetic diversity among pasture and colonized road- ing the diversity at both inter and intraspecific level.
side populations of northwestern part of Mexico using The authors also demonstrated that the isoenzyme
ISSR markers. The genotypic parameters analysis variation between different species of Cenchrus is
demonstrated that populations have low level of gen- affected by the breeding system of the species. The
etic diversity between the roadside populations and perennial species show more polymorphism than the
pasture populations. These high frequencies indicate annual species. Out of nine isoenzymes tested, ester-
that the genotype of the region may be maintained by ase produced more variable loci showing three and
agamospermy or probably due to founders effect, or two polymorphic loci in perennial and annual species,
the populations sampled may represent the original respectively.
introduced genotypes (Gutierrez-Ozuna et al. 2009). Chandra and Dubey (2008) also assessed the differ-
However, low level of polymorphism shown in vege- ential response of eight species of Cenchrus using
tative characters and at the DNA level may be a result drought tolerant physiochemical parameters such as
of occasional sexual reproduction or chance fertili- malondialdehyde (MDA), proline content, specific
zation of unreduced gametes (Hignight et al. 1991). leaf area, and carbon isotope discrimination (CID).
Somatic variation can also be responsible for the low Generally, the level of MDA and proline accumulation
level variation. increases in response to water stress in wheat (Zhang
Griffa et al. (2006) employed AFLP markers in and Kirkham 1994), indicating a higher peroxidation
assessing genetic diversity in buffelgrass. According of lipids. The same result of MDA accumulation was
to the authors, it was the first report of utilization of the observed in most of the eight species subjected to
technique in buffelgrass. Their analysis using three water stress, with the exception of C. ciliaris, which
primer combinations produced 152 scorable bands shows significant drop in MDA accumulation. There is
among six apomictic cultivars, one sexual line and also significant increase in proline content with the
15 putative hybrids of the cross between apomictic exception of C. prieuri, which is an annual species.
male cultivar and sexual female lines. These markers The percent increase in the MDA and proline accumu-
segregate most of the putative F1 hybrids towards the lation was maximum in C. pennisetiformis, thus indi-
female parent with the exception of two putative F1 cating higher tolerance of this species to water stress
hybrids, which cluster neither with their parents nor conditions.
with the other putative hybrids. The closeness of the
hybrids to the female parent indicates that these
hybrids may have originated through self-pollination
of a sexual line. The two putative hybrids, which form 3.9 Genetic Map
a separate cluster, may represent the true hybrids. The
hybrid nature of the two F1 putative hybrids was con- Genetic map has been reported in C. ciliaris, but no
firmed cytologically with observation of aposporous reports could be found in other species. A full genetic
embryo sacs and with the amplification of sequence- map, which covers approximately 70–80% of the
characterized amplified region (SCAR) marker “ugt genome, has been reported in buffelgrass (Jessup et al.
197”. 2003; Fig. 3.3). A total of 100 cDNA probes consisting
Chandra and Dubey (2007) demonstrated that the of clones from a cDNA library from the pistils of an
STS markers developed on the basis of functional gene apomictic buffelgrass and other cDNA clones, which
sequences of other non-related grass species can be show differential expression between sexual and apo-
successfully employed to analyze the level of genetic mictic ovaries in a previous study by Vielle-Calzada
diversity between eight different species of Cenchrus. et al. (1996), and 360 heterologous polymorphic
44 S. Goel et al.

P2C03d MS48x Pca3b

P6C07c CSU706b SB1541 P10H05c P9H08h
18.7 11.3 23.6
16.1 16.4 23.0 22.2
3.3 P12C02d
P10F11d P2E04a 27.4
8.8 P3C80a 7.7 P9F03c P11B11f Pcs1c
P11C07a P7G01d CSU742f
20.5 15.9
16.0 19.4 P9C09a 12.9
25.2 P10C12b 8.7 P6B07g
P9H05f 20.7 P9B07b
RZ478i P7A08g 19.0 P1C05d 20.7
5.4
10.3 P8B05c 19.6 P6B06a C746b 21.2
1.2 16.4 P7A01e M651d
P6B06c P1H06e 14.6
8.3 17.6
P9G08c P9G04a P10H05b P7H04h 19.4 P8H10c
6.4 P7A01f 28.0
P10A04b 12.5 P6G04c P10H08h
P9B07d
20.7 P1C09g 25.3
31.1 16.6 P9B07e
P1H05j 26.2
C356d 36.4
15.6 P10C12a 19.2
27.8 P2C06n P10D08f
P3A04k P7G03e 14.0 P9C09c
P8A05b 13.5 P7A01d 8.0
18.1 P12C03d M848d
23.3 3.5 Pcs1d 4.0
M891b 19.4 P4F04a P7F01h P8H10b
6.5 3.1 P9B07c 14.3 4.9
P10H05g C356g P10H05e P9A09g
3.5 14.6
13.2 3.4 P3H07k P5H09l P3C08b P1C12f 14.7
P3B10c P8F01a 26.7 Pc08b
P9D10i 21.9 10.7
10.7 16.8 CSU523c
12.1 P8A02e 1.7 P10C12c
P7C10o CDSR94k CSU742e
1.0 P10H05d 9.2 P9D10c 26.4 15.9
12.8 1b 1.2 P12C05g P10E03e
CSU410i
CSU410h 116.2 cM 1.6
P8C08i 1b 7b, LOD-4.90 11.7 P2E06d
20.8 P5G09a
12.9 117.5 cM P1D01f
25.5 CSU360i P9B09a P8F01c 13.6
12.5 19.7 17.5 P9B09c
P3H07d
23.0 5.4 P10F10c
13.9 P1C12b P9D09b
P4A06d P12D07a 2.1 22.5
4.0 CDSR94g P7F01e
6.3 CSU5391 P7C03c P10C08c 20.8 15.0
4.7 P7H05e 18.0
P3B106 P5F11h P7D07a P5G09c
P1C12c
16.5 23.4 2a 7.5 P10E03a 2b
P7G03a 228.8 cM P3A06n 190.9 cM
2a
17.7 P4A06b 171.2 cM
10.9 24.7
CDSR94J
6.0 P7G03c
27.7 P8A02d P10D03a
8.1 2b
P10H05f
CDSR94i 192.2 cM
1a 18.5
293.0 cM
CSU527e
12.4
P3C08c
1a
241.9 cM

CSU455d P3E08i P3C01g Pca2d P7D12g P6A08b P9A10i P7C04h

12.7 13.4 8.7 11.6 11.1 12.5
13.5
26.7 3.8 P7D12c P9D04b P5D01m P8D06e
P9D04d 8.9 3.0 P1B11d CSU351aS
P1D01i 5.4 4.6 PSH07e
2.2 P1C03h 14.3 P3C01a P12E06c 6.9
P8A05g 12.7 7.3 P1C03e 2.7 P5H07d 16.8
P6A08e 5.5 P6A08c 7.6 P9A10e
P9D10f 23.5 PSF11a 2.7 P1D04a
16.8 P10C08a P11D10e
31.9 5.7
P1C03d 13.7 P5H02e
24.7 P1B11b 13.3 3.1
P3G08g
P10B12c
8.9 P8D06c P1D03a
2.0 P9H05g P9A10c 24.1 7.2
P9H05e 11.0 P9F06j 1.5 P7C04f P4G04h 27.9
24.8 P6H03a P10H12d 5.5 P9F06h 3.3
P10G12b
SB289c
14.7 5.0 P10F06e 6.0 34.9
17.2 C250d 5a, LOD = 3.17 15.5 C250b
P1A09b
CSU360l P7H07k 9.4 P1D03h 5a, LOD = 4.20
1.4 P11D10c 16.9 5b
2.9 P2B11b P3G10a 5a, LOD = 3.18 12.3
30.7 P9A06d 34.2 3.6 C250c 57.2 cM
5.2 7.8 P10C05d P1D04d P1D04i P10H12c
P9B06d 4.1 14.0 4b, LOD–4.20
5.0 P7B12f P1D03f P1D03g 4b
CDSR7f
P1D01h SB289d 8.8 9.9 74.9 cM P5D10g P3F02d CSU460j P7C04g
P10C05a 1.9 P4B06c P4G04f CSU479g P1D03c
14.1 M737d 1.9
23.9 21.3 8.4 3.5 P10D04e P3G10c
P9F06i 10.2 3.8 P10G02k CDSR7i P9D02c M737e
3.7 2.4
P9D06f P8G08g CDSR7a M737b 2.0 P6A06b
5.7 P7C03e 7.5 P10C05f
P2D11a 6.1 P1D03b 4b 19.5 7.1
6.5 13.5 P10B12a
CDSR7g P4C06d 93.7 cM
8.1 CDSR7j P7B12g
19.3 P10G06i 11.5 21.1
1.8 CSU539b 4a
M891a 5.0 CDSR94d P7E03g 141.6 cM
8.8 P4G04b
P4E01b 13.0
7.6 32.3
P4A06e P4c06e P8D04b
12.5
CDSR94l 17.6 P7H01d 33.2
21.9 4a
P7A03g 200.2 cM
P8D04a 3b P4E03b
3a
153.2 cM 12.8
222.4 cM
P9B03h
16.7

M248b
5a
186.0 cM

Fig. 3.3 (continued)

3 Cenchrus 45

P10B12d CSU351b P1F011 P10G03i P9E10g M869d SB1773f

13.4 8.1
13.4 HHU27d
22.3 24.4 22.8
P10F12h 30.1 HHU27b
6.4 Pcs2b 15.8
CDSR7e P1D03d 1.7
C147d SB1773d M466c
5a 18.6 26.4
16.4 P6H01b
22.3 cM P10G02b P10G07h 21.5 15.8
7.2 P4F01a Sub31f 15.5
P4D05f
CDSR7c 1.5
2.9 P8B08a 5.8 P4D05d 8.3
11.5 Sub31e P10D08h M568b
5.8 P7C04c 8.2 Pcs4d P7E10f
P1C01b 10.4 7.7 14.5 6.2
1.1 P9H09c P9G09d P7G12b
5.0 P3F02c P10F04c P6C02c
CSU460h 10.7 16.4
8.5 18.8
P9A10j 24.1 17.0
P3B11d
P1H11a CSU781c
15.2
17.0 9.6 P4B11b 2a, LOD-4.90
P6A08d P7G04a P4F01c
6b 3.7 7b
14.5 P7G10e 2.8 P8B08c 17.6
76.8 cM 72.9 cM
P9F02c P10D08i
9.9 P8F03a
21.9 26.4 P8D08b 8.5
15.4 Pcs5b
P7H07i 7a
P8C02c P8C06g 100.5 cM
21.8 6a
22.5 111.6 cM
P1F08B
15.9 M812g
P1H02f 6a
5b 131.2 cM
171.4 cM

P8B12b P8H05i CSU410f CSU410b P10E11a P3A07a P3F08e P9E02h

7.1 12.4
SB1773e 14.0 18.7 16.9
19.5 P9C10b P9c10c
25.3 P4D05e 2.8 26.1
CSU781b 2.1 P11B11d
23.4 12.3 P3E01d P8H05h 5.9 C746c
7.3 8b P1C05a
P7H08c P4D05c P12C05b 16.9 cM P8F02f
CSU423f 8.1 28.0 9.6
M466b 20.2 17.6
19.7 2.6 P5G10b P4D05h
Pcs4a 9.0
18.8
HHU27a 16.8 CSU479i M466f
2.1 Pca5c Pca5a
M466a
P9F11f 8.6 14.2 17.2
PAp01 CSU423d 16.5 24.6
1.4 P8C08j P3A07g P3A07e P13G07a Pca8a
25.1 P2F06e 8a
9b
21.7 28.2 8b 59.6 cM P1F02a
9a 61.0 cM
63.2 cM
CDSR94a 67.9 cM
P8C08h
7a P8C03c
7b
74.4 cM 8a
78.8 cM
82.1 cM

P6E08f P6H01a P6D01b P9A08g P11F10d P9A08i P7A05f P10H3d

4.2 7.1
P2B07c
16.6 7.8 15.6 P9A08e 17.3
21.1 24.4 P9E10f 8.8
P2B07b P1H11b 27.5
3.4 8.0 P2F07c P11C07c
P7H09f P9A08c P7H09c
6.3 P3G11l 3.5 P10B07c 3.8 14.5 13.9
3.7 M568c 18.3
P10C11h P3G11a 9.9 P9A08k 3.7 P2A09e
5.2 14.2 1.5 P9G04c P4D08c
P10C11e 12.6 P10C11c P2B07f M568a
8.0 11b 13.6
P9C04a 11.7 P9C04i
P7H09d 8.4 25.0
5.6 7.3 P6E08g 23.4 34.3 cM P13F03d
P7E03j P10C11g 3.0 P9C04g
7.7 P6C04c 12.1
10b 11a 11b
P3G11h P9C04h RZ_478_2 P2C02d
5.7 48.0 cM 48.2 cM 46.4 cM 7.6
P3G11b 11.8 9.5 P7G01c
10a P4D08b
P8G08i 11.9
59.7 cM 9.1
17.8 P7B03a
P9C04f 12b
11a
P6G04b 75.7 cM
76.0 cM 12a
P9H09i P5D01d P5H10h P1H08e 84.2 cM
8.1
P6C07d
24.6 21.7 22.6 22.6 P10E03c P3B11g P6C10g C250e
13.3 2.5
P10A08f
P8G09e P9G04f
2.6 P3E01g CSU413a 21.8 17.7 23.0
P9H09g 2.3 26.5
CSU428e 16.0
13.0 23.7 P7D12f P1B07e
21.4 P1D02k
P7H05f P1E05i 3.2 P6B07c
9.3 P9B09d
P10H08i P1H04d 26.7 25.8 22.7
P1F12b 20.3 14.9
11.0
23.0 P1H04c P10H08e M869b C250f
P4F02b 16 9.6 P8C02d
36.1 19 20
57.6 cM P4E12f 18 46.0 cM 45.7 cM
P10F11e 17
23.6 15 48.5 cM
54.2 cM
69.3 cM
P10D08a CSU525n
14
18.5
83.5 cM
Sub31b
13
100.6 cM

Fig. 3.3 (continued)

46 S. Goel et al.

CSU428a P7E03m P5H01b P8G01g P10A08h P8A07d P5D01c P5H08c

4.4 2.4 M443b
P12E05b 14.0
19.9 18.4 10.4 17.1 18.2
P3E09c 25.0
P4G01l 24.6
P12E05a P5H01a 6.1 P5H10a P9E02i
4.4 P4G04d P4G01e 6.4 15.1
1.7 P8B04f 15.8 RZ478b P1A04c
P7C06g 5.8 P8C08e P11F08c
22.0 P1B07f
21.8 Pca12e 25 18.1 26
18.4 24 27.0 cM
23 29.1 cM
P8C02e 30.4 cM P6A07b
P2C03f 32.9 cM 1.5
RZ166c 22 P8A07j P3E09e
21 40.4 cM 3.5 P8H11a
15.3 8.3
44.4 cM
P9H09h M180a
10.6
P3B04b 26.3
5.5
P7C07e
RZ166f
26.6 25
82.7 cM

P12B03b
P7A05k P1C12d RG463a P7C12c P4G01m P2C02e P1F01m
8.4 11.8
P11F08g P10F12c 13.4
20.8 18.1
7.4 26.5 24.8 23.7
P10H07g P9G04d
1.3 18.6 P7A05c
P9H02h P7A05j 33
9.5 RZ166d
CSU706e 31 32 13.4 cM
M180f P7A09c P8H07d 29 30 20.8 cM 18.1 cM
27 28 22
24.8 cM 23.7 cM
26.6 cM 26.5 cM 131.1 cM

P6G02e CSU705c CSU539d P2A09d P12B03f P9H10h M466d CSU409g

8.1 11.3 9.8
P3G08f 14.3
P9H09d 19.9 P4H07a
30.7 22.8
14.2 11.7 P7D12i
32.3 13.2 7.3
P7D12d Pca8d
P3H12d M568e P1E09g
C250k P7G10c
19.2 16.3 12.6
P10F12f 15.9
5.1 25.6 M869e
28.1 P8H07c 27.3 P6H09d
4.6 M869c CSU413c 41
P7D05c 39 40 34.2 cM
4.4 38
CSU377l 2.5 P12C03e P2A06f 39.1 cM 37.8 cM
P8H04f CDSR18b 40.4 cM
5.5 37
21.3 P1D02j 36
48.0 cM
50.1 cM
P8D02c 17.2
12.1
P6G10l P9H09b
6.3 CSU527a P8G09b P9H08j P9H08e P1A10a
P6B12c 8.8 7.0
9.3 P2D06b 13.8
P9G01i P8G09f 5.0
4.3 P8H06a P12F04n 24.1
36.0 29.5 P12D02c
17.9
14.2 15.1 12.6
C250h M248c
P9B03g P3C04d P1F12c 46
15.8 P1E06b SB41a 43 45
42 44 24.1 cM
C250g 35 27.3 cM 27.1 cM 26.4 cM
108.1 cM 29.5 cM
18.3
P8C02b
CSU399e
16.4 SubEe P10E11b P8C02b P7G01a P1A02c
7.3 6.1
P4E03h 15.5 P7E03k P6G05a
8.0 19.8 18.9 51
P10G2g 50
9.9 P9E02k 7.3 cM 6.1 cM
M834c CSU460c P12C05a 49
47 48 15.5 cM
18.2 19.8 cM 18.9 cM
P8G01h

29.0

P12D02f
34
241.2 cM

Fig. 3.3 Maternal (white linkage groups) and paternal (black linkage groups) RFLP maps of Pennisetum ciliare. Framework map
(LOD > 2.0) markers and map distances (Kosambi map units) are shown to the right and left of the horizontal lines, respectively

cDNAs and gDNAs from across the Poaceae were nal parent with an average marker interval of
hybridized to 87 F1 hybrids derived from a cross 10.8 cm, while 78 SDRFs did not show any linkage.
between heterozygous sexual genotype with another In the paternal map, 245 SDRFs were assembled into
highly heterozygous apomictic genotype. The 460 42 linkage groups spanning over 2,757 cm with an
polymorphic probes yielded segregation data for average marker interval of 11.3, while 53 SDRFs
400 single dose restriction fragments (SDRF) in did not show any linkage. These linkage maps give
female parent and 298 SDRF in male parent. MAP- an overall estimate of 4,309 Centi Mozgan and
MAKER EXP v3.0 assembled 322 SDRFs into 47 3,679 Centi Mozgan maternal and paternal genome
linkage groups spanning over 3,464 cm in the mater- sizes, respectively. More refined genetic maps can be
3 Cenchrus 47

developed by inclusion of more markers, which will provided progenies segregating for sexuality and apo-
assist in locating quantitative trait loci (QTLs) con- mixis, although results of such crosses do not seem to
trolling the major agronomically important traits. be compatible with the models of disomic and allote-
trasomic inheritance of apospory locus (Gustine et al.
1997).
3.10 BAC Libraries Segregation analysis of isozyme, protein, and
RAPD markers within half-sib progenies of both
open-pollinated sexual and apomictic lines by Gustine
Roche et al. (2002) constructed BAC libraries from the
et al. (1996) does not reveal any marker co-segregating
apomictic line B-12-9. This buffelgrass bacterial arti-
with apospory, although subsequent bulked segregant
ficial chromosome (BAC) library contains 68,736
analysis using RAPD markers in two populations
clones with an average insert size of 109 kb, which
raised through crossing of sexual B-2s  apomictic
covers 4.8 haploid genome equivalents. These BAC
Higgins and B-2s  B-12-9 identified two RAPD mar-
clones will serve as a useful tool for more refined
kers (J16-800, M02-680) tightly linked to aposporus
mapping of buffelgrass genome through the applica-
locus (Gustine et al. 1997).
tion of more advanced techniques such as FISH, which
Ozias-Akins et al. (1998) reported 12 co-segregat-
will ultimately help in resolving physical location of
ing SCAR markers linked to apomixis in Pennisetum
genome complexes.
squamulatum. Roche et al. (1999) analyzed these
markers in two Cenchrus populations raised earlier
by Gustine et al. (1997). Out of these 12 markers, 9
3.11 Genetics and Molecular Mechanism
markers (UGT-197, P16R, Q8M, U12H, V4, X18R,
of Apomixis in Buffelgrass A10H, C4, and 07M) and another putative marker
C16 were found to be associated only with aposporous
The genus Cenchrus has been studied extensively due mode of reproduction in buffelgrass populations. All
to the existing trait of apomixis and hence needs a markers are segregated as dominant locus and these
special mention. This trait is promising and can play markers are also conserved between P. squamulatum
rich economic dividends, if deciphered. Studies focus- and apomictic C. ciliaris. They concluded that all the
ing on this trait have led to interesting information in markers clustered together at what was called “apos-
the genus Cenchrus. The discovery of sexual lines in pory specific genomic region (ASGR)”. ASGR has
C. ciliaris provides an opportunity to study the genet- been shown to be hemizygous with suppressed recom-
ics of apomixis in this grass species. Taliaferro and bination in P. squamulatum (Ozias-Akins et al. 1998).
Bashaw (1966) assayed the inheritance pattern of apo- Jessup et al. (2002) reported two markers in C. ciliaris,
mixis in selfed progenies of sexual buffelgrass (B-1s) HHU27 and UGT-197, flanking the aposporus locus at
and F1 progenies of a cross between sexual (B-1s) and a distance of 1.4 cm and 10.7 cm, respectively,
two apomictic lines. It was concluded in this study that although Goel et al. (2006) did not find any recombina-
the gene controlling apomixis is dominant and sexual- tion between these markers and ASGR in a population
ity is controlled by a gene epistatic to gene controlling of 94 individuals from a cross between P. glaucum
apomixis. The genotype of apomictic plants will be (induced tetraploid, 2n ¼ 28) and P. squamulatum
“A-bb” and those for sexual will be “aabb,” suggesting (2n ¼ 56).
that the sexual plant originated as a result of nat- Roche et al. (2002) isolated the BAC clones from
ural mutation of the dominant apomictic gene. A the library of two apomictic grasses, P. squamulatum
subsequent study by Sherwood et al. (1994) from self- and C. ciliaris, using six low copy SCAR markers
ing and intercrossing of sexual B-2s lines and progeny tightly linked with ASGR. Fingerprinting analysis of
of open-pollinated sexual B-2s and other two apomic- the BAC clones indicates that the markers, although
tic lines concluded that inheritance pattern in buffel- clustered together in a single genomic region, are
grass shows two loci tetrasomic model. These authors duplicated several times within the ASGR in both
also suggested that the sexual line used by Taliaferro the species (Roche et al. 2002). FISH analysis using
and Bashaw in 1966 was highly sexual but facul- the BAC clones containing SCAR markers as probes
tatively apomictic because selfing of these lines confirmed the results from the earlier genetic and
48 S. Goel et al.

molecular mapping that apomixis is controlled by a 3.12 Genomic Database

single dominant, hemizygous locus in both P. squa-
mulatum and C. ciliaris (Goel et al. 2003; Akiyama
The NCBI database has the record of 87 core nucleo-
et al. 2005). The FISH results revealed that ASGR in
tide sequences of Cenchrus (36 of C. ciliaris, 12 of
C. ciliaris is located on the short arm, close to the
C. echinatus, 9 of C. setiger, 9 of C. pilosus, 8 of
centromere of a chromosome that also contains the
C. myosuroides, 5 of C. calyculatus, 4 of C. incertus,
18S rDNA locus (Goel et al. 2003; Akiyama et al.
2 of C. argimonoides and 2 of C. brownii), 21,729
2005). The ASGR carrying chromosome does not
expressed sequence tag (EST), 2,145 genome survey
appear to have a morphologically similar chromosome
sequence (GSS) starting from Genebank accession
in C. ciliaris genome. Despite its difference in mor-
no ED544199 to ED 546344 and 45 amino acid
phology, ASGR carrying chromosome pairs with other
sequences (28 of C. ciliaris, 9 of C. echinatus and
chromosomes during meiosis I forms a quadrivalent
8 of C. pilosus). The core nucleotide sequences consist
(Akiyama et al. 2005). This suggests that ASGR car-
of published and unpublished diverse sequences rang-
rying chromosome have at least one homeologous
ing from apomixis-associated mRNA, tRNA, ribo-
chromosome that may undergo synapsis and recom-
somal protein gene, chloroplast ndhF gene, knotted 1
bination during meiosis (Akiyama et al. 2005). Physi-
gene, rbcL, phytochrome B genes to signaling protein-
cal mapping studies also suggest that suppression of
like mRNA sequences. Out of 36 nucleotide sequences
recombination is mainly due to hemizygocity of ASGR
in C. ciliaris, 14 are of apomoixis-associated mRNA
(Goel et al. 2006). Lack of recombination in ASGR
submitted by Gustine et al. (1996), Conner et al.
hampered the construction of a genetic map and
(2008), and Singh et al. (2007), and six are of chloro-
further isolation of novel genes controlling the apo-
plast marker genes ndhF (Giussani et al. 2001) and
mixis in buffelgrass because it is unlikely that fur-
knotted 1 (Doust et al. 2007) . The unpublished 21,729
ther recombination studies will provide a finer map
ESTs in the database represent the total transcriptome
of ASGR (Akiyama et al. 2005).
of the apomictic pistils of C. ciliaris, submitted by A.
An alternative approach to identify the apomictic
H. Paterson and S. R. Schulze, Plant Genome Mapping
genes in C. ciliaris has been to analyze genes differen-
Laboratory, University of Georgia. In addition to these
tially expressed during the sexual and apomictic repro-
ESTs, A. Hussey Mark submitted eight ESTs that
duction (Vielle-Calzda et al.1996). Two cDNA clones
show differential expression between apomictic and
(Pca-2 and Pca-3) were found to be expressed specifi-
sexual ovaries in P. ciliare. These sequences appear
cally in apomictic ovaries, one clone (Pca-2) only in
in GeneBank, EMBL and DDBJ under the acces-
sexual ovaries and one fragment (Pca-1) in both sexual
sion numbers U65082, U65383, U65384, U65385,
and apomictic ovaries. Two more genes Pca 21 and
U65386, U65387, U65388, and U65389 (Vielle-
Pca 24 that expressed preferentially (Pca 21) and
Calzada et al. 1996).
specifically (Pca 24) in apomictic ovaries have been
GSS data in the GeneBank are the short-gun
isolated through subtracted cDNA microarray analysis
sequenced high quality BAC clone reads of “ASGR”
(Singh et al. 2007). Pca 21 shows homology with two
of C. ciliaris submitted by Peggy Ozias-Akins, Uni-
wheat genes expressed specifically in flowers and in
versity of Georgia, Tifton Campus. These sequences
inflorescence and Pca 24 with a maize cDNA that
are also available in FUNGEN ASGR database at the
encode putative CHCH (coiled-coil-helix–coiled-
web portal http://asgr.uga.edu (Conner et al. 2008).
coil-helix) gene, which have developmental roles
After assembling these sequences in contigs, BLASTX
(Singh et al. 2007), but none of the genes have yet
analysis with the Uniport TrEMBL database shows 25
shown to be linked with ASGR in buffelgrass (Ozias-
sequence contigs from C. ciliaris similar to proteins
Akins et al. 2003; Singh et al. 2007). These results
with diverse functions reported from other species
show that even the gene expression is conserved
(Conner et al. 2008). These putative protein coding
between sexual and apomictic ovaries. It may be
regions were mostly from the BACs mapped to low
because of the same pattern of megaspore mother
copy region of ASGR.
cell differentiation and meiosis in sexual and apos-
The EMBL database has 26,011 nucleotide
porus ovules up to the tetrad stage (Vielle-Calzada
sequence entry out of which 63 sequences are of
et al. 1996).
3 Cenchrus 49

coding sequence. This database also has 178 amino C. ciliaris accessions predominates. The list of germ-
acid sequence entry. These database also include the plasm hold can be accessed through the website http://
sequences of the related genus of Cenchrus and nucle- www2.dpi.qld.gov.au/extra/asp/auspgris/. It provides
otide sequences of the pathogens, which infects these passport information on germplasm collections related
plants. to the identity, origin, characteristics, and evaluation
of accessions held. Germplasm samples are conserved
under long-term storage conditions and made avail-
able throughout the world for research, breeding, and
3.13 Germplasm Banks conservation.
Genetic Resources Program of the International
National Bureau of Plant Genetic Resources Centre for Tropical Agriculture (CIAT) is another
(NBPGR), New Delhi, is the nodal organization in genebank, which is among the top germplasm hold-
India for exchange, quarantine, collection, conserva- ings of tropical forages obtained or collected from
tion, evaluation, and the systematic documentation of over 141 countries. Germplasms are conserved under
plant genetic resources. The genebanks of NBPGR an agreement with the International Treaty on Plant
and its regional research centers including Indian Genetic Resources for Food and Agriculture of the
Grassland and Fodder Research Institute (IGFRI), FAO. The genebank has listed 21 accessions of differ-
Jansi, Central Arid Zone Research Institute, Jodhpur, ent species of Cenchrus available for distribution. The
and Agricultural Research Institute, Bikaner, conserve accessions can be accessed through the website http://
the germplasm collected from different regions of the www.ciat.cgiar.org/urg.
world, and the samples of the conserved germplasm A total of 444 accessions of different species of
are made available to the interested sectors through Cenchrus have been listed the Genetic Resources Unit
Germplasm Exchange Division of NBPGR. of the Institute of Grassland and Environmental
A total 750 accessions of wild species of the genera Research (IGER). This unit is primarily concerned
Pennisetum and Cenchrus, assembled from 50 with the collection, storage, characterization, and doc-
countries are conserved at the genebank of ICRISAT umentation of temperate forage grasses and legumes.
(India). Many of the wild relatives have evolved sur- IGER is also responsible for the European Central
viving drought, floods, extreme heat and cold, and in Crop Databases for Lolium (rye-grass) and Trifolium
the process, they have become adapted or developed repens (white clover), and the information of the crops
resistance to the pests and diseases, which cause heavy can be accessed from IGER’s web page.
losses to the crops. These accessions can be a valuable
source for research and development.
International Livestock Research Institute (ILRI)
situated in Kenya and Ethiopia held 294 accessions 3.14 Potential and Pitfalls
of Cenchrus. Most of the accessions were collected
from the tropical African countries and are predomi- Because of its low cost of establishment in low nutri-
nated by C. ciliaris. The list of germplasm held at this ent habitats and high forage production and relative
institute can be accessed through the website http:// resistance to drought conditions and overgrazing,
www.ilri.org/forage/Cenchrus.pdf. C. ciliaris serves as an important fodder grass for the
The National Germplasm Resources Laboratory, arid and semiarid tropical regions of the world. It has
Beltsville, Maryland (GRIN, USDA), has listed 600 also been reported that the biomass production and
accessions of P. ciliare (syn. C. ciliaris) and 12 acces- water use efficiency of this grass is better than other
sions of other species of Cenchrus available in its arid zone grasses like Lasiurus scindicus, Panicum
genebank. GRIN provides information of the acces- turgidum, and Coelachyrum piercei (Osman et al.
sions and its availability for distribution. 2008). Another aspect prevailing in the genus Cen-
AusPGRIS – the Australian Plant Genetic Resource chrus is the apospory trait. At the first conference on
Information Service holds 453 accessions of Cenchrus apomixis held in September 1995 at the College Sta-
collected from 33 countries of the world. The acces- tion of the Texas A&M University, it was stated that
sions represent ten species; however, the number of “harnessing apomixis genes for plant improvement
50 S. Goel et al.

offers the potential for quantum in agriculture produc- Bogdan AV (1977) Tropical pastures and fodder plants. Long-
tion – an ‘asexual revolution’, the benefits of which man, London, UK
Chandra A, Dubey A (2007) Transferability of STS markers
could dwarf those of green revolution,” C. ciliaris for studying genetic diversity within the genus Cenchrus
along with a relative species P. squamulatum is more (Poaceae). Curr Sci 92:961–967
extensively studied to harness the benefit of apomixis Chandra A, Dubey A (2008) Evaluation of genus Cenchrus
mechanism. based on malondialdehyde, proline content, specific leaf
area and carbon isotope discrimination for drought tolerance
Most studies in the genus Cenchrus has been con- and divergence of species. Acta Physiol Plant 30:53–61
ducted considering economic impacts limiting most Chen C-H, Kuoh C-S (2004) Cenchrus ciliaris L., a newly
studies to C. ciliaris and little has been done to study naturalized grass in Taiwan. Taiwania 49(4):232–236
other species in genus. This leaves a huge scope to Clayton WD, Harman KT, Williamson H (2006 onwards) Grass-
Base – the online world grass flora. http://www.kew.org/
enrich the information in other species of this genus. data/grasses-db.html. Accessed 3 Feb 2009
Many species in genus Cenchrus are potential invasive Conner JA, Goel S, Gunawan G, Marie-Michele C-P, Virgil Ed
weeds threatening many keystone habitats of many J, Chun L, Haiming W, Lee HP, John EM, Jeremy D, Lixing
plants and animal species. More in-depth knowledge Y, Jeffrey LB, Patricia EK, Ozias-Akins P (2008) Sequence
analysis of bacterial artificial chromosome clones from
of these species will also be helpful in controlling the apospory-specific genomic region of Pennisetum and
spread of these weeds. In conclusion, C. ciliaris is a Cenchrus. Plant Physiol 147:1396–1411
crop, which has not gone through the bottleneck of Cook BG, Pengelly BC, Brown SD, Donnelly JL, Eagles DA,
domestication and still open for exploitation of natu- Franco MA, Hanson J, Mullen BF, Partridge IJ, Peters M,
Schultze-Kraft R (2005) Tropical forages: an interactive
rally occurring variation in the species, but occurrence selection tool. CSIRO, DPI&F (Qld), CIAT and ILRI,
of both sexual and apomictic genotypes provide an Brisbane, Australia. http://www.tropicalforages.info/.
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ring in other closely related species from the genus. Correlld S, Johnston MC (1970) Manual of the vascular plants
of Texas. Texas Research Foundation, Renner, TX, USA
For exploiting these species, a systematic study Cox JR, Martin-R MH, Ibarra-F FA, Fourie JH, Rethman JFG,
involving various aspects and carried out on all the Wilcox DG (1988) The influence of climate and soils on the
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DeLisle DG (1963) Taxonomy and distributon of the genus
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Chapter 4
Cynodon

Yanqi Wu

4.1 Introduction Common bermudagrass was introduced to colonial

America at least by the mid-1700s and spread rapidly
with the development of agriculture initially in the
Cynodon L. C. Rich. is a small genus of warm-season
southeast and subsequently in southcentral and south-
species in the tribe Cynodonteae, subfamily Chlori-
west regions. Early reactions of farmers to bermuda-
doideae of the grass family Poaceae (Gramineae).
grass were mixed as the grass was a serious weed in
Though small in number of species, the genus contains
row crops, especially cotton (Nelson and Burns 2006).
species that are economically immensely important as
Many southern and central states listed it as a noxious
grazed or stored forages, for soil stabilization, and turf.
weed. It was not until the middle of the twentieth
The focus of this chapter is on use of Cynodon species
century that attitudes changed regarding its value to
for forage. Although Cynodon species have inciden-
agriculture, and circumstances led to its widespread
tally provided forage for herbivores for many centu-
adoption as a cultivated pasture and hay crop. Harlan
ries, it was not until the middle of the twentieth
(1970) described “the development of C. dactylon var.
century that large scale monoculture use developed
dactylon as an improved hay and pasture grass suitable
to provide forage for domesticated livestock. This
for intensive management as almost entirely an Amer-
development was in the southern USA and occurred
ican phenomenon.” He cited several factors contribut-
in large measure due to breeding gains, leading to
ing to bermudagrass becoming a major perennial grass
widely adapted cultivars with greatly enhanced forage
for animal grazing and hay production in the southern
yield potential. The most important of the Cynodon
USA. Prominent among the factors was the release of
species as a forage is C. dactylon (L.) Pers. This
“Coastal” bermudagrass in 1943. Coastal was devel-
species consists of six taxonomic varieties that vary
oped by USDA-ARS Geneticist G.W. Burton at the
substantially in magnitude of natural distribution,
Coastal Plains Experiment Station, Tifton, GA. Re-
genetic diversity, and value for forage. The taxon of
lease of this high yielding variety coincided with
greatest importance is C. dactylon (L.) Pers. var. dac-
other factors, leading to its widespread adoption in
tylon. This taxon is distributed across all continents
the southern USA. These factors included (1) a
and on most ocean islands between latitudes of
decrease in the cotton industry in the southern USA,
approximately 45
North and South. It is enormously
(2) a concurrent increase in the cattle industry in the
polymorphic with forms adapted to a range of climatic
same region, and (3) greater availability of nitrogen
and edaphic conditions. It is generally known as “com-
fertilizer. These circumstances led to greatly increased
mon bermudagrass” because of its prevalence.
research effort by state agricultural experiment sta-
tions and other research entities on the use of bermu-
dagrass as a pasture and hay crop. The development of
specialized sprig harvesting and planting equipment
Y. Wu
also increased the use of bermudagrass (Taliaferro
Department of Plant and Soil Sciences, Oklahoma State
University, 368 Ag Hall, Stillwater, OK 74078, USA et al. 2004a). Currently, forage bermudagrass is grown
e-mail: [email protected] on about 10–12 million ha in the USA as a major

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 53
DOI 10.1007/978-3-642-14255-0_4, # Springer-Verlag Berlin Heidelberg 2011
54 Y. Wu

perennial grass feed base for animal husbandry (Talia- racemes, which usually are dark red in color, similar
ferro et al. 2004). Bouton (2007) estimated the current to those of C. plectostachyus, but relatively smaller.
economic value of the forage systems in the southeast- Bright red racemes are digitate from a few to 15, with
ern 14 States to be approximately US$11.6 billion each raceme being straight and 4–8 cm in length.
annually, in which forage bermudagrass is a major Spikelets are sessile, normally two rows on a rachis.
component along with tall fescue [Festuca arundinacea Glumes are persistent and similar but about 3/4
Schreb., recently Schedonorus phoenix (Scop.) Holub] spikelet length. Florets are laterally compressed,
and bahiagrass (Paspalum notatum Flugge). 2.5–3.0 mm long, and red or purple. Lemma is keeled
The remainder of this chapter focuses on the taxon- and hairy on the keel. Leaf blades of C. aethiopicus is
omy, distribution, origin and evolution, and germ- 3–25 cm long and 3–7 mm wide, linear lanceolate,
plasm conservation of C. dactylon var. dactylon and coarse, stiff and harsh, glabrous or sparsely hairy, and
related species relative to their use in the development conduplicate (Harlan et al. 1970b; Clayton et al.
of new forage cultivars. 2009). Culms are rather woody at maturity, can be
40–100 cm long, and 2–6 mm in diameter. Stolons
have long internodes and are stout (Harlan et al.
1970b). Harlan et al. (1970b) indicated that C. aethio-
4.2 Basic Botany of the Species picus was distributed in wet habitat on coastal plains to
highlands from Ethiopia to Transvaal of South Africa.
In addition to its distribution in Africa, it is distributed
4.2.1 Taxonomy, Morphology, and
in Australia (Clayton et al. 2009). Taliaferro et al.
Distribution (2004a) indicated that stargrass species, including
C. aethiopicus, are indigenous to tropical Africa but
The genus Cynodon consists of nine species in the have been exported to many other tropical and semi-
revised taxonomic classification of Harlan et al. tropical regions of the world. C. aethiopicus cultivars
(1970b). The species are C. aethiopicus Clayton et have been developed by biotype selection (Taliaferro
Harlan, C. arcuatus J. S. Presl ex C. B. Presl, C. barberi et al. 2004a). Available evidence indicates some
Rang. Et Tad., C. dactylon (L.) Pers., C. incompletus hybridization potential between C. aethiopicus and
Nees, C. nlemfuensis Vanderyst, C. plectostachyus C. dactylon (Harlan et al. 1969). Harlan et al. (1969)
(K. Schum.) Pilg., and C. transvaalensis Burtt-Davy attempted many crosses between plants of the two
(de Wet and Harlan 1970). de Wet and Harlan (1970) species with only six triploid hybrids produced from
did not include C. x magennisii in the revised system diploid (2n ¼ 2x ¼ 18) C. aethiopicus by tetraploid
while they listed the taxon as a species in “A guide to (2n ¼ 4x ¼ 36) C. dactylon. No hybrids were gener-
the species of Cynodon (Gramineae)” (Harlan et al. ated by crossing diploid C. dactylon with diploid and
1970b). C. x magennisii is a naturally occurring trip- tetraploid C. aethiopicus (Harlan et al. 1969). de Wet
loid hybrid between C. dactylon and C. transvaalensis and Harlan (1970) subsequently reported that tetra-
(Harlan et al. 1970a). Clayton et al. (2009) lists 10 ploid C. aethiopicus crossed readily with tetraploid
Cynodon species in Kew’s online grass database. In C. dactylon var. dactylon to produce hybrids, which
this list, C. coursii is ranked as a species, while it was a were partially fertile. Based on this information, it
variety of C. dactylon in the classification of Harlan would be possible to use C. aethiopicus as a genetic
et al. (1970b); C. radiatus Roth ex Roem. and Schult source to improve C. dactylon as a forage grass using
was described as an annual species and as a synonym traditional breeding procedures.
of C. arcuatus (Clayton and Harman 2002), while C. arcuatus is a sod-forming grass without rhi-
Harlan et al. (1970b) described it as a perennial. zomes (Harlan et al. 1970b). Its inflorescence nor-
C. parviglumis Ohwi is listed as a species by Clayton mally has 4–5 racemes, which are long, slender, and
et al. (2009) but not described by Harlan et al. (1970b). flexuous. Spikelets are rather densely arrayed on
C. aethiopicus is a perennial sod-forming species racemes. Plants of C. arcuatus in our greenhouse are
without rhizomes (Harlan et al. 1970b). The species perennial and readily set seed. Glumes are subequal
has low tolerance to freezing temperatures. Its inflo- and about one-half the spikelet length. Lemma is hairy
rescence is composed of one to three whorls of on keel. Stolons are slender and have short internodes.
4 Cynodon 55

One distinct character of C. arcuatus is that its leaf has worldwide distribution in warm and temperate
blade is very broadly lanceolate. The grass is climates, var. aridus is relatively widely distributed,
distributed in northern Madagascar, adjacent islands and vars. afghanicus, coursii, elegans, and polevansii
to Sri Lanka (Ceylon), South and East India, Southeast are narrow endemics (Harlan and de Wet 1969; Harlan
Asia, Southern China, Philippines, Indonesia, and et al. 1970b; de Wet and Harlan 1970).
Northern Australia (Harlan et al. 1970b). Habitat of Var. afghanicus is an Afghanistan endemic taxon
the species is in lowland wet areas. C. arcuatus is recognized by Harlan and de Wet. The characteristics
genetically completely isolated from all other Cyno- of var. afghanicus include no rhizomes in the diploid
don species (Harlan et al. 1969). The grass has limited race but very short rhizome-like structures in tetra-
forage value, but its genetic isolation from other Cyno- ploid race, and its racemes bear closely imbricated
don species precludes its use in interspecific hybridi- spikelets. Even without rhizomes, living plants of
zation as a means of breeding (Harlan 1970). var. afghanicus overwintered in Stillwater, OK, for
C. barberi plants are small in size, rare in abun- several years, demonstrating tolerance to freezing
dance, and confined in wet areas and edges of perma- temperatures (Harlan et al. 1970b). The variety is
nent water courses. The species is endemic to southern found only in lowland sites of steppes and along irri-
India and of little value as forage, but it has some local gation ditches. It seems that tetraploids have a wider
use as turf (Harlan 1970). The grass is perennial and natural distribution than diploids.
sod-forming, but has no rhizomes, and has low toler- Var. aridus is diverse in morphology, but it has less
ance to freezing temperatures. Culms can ascend up to genetic variability than var. dactylon. A race with
25–40 cm tall (Clayton et al. 2009), but stolons form relatively small plants is found in southern India
loose mats less than 10 cm in height (Harlan et al. while a race with much larger plants occurs in South
1970b). An inflorescence normally consists of 2–5 Africa. Two to several racemes constitute an inflores-
short, slender and delicate racemes. Glumes are cence with one or rarely two whorls. Glumes are about
unequal, and longest glumes can slightly exceed the 2/3 the length of spikelets, which are loosely arranged
length of the lemma, which is somewhat hairy on keel. on the racemes. Var. aridus plants have little winter
Leaf blades of C. barberi are short, broadly lanceolate hardiness in Stillwater, OK, but their rhizomes can
with long, sparse hairs. Harlan et al. (1970b) indicated grow deep enough in soil to escape freezing tempera-
that this species is somewhat a miniature version of tures (Harlan et al. 1970b). Harlan and de Wet (1969)
C. arcuatus, except that inflorescences of the two indicated that the variety is distributed in a vast geo-
species are very different. C. barberi is genetically graphic area from South Africa, Zambia, Tanzania, to
well isolated from other species in the genus Cynodon Israel and then to India and Sri Lanka (Ceylon). The
(Harlan et al. 1970c). variety is called “giant” bermudagrass in the USA
C. dactylon is the species of greatest genetic diver- where seed is produced in Arizona and California.
sity, widest geographic distribution, and greatest eco- Var. coursii is relatively coarse. It can grow
nomic value within the genus. Harlan et al.’s (1970b) 40–50 cm tall. Morphology of its inflorescences, race-
taxonomic classification includes six varieties within mes, spikelets, and glumes is similar to var. dactylon
the species. They are var. afghanicus Harlan et de Wet, while lemmas are pilose on keel (Harlan et al. 1970b).
var. aridus Harlan et de Wet, var. coursii Harlan et de Var. coursii has no rhizomes, and leaves are often
Wet, var. dactylon (L.) Pers., var. elegans Rendle, and bunched and appressed at the tip of culms. The variety
var. polevansii (Stent) Harlan et de Wet (Harlan et al. is distributed in the central plateau of Madagascar. It
1970b). The intraspecific classification was based on has no winter hardiness in Stillwater, OK.
data for natural distribution, morphological distinct- Var. dactylon is enormously variable with plants
ness, cytogenetic behavior, and ecology (Harlan and ranging in size from very small, fine textured types
de Wet 1969; Harlan 1970). In addition to a living suitable for turf use to large and robust ones best suited
collection of 453 accessions from 28 countries grown for forage production (Harlan and de Wet 1969). The
at Stillwater, Oklahoma, for the data collection, speci- variety is distributed across all continents and on
mens were examined in Kew, British Museum, Edin- most ocean islands between North and South latitudes
burgh, Paris, Honolulu, and Washington (Harlan and of approximately 45
. In Europe, it can extend to
de Wet 1969). Among the six varieties, var. dactylon locations, such as Yorkshire in England, Friesland in
56 Y. Wu

Netherlands, Potsdam in Germany, and southern of C. incompletus have fine, slender, reddish stolons,
Siberia in the Russian Federation (formerly USSR), linear-lanceolate somewhat hairy leaf blades, and one
all at approximately 53
N latitude (Harlan and de Wet whorl of racemes in inflorescence. The two varieties
1969). Inflorescences contain racemes arranged in one are winter hardy in Stillwater, OK (Harlan et al.
whorl, rarely two. Glumes are usually ¾ the spikelet 1970b). The species is distributed in southern Africa,
length, occasionally slightly longer. The stolons of primarily in South Africa as far north as 23
South
var. dactylon are never as large as those of the star- latitude. Var. incompletus predominates in the arid
grass species, C. plectostachyus, C. aethiopicus, or regions, while var. hirsutus predominates in the Bush-
C. nlemfuensis (Harlan et al. 1970b). The variety has veld, but their distribution ranges overlap extensively
rhizomes and likes disturbed habitats. Harlan and de in the western Transvaal (de Wet and Harlan 1971).
Wet (1969) reported finding natural spread of var. C. nlemfuensis is an important forage species with
dactylon plants in valleys about 3,000 m above sea robust plants capable of rapid growth and high biomass
levels in the southern Tibetan region but not on the yield. It is one of the stargrass species from which
high plateau of the northern Tibetan region. Harlan forage cultivars have been developed, and it has been
and de Wet (1969) recognized three races as tropical, used in interspecific hybridization with C. dactylon to
temperate, and seleucidus on the basis of appearance, produce forage cultivars. Typical plants of the species
adaptation, and geographic distribution. This variety is have inflorescences in one or rarely two or three whorls
extremely valuable and extensively used directly or as of racemes, which are very long, spreading, and flexu-
a parent to develop interspecific varieties as commer- ous, usually 5 or more, about 4 or more centimeters in
cial products for forage and turf. length (Harlan et al. 1970b). Glumes are subequal,
Var. elegans is a medium coarse variety confined to about ¾ of the length of a spikelet. The species has
southern Africa (Harlan and de Wet 1969). It has a no rhizomes and has low tolerance to freezing tem-
distinctive appearance from sympatric var. aridus. Var. peratures. Two botanical varieties were recognized in
elegans has green foliage rather than glaucous one of var. Harlan et al.’s (1970) taxonomical classification, var.
aridus (Harlan and de Wet 1969). Plants of var. elegans nlemfuensis and var. robustus. The nlemfuensis variety
can form a lax, loose sod, and its culms are decumbent is widely distributed in the wetter parts of East and
with ascending appressed leaves (Harlan et al. 1970b). Central Africa from Ethiopia to Zimbabwe (Rhodesia)
The variety has no winter hardiness in Stillwater, OK. It is and Angola. Compared to variety nlemfuensis, var.
valuable in breeding forage bermudagrass. robustus is taller and coarser and has bigger stolons.
Var. polevansii is small in plant size. Stolons are The robustus variety is mainly distributed in high rain-
rather fine with short internodes forming a dense turf, fall areas of eastern Africa, ranging from Ethiopia to
and leaves are very harsh, rigid, and erect (Harlan Zimbabwe and westward to Angola.
and de Wet 1969). The variety has only been collected C. plectostachyus is another non-rhizomatous large
from one site near Barberspan, South Africa (Harlan East African Cynodon species included in the stargrass
et al. 1970b). It has good winter hardiness in Oklahoma grouping. It is distinct from all other Cynodon species
and is dark green in color and therefore could be because of its minute glumes, which are at most 1/4 to
valuable in the development of turf type cultivars. 1/5 of the length of the spikelet (Clayton and Harlan
C. incompletus is a South African species. Like 1970). Its inflorescence is normally arranged in 2–5
several other African Cynodon species, C. incompletus whorls. It has robust, erect culms reaching heights of
has no rhizomes. The species is subclassified into two 30–90 cm. The species has low tolerance to freezing
varieties, var. incompletus and var. hirsutus (Stent) de temperatures. Its habitat is in areas of relatively high
Wet et Harlan. de Wet and Harlan (1971) indicated rainfall, particularly along the highlands of East
that var. incompletus plants have leaves, which are at Africa. The species is distributed from Ethiopia to
most moderately pilose, and lower and upper glumes Kenya, Tanzania, Zambia, Malawi, and Uganda to
are about 1/4 and 1/2 the spikelet length, respectively, Eastern Congo (Harlan et al. 1970b).
while var. hirsutus has hirsute leaves and relatively C. transvaalensis is an endemic species in South
longer glumes, which are 1/2 to 3/4 as long as the Africa. Typical plants of the species are small in size
spikelet. Lemmas are often somewhat blunt with hairs with yellowish-green, erect leaves and small inflores-
on keels (Harlan et al. 1970b). Morphologically, plants cences with the spikelets loosely arranged on the
4 Cynodon 57

racemes (de Wet and Harlan 1970). Its leaf blades are Table 4.1 Cynodon species and their somatic chromosomes
rarely more than 2 mm wide, and its stolons fine, (Harlan et al. 1970b)
slender, with short internodes. Plants of C. transvaa- Taxon Somatic
chromosome
lensis are usually found in damp areas around perma- number
nent waterholes and along stream banks (Harlan et al. C. aethiopicus Clayton et Harlan 18, 36
1970b). Its natural distribution is in a range from C. arcuatus J. S. Presl. Ex C. B. Presl. 36
southwestern Transvaal, Orange Free State to the C. barberi Rang. Et Tad. 18
northern part of the central Cape Province of South C. dactylon (L.) Pers. var. dactylon 36, 45a, 54a
Africa. It is known as “African bermudagrass” and has var. afghanicus Harlan et de Wet 18, 36
been widely used for turf and as a parent in crosses var. aridus Harlan et de Wet 18
var. coursii (A. Camus) Harlan et de Wet 36
with common bermudagrass in the development of F1
var. elegans Rendle 36
hybrid cultivars. African bermudagrass has little value var. polevansii (Stent) Harlan et de Wet 36
for forage due to its small size and management C. incompletus Nees var. incompletus 18
requirements. var. hirsutus (Stent) de Wet et Harlan 18, rarely 36
C. nlemfuensis Vanderyst var. nlemfuensis 18, rarely 36
var. robustus Clayton et Harlan 18, 36
C. plectostachyus (K. Schum.) Pilger 18, rarely 36a
4.2.2 Cytology, Karyotype, and
C. transvaalensis Burtt-Davy 18
Genome Size a
Sources: Assefa et al. (1999) and Wu et al. (2006a)

Hurcombe (1947) reported chromosome numbers of

several cultivated Cynodon species in South Africa They found that diploid (2n ¼ 2x ¼ 18), tetraploid
using root tip sections to count chromosomes. (2n ¼ 4x ¼ 36), and hexaploid (2n ¼ 6x ¼ 54)
Her investigations indicated that chromosomes of races were sympatric. de Silva and Snaydon (1995)
C. bradleyi (now considered a hybrid between studied chromosome numbers in C. dactylon of Sri
C. incompletus var. incompletus and var. hirsutus), Lanka. They reported that most populations were tet-
C. transvaalensis, C. dactylon, and C. magennisii raploids, while plants from roadsides and lawns in the
were 18, 20, 40, and 30, respectively. She believed wet region and from forests in the hill country were
that the base chromosome number of Cynodon was 10. diploids. They also added that populations from paddy
She speculated that the determination of 18 somatic fields in the wet region contained both diploid and tetra-
chromosomes for C. bradleyi was due to aneuploidy ploid plants. Surprisingly, they did not find triploids in
(Hurcombe 1947). Forbes and Burton (1963) reported any environments (de Silva and Snaydon 1995).
chromosome numbers of six Cynodon species using Wu et al. (2006a) reported four ploidy levels, trip-
root tip smears. Their study confirmed the base num- loid, tetraploid, pentaploid, and hexaploid in 119
ber of Cynodon to be 9 rather than 10. Their results Chinese C. dactylon accessions. The few triploid
indicated that C. bradleyi, C. incompletus, C. plectos- plants in the collection were considered likely inter-
tachyus, C. transvaalensis, and C. x magennisii had specific hybrids that had been introduced to the col-
respective somatic chromosome numbers of 18, 18, lection area. Tetraploids were the most common
18, 18 and 27, while C. dactylon had both diploid (88%) cytotype in the collection, which was consistent
(2n ¼ 2x ¼ 18) and tetraploid (2n ¼ 4x ¼ 36) plants. with previous reports of ploidy levels in Chinese
They indicated that some chromosome satellites could Cynodon germplasm. Pentaploids and hexaploids in
cause misinterpretation of chromosome numbers. the collection were only from eastern provinces of
Harlan et al. (1970b) published a taxonomic revi- China. They did not find diploids in the collection.
sion of the genus Cynodon, which is presented in Similarly, Kang et al. (2008) reported triploid, tetra-
Table 4.1. Their revision was based on comprehen- ploid, pentaploid, and hexaploid cytotypes among 43
sive cytotaxonomic investigations and has been Korean Cynodon accessions collected in the southern
widely adopted. regions of the Korean peninsula between 34 and 36
N
Malik and Tripathi (1968) reported three cytotypes latitude and from 126 to 129
E longitude. It was specu-
of naturally occurring C. dactylon in Udaipur, India. lated that triploids were introduced since no diploid
58 Y. Wu

plants were ever reported in the nation. More recently, 4.2.3 Agriculture Uses
Gulsen et al. (2009) reported diploids, triploids, tetra-
ploids, pentaploids, and hexaploids in 182 naturally
Several of the Cynodon species have economic value
occurring Cynodon accessions in the southern provinces
derived from their agricultural use, principally as
in Turkey. They further commented that diploids
grazed or conserved fodder for livestock. The most
occurred in three provinces, indicating their indigenous-
important of the species is C. dactylon, which includes
ness. Their results supported the statement made by
the ubiquitous common bermudagrass, C. dactylon var.
Harlan (1970) that the region from West Pakistan to
dactylon that is of paramount importance. Stargrasses
Turkey is the evolutionary center of C. dactylon.
are also of substantial importance in tropical and sub-
Ourecky (1963) reported the pachytene chromo-
tropical regions. In addition to livestock fodder, these
some morphology of a diploid C. dactylon plant from
grasses have value in controlling soil erosion and in
Afghanistan, which probably was var. afghanicus.
the remediation of spoil soils. They are also effective
Pachytene chromosomes of the Cynodon plant varied
sinks for removing excess nutrients from soils receiv-
from 16.8 to 36.3 mm in length. The shortest one in the
ing large amounts of animal effluent. They also have
genome complement was designated as chromosome 1
potential as bioenergy feedstock crops because of their
and the longest as chromosome 9 (Ourecky 1963).
high biomass production capability (Casler et al.
Chromosome 4 contains the nucleolar organizing
2009). The use of bermudagrass as turf rivals or
region. Brilman et al. (1982) reported pachytene chro-
exceeds its use as livestock fodder.
mosome morphology of diploid C. dactylon var. ari-
Bermudagrass has had a dramatic effect on the
dus and plants of var. afghanicus x var. aridus. The
livestock industry in the southern USA (Nelson and
chromosomes ranged in mean length from 20.5 to
Burns 2006). Bermudagrass pastures are managed for
44.6 mm, which was reasonably consistent with the
grazing or haying or dual purpose in the South. Star-
results of Ourecky (1963). Arm ratios were from
grass is cultivated and used in Florida, Central and
about 1.5 to 2.14 (Brilman et al. 1982). Brilman et al.
South America, the Caribbean, and tropical Africa
(1982) suggested that the genomes of var. afghanicus
(Taliaferro et al. 2004a). Taliaferro et al. (2004a)
and var. aridus are completely homologous.
pointed out many characteristics making bermuda-
Knowledge of genome size is important in plant
grass valuable for use as grazed or harvested forages.
species being investigated with molecular genetic
These include their relative ease of establishment,
techniques. The use of flow cytometry to estimate
high biomass production potential per unit area, rela-
nuclear genome size, or genomic DNA content, of
tively good palatability, and high tolerance to biotic
Cynodon species was first reported by Taliaferro
and abiotic stresses. Improved cultivars coupled with
et al. (1997). The mean nuclear DNA contents of
advanced management techniques enable bermuda-
diploids, triploids, tetraploids, and hexaploids in their
grass forage production to be a low cost system.
study were 1.11  0.04 (1,087.8  39.2 Mbp), 1.60 
Bermudagrass is a major crop of the turf industry in
0.04 (1,568  39.2 Mbp), 2.25  0.13 (2,205 
southern USA and many other countries in the world,
127.4 Mbp), and 2.80  0.14 (2,744  137.2 Mbp)
where climates are tropical, subtropical, and warm
pg/2C, respectively. Their research indicated that
nuclear DNA content can be used to estimate ploidy Table 4.2 Genome size for Cynodon native in China, Korea,
level in Cynodon. Genome sizes of diploid African, and Turkey
triploid Tifway, triploid Tifgreen, and tetraploid Ploidy (2n) Genome size range (pg/2C)
“Savannah” as reported by Arumuganathan et al. Chinese Korean Turkish
(1999) were 1.03  0.01 (1,009.4  9.8 Mbp) pg/2C, Cynodon by Cynodon Cynodon
Wu et al. by Kang et al. by Gulsen
1.61  0.00 (1,577.8  0.0 Mbp), 1.37  0.01 (2006a) (2008) et al. (2009)
(1,342.6  9.8 Mbp) pg/2C, and 1.95  0.01 (1,911  Diploid (18) N/A N/A 1.03–1.14
9.8 Mbp) pg/2C, respectively. Recently, Wu et al. Triploid (27) 1.55–1.65a 1.42–1.56a 1.44–1.62
(2006a), Kang et al. (2008), and Gulsen et al. (2009) Tetraploid (36) 1.96–2.30 1.94–2.19 1.95–2.36
reported genome sizes for various ploidy levels of Pentaploid (45) 2.37–2.49 2.54 2.56–2.75
native Cynodon accessions in China, Korea, and Tur- Hexaploid (54) 2.90–3.13 2.77–2.85 3.13–3.44
a
key using flow cytometry (Table 4.2). Likely introduced plants
4 Cynodon 59

temperate. Taliaferro (2003) indicated the use of bermu- (1970). Additionally, they have little potential for con-
dagrass as turf rivals its use as a forage grass. As a matter tributing to the development of improved forage culti-
of fact, bermudagrass used as a turfgrass is much more vars as genetic sources. It should be reemphasized that
extensive than for forage on a global scale. Because of C. transvaalensis is crucial in the breeding of hybrid
its relatively high drought tolerance (Beard 1973), ber- turf bermudagrass cultivars and also has direct use as
mudagrass has replaced other turf grasses, particularly for turf. The most valuable species for forage use
cool-season species, in turf applications as the availabil- include the stargrass species, C. aethiopicus, C. nlem-
ity of irrigation water becomes limited in many parts of fuensis, and C. plectostachyus, in addition to the cos-
the world. Currently, most bermudagrass turf is estab- mopolitan and ubiquitous species C. dactylon. The
lished using clonal hybrid cultivars, such as “Tifway”, Stargrass species are generally well adapted to tropical
“Patriot”, etc., derived from crossing C. dactylon with climates receiving 800 mm or more annual rainfall
C. transvaalensis. However, seeded cultivars of C. dac- (Taliaferro et al. 2004a).
tylon have been bred and commercialized in an acceler- C. aethiopicus has wider distribution than the other
ated pace in the recent years. Much of this latter effort is two robust East African species. Harlan (1970) noted
by private industry (Taliaferro 2003). Turf bermuda- that the species is very robust and can grow up to 2 m
grass is used on lawns, parks, roadsides, sports fields, tall. At maturity, woody culms of C. aethiopicus rattle
and golf courses, to name a few. in the wind. Because of its abundance and apparent
Bermudagrass has an exceptional ability to develop productivity, the species had probably been used con-
a dense vegetative cover, which can protect the under- siderably as a forage grass before it was recognized as
lying soil from erosion caused by wind, water, or other a species (Harlan 1970). Under heavily grazed condi-
physical forces. The sod-forming ability of bermuda- tions, C. aethiopicus and C. dactylon may look similar,
grass comes from aggressive growth of stolons and but the absence of rhizomes in C. aethiopicus and their
rhizomes, which work together with its fibrous root presence in C. dactylon can be used to distinguish
system in binding the topsoil tightly against erosion them. Genetically, the species is well isolated from
movement of soil particles. Bermudagrass can with- the other species, but at the diploid level, C. aethiopi-
stand inundation for an extended period so that it is an cus crosses with C. nlemfuensis Vanderyst var. nlem-
ideal plant species to grow along water ways, on river fuensis, and at the tetraploid level, it crosses readily
banks, and other flooded areas. Bermudagrass gener- with C. dactylon var. dactylon to produce partially
ally grows well in disturbed habitats, such as right-of- fertile hybrids (de Wet and Harlan 1970). Accord-
ways, roadsides, and urban areas including parks. The ingly, C. aethiopicus has potential to contribute to
ability of bermudagrass to provide an attractive turf the genetic improvement of bermudagrass and star-
with minimal adverse effects due to biotic and abiotic grass via its hybridization potential with forage
stresses and protect the environment through preven- C. dactylon and C. nlemfuensis var. nlemfuensis. Forage
tion of soil erosion, removal of greenhouse gasses, and cultivars of C. aethiopicus were released from ecotype
release of oxygen make it a very valuable grass. selections of natural collections (Table 4.3).
C. nlemfuensis is the most promising of the star-
grasses used directly for forage (Harlan 1970) and is
4.3 Conservation Initiatives highly valuable in the development of hybrid cultivars
from interspecific crossing with C. dactylon (Burton
1972; Burton et al. 1993). Harlan (1970) noted that
4.3.1 Species for Forage C. nlemfuensis var. robustus was used to establish
pastures at the Muguga Research Station near Nairobi
As previously mentioned, the value of Cynodon spe- and had high yield in field trials at the Henderson
cies and varieties as grazed or stored forage for rumi- Research Station. Compared to var. robustus, var.
nant animals, and their potential value in genetic nlemfuensis is finer, less robust, and less hairy. “IB-8”
improvement programs, varies from high to negligent. is a cultivar of C. nlemfuensis var. nlemfuensis, selected
The taxa C. barberi, C. arcuatus, C. transvaalensis, and released by H. R. Chheda at the University of
and C. incompletus are of minor value in grazing and Ibadan, Nigeria, in 1968 (Harlan 1970; Crowder and
have no value in hay production according to Harlan Chheda 1982). Characteristics of IB-8 are high forage
60 Y. Wu

Table 4.3 Forage cultivars of stargrass and hybrid cultivars from bermudagrass by stargrass crosses [after Taliaferro et al. (2004a)]
Cultivar Taxon Year of Origin References
release
Cv No 2 (Rhodesian Cynodon aethiopicus Unknown Zimbabwe Bogdan (1977),
No. 2 stargrass) Clatworthy (1985)
McCaleb C. aethiopicus 1975 Florida Agricultural Experiment Hodges et al. (1975)
Station (FAES)
IB-8 C. nlemfuensis var. nlemfuensis 1968 University of Ibadan, Nigeria Crowder and Chheda
(1982)
Muguga C. nlemfuensis var. nlemfuensis Unknown Zimbabwe Clatworthy (1985)
Ona C. nlemfuensis var. nlemfuensis 1979 FAES Hodges et al. (1984)
Tifton 68 C. nlemfuensis 1984 USDA-ARS and Georgia AES Burton and Monson
(1984)
Florico C. nlemfuensis var. nlemfuensis 1988 Florida and Puerto Rico AES Mislevy et al. (1989a)
and USDA-ARS
Florona C. nlemfuensis var. nlemfuensis 1988 FAES Mislevy et al. (1989b)
Coastcross-1 C. dactylon  C. nlemfuensis 1967 USDA-ARS and Ga. AES Burton (1972)
var. robustus
Brazos C. dactylon var. 1982 Tex, La., and Okla. AESs Eichhorn et al. (1984),
afghanicus  C. nlemfuensis Taliaferro (1986)
var. robustus
Tifton 85 C. dactylon  C. nlemfuensis 1992 USDA-ARS and Ga. AES Burton et al. (1993)

yield, good drought tolerance, and outstanding animal cultivars. Var. aridus ranges from small plants in
weight gain compared to local bermudagrass. Later in South India to large ones in South Africa. Forms of
the USA, several C. nlemfuensis cultivars and hybrids var. aridus from East Africa are larger and more vig-
derived from crosses of C. dactylon by C. nlemfuensis orous (Harlan 1970). Cooper and Burton (1965) eval-
were developed/selected and released (Table 4.3). uated forage and turf potential of giant bermudagrass
C. dactylon is the most valuable and important (var. aridus) in southeastern USA. They reported that
species as a forage grass for grazing and/or hay pro- giant bermudagrass yielded no more forage than com-
duction and in the development of improved cultivars mon bermudagrass and only around 55% of Coastal
using intraspecific and interspecific hybridizations. bermudagrass (Cooper and Burton 1965). Var. dacty-
However, according to Harlan’s evaluation, four vari- lon is highly important as a herbage grass as it is so
eties in C. dactylon, vars. afghanicus, coursii, elegans, widely distributed and has been used to develop
and polevansii, are of relatively minor value as herb- numerous improved cultivars for forage use. Harlan
age grasses, as they are endemic when compared to (1970) stated that the release of Coastal in 1943 repre-
var. dactylon and aridus (Harlan 1970). Plants of var. sented a hallmark in the records of plant breeding.
afghanicus are relatively robust, vigorous, and winter Coastal, yielding 2–4 times the biomass yield of nat-
hardy but are endemic to the lowland and moist sites in uralized common strains, changed the attitude towards
Afghanistan (Harlan 1970). Var. coursii and polevansii bermudagrass among the southern row crop farmers
are endemic to Madagascar and South Africa, respec- around the middle of twentieth century (Harlan 1970).
tively, containing no germplasm attributes that will More efforts have been initiated on the forage bermu-
likely be of importance in breeding new forage culti- dagrass breeding since then, and consequently more
vars (Taliaferro 1986). Var. elegans is indigenous in cultivars have been released and used in commercial
southern Africa and relatively common in the south of production in the USA (Table 4.4).
latitude 13
S. Plants of var. elegans have good yield
potential but have moderate to low winter hardiness
4.3.2 Germplasm Collection and
(Taliaferro 1986). The development of Coastal having
var. elegans as a parent and “Midland 99” and “Bra- Conservation
zos” both having genetic contributions from var.
afghanicus indicates that vars. elegans and afghanicus The availability of a genetically diverse germplasm
are of high valuable potential in breeding new forage collection is often key to the success of a breeding
4 Cynodon 61

Table 4.4 Forage bermudagrass cultivars released in the USA [after Taliaferro et al. (2004a)]
Cultivar Year of release Origin References
Coastal 1943 USDA-ARS and Ga. AES Myers (1951), Burton
(1948, 1954)
Midland 1953 USDA-ARS and Okla. AES Hein (1953), Harlan
et al. (1954)
Suwannee 1953 USDA-ARS and Ga. AES Burton (1962)
Greenfield 1954 Okla. AES Elder (1955)
NK-37 1957 Northrup, King & Co Hanson (1972)
Alicia Mid-1960s Cecil Greer Grass Farm, Edna, TX Taliaferro et al. (2004a)
Hardie 1974 Okla. AES Taliaferro and
Richardson (1980)
Callie 1974 Miss. AES Watson (1974)
Tifton 44 1978 USDA-ARS and Ga. AES Burton and Monson
(1978)
Guymon 1982 Okla. AES and USDA-ARS Taliaferro et al. (1983)
Tifton 78 1984 USDA-ARS and Ga. AES Burton and Monson
(1988)
Grazer 1985 La. AES and USDA-ARS Eichhorn et al. (1986)
Cheyenne 1989 Jacklin Seed Co. Samudio and Brede
(1998)
Russell 1994 Ala. and La. AESs Ball et al. (1996)
Florakirk 1994 Fla. AES Mislevy et al. (1999)
Wrangler 1999 Johnston Seed Co. Taliaferro et al. (2004a)
Midland 99 1999 Okla., Kan. Mo., Ark. AESs, Noble Foundation, Taliaferro et al. (2002)
and USDA-ARS
Goodwell 2007 Okla. AES Wu et al. (2008)

program. Genes influencing a range of performance characterized, but their limited distributions and pop-
traits can often be found in relatives of commercial ulation sizes likely make them vulnerable, certainly
cultivars or the breeding populations from which those more so than in C. dactylon.
cultivars originated. Though giant strides have been Morphological and genetic diversity of C. dactylon
made in Cynodon breeding, the scope of the breeding var. dactylon is well documented (Harlan and de Wet
effort and the degree to which the total germplasm 1969; Harlan 1970; Harlan et al. 1970a; Wu et al.
base has been utilized are small. According to Harlan 2007). Native collections of C. dactylon from China,
(1970), the most extensive collections of Cynodon Korea, and Turkey studied by Wu et al. (2006a, b,
germplasm in East Africa were assembled at the Kitale 2007), Kang et al. (2008), and Gulsen et al. (2009),
Research Station in Kenya from about 1951 to 1962 by respectively, all provided results indicating tremen-
A.V. Bogdan. Bogdan described his and other’s work dous genetic variation within the taxon. Additional
with herbage Cynodon germplasm in Tropical Pasture reports indicating substantial variation within C. dac-
and Fodder Plants (Bogdan 1977). The collections of tylon var. dactylon, indigenous to China, were made by
Bogdan included C. nlemfuensis, C. plectostachyus, Liu and colleagues at the Institute of Botany, Nanjing
and C. aethiopicus. Other significant stargrass collec- (Liu et al. 2003), by Abulaiti and colleagues at Xing-
tions were housed at the Frankenwald Botanical jiang Agricultural University (Abulaiti et al. 2003),
Research Station in South Africa and the Henderson and by Wu before 2001, and now Zhang and collea-
Research Station in Harare, Zimbabwe (Taliaferro gues at Sichuan Agricultural University (Wu et al.
et al. 2004a). However, very little information is avail- 2001; Yi et al. 2008).
able on the magnitudes of phenotypic and genetic Cynodon plants may have been introduced to the
diversity in the three East African Cynodon species. Americas soon after the discovery of the new world by
Although endemic to Africa, each of the species is not Columbus in 1492 (Taliaferro et al. 2004a). The first
widely distributed in natural environments. Genetic written report of bermudagrass in the contiguous USA
erosion within the respective species has not been was made in 1751 by the then Georgia Governor
62 Y. Wu

Henry Ellis as noted by Kneebone (1966) and Burton (NPGS) of the United States Department of Agricul-
and Hanna (1995). By the early 1800s, bermudagrass ture (USDA) as indicated by the Germplasm Re-
had become one of the most important grasses in the sources Information Network (GRIN) database
South (Taliaferro et al. 2004a). Today, bermudagrass records as of June 2009 (GRIN 2009). Compared to
is the major warm-season perennial used in the south- the number of accessions in the collection in 1995, the
ern US for forage and turf, and it has spread to the current USDA NPGS Cynodon collection contains
central and northern states. However, organized col- 22% fewer accessions. Among the US Cynodon col-
lections of Cynodon germplasm for cultivar develop- lection, 379 accessions can be listed as forage germ-
ment and genetic research in the USA occurred in the plasm (Table 4.5) as discussed above. C. aethiopicus,
twentieth century, more intensively after the1930s. In C. nlemfuensis, and C. plectostachyus are represented
the bermudagrass breeding program initiated by Glenn by one, three, and 32 accessions, respectively, while
Burton in 1936 at the Coastal Plain Experiment Station C. dactylon is represented by 318 accessions. It is
of the Crops Research Division, USDA-ARS in Tif- evident that the national collection does not contain
ton, GA, bermudagrass and stargrass germplasm from ample genetic diversity in the East African Cynodon
Africa and other countries were assembled via collec- species. Of the 318 C. dactylon accessions, 272 belong
tion trips, gifts, and exchange (Burton 1951). The to var. dactylon and have been collected from 36
efforts of Burton and subsequently Wayne Hanna countries. C. dactylon varieties afghanicus, aridus,
and Bill Anderson and their colleagues have accumu- coursii, elegans, and polevansii are represented by 1,
lated a worldwide collection of more than 600 forage 1, 5, 2, and 4 accessions, respectively, and accessions
bermudagrass accessions (Anderson 2005). The col- from each taxon originated from only one country.
lection is highly genetically diverse. It has contributed Even with C. dactylon var. dactylon, the potential to
to the development and release of numerous hybrid increase the diversity by more comprehensive germ-
cultivars over the last half century and continues to plasm collection is large. The data further indicate
be used for improvement in forage traits. Anderson that comprehensive germplasm collection in major
(2005) selected a core collection from the large forage geographic regions is required to sample the full
bermudagrass accessions using phenotypic data sam- extent of the available variation. In both South and
pled in the field in cluster analysis. The core collection North Americas, introduced Cynodon plants have
was to be used for the assessment of chemical compo- spread over wide geographic expanses and have
sition or stress tolerance. become naturalized to specific climatic and edaphic
Jack R. Harlan, Wayne W. Huffine, J.M.J. de Wet, conditions. Consequently, genetic diversity has
and colleagues assembled a worldwide bermudagrass increased in these populations due to gene recombi-
collection at the Oklahoma State University (OSU) in nation, gene mutation, new gene generation, and natu-
the late 1950s and early 1960s by making germplasm ral selection under diverse environments since and
collection trips to Africa, Asia, Australia, and southern during their introduction and dispersion, respectively.
Europe (Harlan and de Wet 1969). The collection Thus, germplasm collected from these new world
contained about 700 accessions and was used in an environments is of potential value to genetic enhance-
extensive and comprehensive biosystematic investiga- ment efforts. Certainly, there should be effort to
tion of Cynodon (Wu and Taliaferro 2009). Charles M.
Taliaferro initiated and directed the bermudagrass
breeding program at OSU from 1968 to 2006 in the Table 4.5 Forage Cynodon collections in the US National Plant
Germplasm System by species and origin as of June 2009
development of forage and turf bermudagrass culti-
Species Accessions Continental origin
vars. He and colleagues added more Cynodon acces-
C. aethiopicus 1 Africa
sions from Africa, Australia, Europe, and Asia to the
C. dactylon 318 Africa, Asia, Australia,
OSU germplasm pool by Harlan (Taliaferro et al. Europe, N. America,
2004b). The OSU bermudagrass germplasm has been S. America
used to develop and release numerous elite forage and C. nlemfuensis 3 Africa, N. America
turf cultivars by Taliaferro and colleagues. C. plectostachyus 32 Africa, Asia
A total of 435 Cynodon accessions are currently C. species 25 Africa, Asia, Australia
Total 379
maintained in the National Plant Germplasm System
4 Cynodon 63

increase the sample size of Cynodon taxa now poorly the United States, as cited by Tracy (1917), as one of
represented in the NPGS collection, but even the num- the most important grasses in the southern States.
ber of C. dactylon var. dactylon accessions in the Tracy believed bermudagrass was a native grass in
collection likely poorly samples the total germplasm the “Old World,” probably India. Tracy (1917) specu-
pool of that taxon. lated that bermudagrass probably was introduced to
Preservation and maintenance of a large bermuda- the American colonies by way of the Bermuda Islands
grass germplasm collection is essential to a breeding (Tracy 1917). Later, Burton in 1951 noted that
program and provide research materials for scientific Thomas Spalding of Sapaloe Island documented in
efforts from basic to applied investigations. Widely his diary that Henry Ellis, the then Governor of Geor-
used techniques for the preservation include mainte- gia, introduced bermudagrass to Savannah, Georgia, in
nance of living clonal plants in the greenhouse or field 1751. But Spalding did not indicate where Henry
germplasm nursery, or both, and storage of seed sam- Ellis obtained the bermudagrass from (Burton 1951).
ples in a dry and cold room at 5–10
C. Viability of Burton (1951) assumed that bermudagrass came to
seed can be maintained for a long term if stored at Americas much earlier than 1751, probably in the
18
C or lower. However, genetic preservation of hay brought by the Spanish conquistadors to feed
clonal germplasm requires daily care of potted plants, their horses. He believed Africa was the primary cen-
such as watering, trimming, and fertilizing in the ter of origin as the bermudagrass introductions from
greenhouse, or careful management of individual Africa showed greater diversity.
plots in the field to avoid contamination due to Harlan et al. (1970b) noted that Africa has more
encroachment of plants from adjacent plots or by Cynodon species than other continents (Table 4.6).
dissemination of seed from plot to plot. Herbicides Clayton et al. (2009) reported that there are seven
such as glyphosate are effective in maintaining alleys Cynodon species in the African continent, surpassing
between neighboring plots. More challenging is to the number found on all other continents. Cynodon
control the contamination from volunteer seedlings diversity at species level is obviously highest in Africa
derived from seed produced on the plant through and appears to be second highest in Asia, third in
outcrossing. One way to reduce or eliminate seed Australia, and least in Europe. Beard and Watson
set is mowing the plants before seed is produced on (1982) stated that lower East Africa is generally recog-
the plant. Alternative forms of preservation techni- nized as the center of origin of bermudagrass.
ques have been studied. Reed et al. (2005) reported a Cosmopolitan tetraploid C. dactylon var. dactylon
cryopreservation protocol for storing bermudagrass was likely derived from diploid progenitors (Wu and
shoot tips in liquid nitrogen (196
C). The protocol Taliaferro 2009). Diploid Cynodon taxa include
is most effective when combined with a 1–4 weeks C. aethiopicus, C. barberi, C. dactylon var. afghanicus,
cold acclimation period and dehydration to 19–23% C. dactylon var. aridus, C. incompletus, C. plectosta-
moisture before shoot tips are exposed to liquid nitro- chyus, and C. transvaalensis. Collectively, those
gen (Reed et al. 2005). The protocol was used to diploids are distributed in a range from South Africa,
store 25 bermudagrass germplasm accessions at the to East Africa, then Eastern Mediterranean region, and
National Clonal Germplasm Repository, Corvallis, then West and South Asia (Fig. 4.1). Among the dip-
Oregon, and at the National Center for Germplasm loid Cynodon species, C. aethiopicus, C. barberi,
Resources Preservation in Fort Collins, Colorado.

Table 4.6 Continental distribution of native Cynodon species

Continent Number Species name
4.4 Origin and Evolution of species
Africa 7 C. aethiopicus, C. dactylon,
C. incompletus, C. x magennisii,
The geographic center of origin of common bermuda- C. nlemfuensis, C. plectostachyus,
grass [C. dactylon (L.) Pers. var. dactylon] has long C. transvaalensis
been of scientific interest because the grass is so Asia 3 C. arcuatus, C. barberi, C. dactylon
widely distributed. About 200 years ago, bermuda- Australia 2 C. arcuatus, C. dactylon
Europe 1 C. dactylon
grass was recorded in Mease’s Geological Account of
64 Y. Wu

C. nlemfuensis, C. plectostachyus, and C. incompletus oped rhizomes and its natural distribution ranges from
may not contribute to the formation of rhizomatous South Africa, Zambia, Tanzania, Israel, to India and
tetraploid C. dactylon var. dactylon in any way due to Sri Lanka. The contributions of var. aridus to the
the absence of rhizomes in the five species. The formation of tetraploid var. dactylon are basic (Harlan
research of Harlan and colleagues indicated that dip- and de Wet 1969).
loid C. aethiopicus, C. barberi, C. nlemfuensis vars. Harlan and de Wet (1969) believed that introgres-
nlemfuensis and robustus, and C. plectostachyus have sion between species was not significant in the evolu-
limited or no hybridization potential with C. dactylon tion of C. dactylon var. dactylon. Vars. aridus and
var. dactylon, further indicating that these likely did afghanicus are the only two diploid forms within
not contribute to the formation and diversity of tetra- C. dactylon. As mentioned above, var. aridus is rhizo-
ploid C. dactylon var. dactylon. Harlan and de Wet matous, relatively widely distributed, and ranges from
(1969) summarized that C. dactylon is completely small plants in India to tall plants in South Africa,
isolated genetically from C. barberi and C. plectosta- while var. afghanicus is endemic and has no rhizomes.
chyus and that it is difficult to cross C. dactylon with Presumably, var. aridus or its direct ancestor is the
C. aethiopicus. Harlan and de Wet (1969) also diploid progenitor of the cosmopolitan tetraploid var.
believed that C. nlemfuensis was an unlikely contribu- dactylon, and var. afghanicus also contributed to the
tor to the variability of C. dactylon. Diploid C. trans- great variability in var. dactylon (Harlan and de Wet
vaalensis can readily hybridize with C. dactylon 1969) (Fig. 4.2).
artificially and in nature (Harlan and de Wet 1969). According to its variation patterns, morphological
But the species may not have contributed much to the variation, adaptation, geographic distribution, and pos-
formation of C. dactylon var. dactylon and its enor- sible genetic origin, the cosmopolitan var. dactylon was
mous diversity since the South African species is subdivided into tropical, temperate, and seleucidus
narrowly endemic in distribution. However, C. dacty- races by Harlan and de Wet (1969) (Fig. 4.2). The
lon var. aridus is different. The taxon has well devel- tropical race presumably originated from diploid

Fig. 4.1 Geographic distribution of diploid Cynodon species

4 Cynodon 65

Union of
unreduced
gametes
Tropical race of var.
Var. aridus dactylon (4x = 36)
(2x = 18)
Hexaploid var.
Union of unreduced dactylon (6x = 54)
and reduced gametes

Evolution
Union of reduced Temperate race of
gametes var. dactylon (4x = 36) Union of reduced
gametes

Diploid Triploids Pentaploid var.

Cynodon (3x = 27) dactylon (5x = 45)
ancestor Seleucidus race of var.
(2x = 18) dactylon (4x = 36)

Evolution
Union of
unreduced
gametes
Var. afghanicus
Var. afghanicus (4x = 36)
(2x = 18)

Fig. 4.2 Cynodon dactylon var. dactylon and its genetic varia- dactylon, union of reduced gametes of diploid var. aridus and
bility are derived from two putative diploid progenitors, tetraploid var. dactylon, and union of reduced gametes of
C. dactylon var. aridus and var. afghanicus, and genetic hexaploid and tetraploid var. dactylon (Harlan and de Wet
events, such as union of unreduced gametes of diploid ances- 1969; Wu et al. 2006a; Gulsen et al. 2009)
tors, union of reduced and unreduced gametes of tetraploid var.

var. aridus. Therefore, the geographic origin of tropical Kang et al. 2008). The origin of the hexaploid forms
var. dactylon might be in the region along the Indian most likely resulted from the union of reduced and
Ocean from southern India to East Africa. The temper- unreduced gametes of tetraploid parents (Wu et al.
ate race possibly was derived from evolution of tropical 2006a). Obviously, the evolutionary events in pro-
var. dactylon as it spread beyond tropical climates. The ducing hexaploid var. dactylon are internal within
seleucidus race may have originated from hybridization C. dactylon var. dactylon. Numerous hexaploid Cyno-
between temperate tetraploid var. dactylon and tetra- don plants were reported in collections from a region
ploid var. afghanicus. Tetraploid var. afghanicus likely south of the Taurus Mountains along the Mediterranean
originated directly from diploid var. afghanicus by coast in Turkey as well (Gulsen et al. 2009).
union of unreduced gametes of parent plants. Harlan
observed various intermediate forms between temper-
ate var. dactylon and var. afghanicus in the region from
4.5 Cynodon Genetic Diversity Revealed
West Pakistan to Turkey, where the Seleucid Empire
existed (Harlan et al. 1970a). From this evolutionary by Molecular Markers
center, the seleucidus race spread outward and merged
into a temperate race in Europe (Harlan and de Wet DNA molecular markers, neutral to environment, have
1969). More recently, hexaploid cytotypes of the var. been extensively used to measure genetic diversity
dactylon were reported in the southeast part of China of Cynodon species. Assefa et al. (1999) reported
and the southern region of Korea (Wu et al. 2006a; high genetic diversity in the genus using DNA
66 Y. Wu

amplification fingerprinting (DAF) to assess molecular gene flow between natural populations, which likely
marker polymorphisms of eight species and seven prevents formation of distinctly differentiated genetic
varieties, including C. aethiopicus, C. arcuatus, groups. Although morphologically, hexaploids and
C. barberi, C. dactylon var. dactylon, C. dactylon pentaploids were similar to tetraploids in the Chinese
var. afghanicus, C. dactylon var. aridus, C. dactylon C. dactylon collection, genetic differentiation among
var. coursii, C. dactylon var. elegans, C. incompletus the three ploidy levels was discernable but not fully
var. incompletus, C. nlemfuensis var. robustus, C. plec- separate. The close genetic relatedness of the three
tostachyus, and C. transvaalensis. Of 539 bands scored, different cytotypes indicates that they have a recent
92% (496) bands were polymorphic, indicating enor- common ancestry.
mous genetic diversity in Cynodon at the genomic Kang et al. (2008) reported genetic diversity of 40
DNA sequence level. Respective accessions of C. dacty- Korean bermudagrass (Cynodon spp.) accessions as
lon var. dactylon and C. dactylon var. afghanicus were assessed using the AFLP marker system. They scored
clustered in separate groupings suggesting higher genetic 2,256 bands generated from PCR reactions of 29
diversity in the two varieties while other accessions from selective primer combinations. Among the scored
the same taxa were generally clustered. They reported AFLP markers, 87.8% (1,982) were polymorphic.
accessions within taxa, differing in ploidy (2x vs. 4x) Genetic diversity was high in the Korean Cynodon
clustered in all instances indicating that diploid and accessions as evidenced by their genetic similarity
tetraploid forms are closely related. coefficients ranging from 0.42 to 0.94. As the tri-
Using amplified fragment length polymorphisms ploids included in the Korean collection likely are
(AFLP), Wu et al. (2004) assessed genetic diversity introduced, the true genetic diversity of indigenous
and relatedness among 28 disparate C. dactylon var. Korean bermudagrass could be much lower. There
dactylon accessions originating from 11 countries were no native diploid Cynodon reported in Korea
(Australia, Bulgaria, China, Germany, France, Italy, (Kang et al. 2008). Genetic separation of tetraploids,
Japan, South Africa, Spain, Zimbabwe, and the United pentaploids, and hexaploids did not occur in the
Arab Emirates) on 4 continents. Of the 590 bands AFLP marker data, though their genome sizes were
scored, 75% (443) were polymorphic. Genetic similar- significantly different.
ity coefficients ranged from 0.53 to 0.98 for the 28 Gulsen et al. (2009) reported genetic diversity in
accessions. Accessions originating from Australia, 182 Turkish Cynodon accessions and its variance par-
Asia, Africa, and Europe were placed in distinct titioning for ploidy, geographic region, and province.
groups, indicating that geographic origin was a signif- The Cynodon accessions were genotyped by combina-
icant factor in their genetic differentiation. This sug- tional use of four molecular marker systems, encom-
gests that the genetic isolation of plant populations passing sequence related amplified polymorphism,
would provide opportunities for the respective popula- peroxidase gene polymorphism, intersimple sequence
tions to be further genetically differentiated by selec- repeat, and random amplified polymorphic DNA. The
tive forces for adaptation to specific environments. accessions were considerably diverse and their genetic
Later, their expanded AFLP study continued on a similarity coefficients ranged from 0.50 to 0.98. Major
C. dactylon var. dactylon collection of 119 accessions portions of the genetic variation in the large collection
from 11 provinces in China (Wu et al. 2006a). Among resided within ploidy level (91%) and provinces
763 scored AFLP bands generated by 13 primer com- (94%), respectively. Genetic differentiation was not
binations, 61.1% (466) were polymorphic. Their significant among ploidy levels. Similarly, genetic
results indicated the Chinese tetraploid accessions variation pattern in adjacent regions was not signifi-
contained much higher genetic variation than hexa- cant as well. The results were basically consistent with
ploid accessions while pentaploids had very those of Wu et al. (2006a). Interestingly, several dip-
low genetic diversity. Though the accessions from loid Cynodon accessions existed in the Turkish collec-
the same or nearby regions tended to cluster, genetic tion of Gulsen et al. (2009) but not in the Chinese and
differentiation among accessions from distinct regions Korean collections by Wu et al. (2006a) and Kang
was not evident. The results may be associated with the et al. (2008), respectively.
cross-pollination and self-incompatibility reproductive Karaca et al. (2002) assessed genetic diversity for
behavior of bermudagrass. Cross-pollination results in some released forage bermudagrass cultivars and
4 Cynodon 67

related selections using four molecular marker systems the use of clonal bermudagrass cultivars. Vegetative
including AFLP, chloroplast-specific simple sequence reproduction in bermudagrass maximized profitability
repeat length polymorphism, random amplified poly- of the forage industry for cattle (Bos taurus) by using
morphic DNA, and directed amplification of mini- the very best individual genotypes for farmers and
satellite-region DNA. Among a total of 1,423 DNA ranchers to plant for grazing and hay (Hanna and
fragments scored, 472 (33%) were polymorphic, indi- Anderson 2008). Burton (1956) clearly indicated
cating low genetic diversity in the forage cultivars. The using the F1 hybrids provided the means to take advan-
narrow genetic base of forage bermudagrass cultivars tage of the heterosis exhibited by superior plants in
was also evidenced by the high genetic similarity coef- segregating populations. F1 hybrids are fixed in gen-
ficients ranging from 0.608 to 0.977. Comparing the etic make up, if they are reproduced clonally.
genetic diversity of the forage bermudagrass cultivars The systematic interspecific hybridization research
with the above mentioned genetic diversity in Cynodon among Cynodon species of Harlan and colleagues in
germplasm, we can see that high potential exists to the 1960s provided detailed insights and useful infor-
develop new cultivars if the naturally occurring diverse mation to guide the future development of interspe-
germplasm is used in fuller extent. cific forage bermudagrass cultivars. As discussed
above, C. aethiopicus, C. dactylon, C. nlemfuensis,
and C. plectostachyus are deemed to be valuable for-
age germplasm resources, since they are productive in
biomass. Since the remaining Cynodon species are not
4.6 Breeding Interspecific Hybrid
productive, they will not be included in the following
Cultivars discussion. Harlan et al. (1969) indicated that C. plec-
tostachyus is genetically well isolated from C. aethio-
Development of improved bermudagrass cultivars for picus, C. dactylon, and C. nlemfuensis. The conclusion
forage and turf utilization has been tremendously suc- was made on the basis of extensive hybridizations.
cessful since the 1940s. In the review report on the More than 500 inflorescences containing 18,555 spi-
50-year grassland science achievements, Nelson and kelets emasculated by hand were pollinated with pol-
Burns (2006) note that improved forage bermudagrass len sources of other Cynodon species and those
has had a dramatic effect on the livestock industry in produced just two presumptive hybrid plants (Harlan
the southern USA. In a sister report of the 50-year et al. 1969). Therefore, the potential of using C. plec-
turfgrass research in the USA, Shearman (2006) indi- tostachyus in an interspecific breeding program would
cates that improved turf bermudagrass is the most be quite low.
widely grown warm-season turf grass internationally. The latter C. aethiopicus, C. dactylon, and C. nlem-
The breeders have extensively and effectively used fuensis species are genetically related but have varying
interspecific and intraspecific crosses to create segre- hybridization barriers, which, however, do not
gating populations for selection and evaluation. For completely prevent intercrossing among them. In the
forage cultivar development, elite parents from hybridization research of Harlan et al. (1969), only
C. dactylon and C. nlemfuensis have been used in five hybrid plants were obtained from numerous
interspecific crosses. Similarly, selected plants of crosses between a diploid C. aethiopicus and a tetra-
C. dactylon and C. transvaalensis have been routinely ploid C. dactylon. They reported six hybrid plants
used in interspecific crosses to develop interspecific from two pollinated inflorescences between a tetra-
hybrid turf bermudagrass cultivars. ploid C. aethiopicus and diploid C. nlemfuensis. The
Bermudagrass can reproduce asexually and sexu- research indicates the great difficulty in crossability of
ally. Vegetative propagation of bermudagrass is rea- C. dactylon with C. aethiopicus but relative ease with
lized using its shoot cuttings, stolons, rhizomes, or C. nlemfuensis. However, the crossability between
combinations of those. In the turf industry, bermuda- diploid and tetraploid C. nlemfuensis and tetraploid
grass sod is an important asexual form used in the C. dactylon is relatively high. Crosses of 91 emascu-
planting of improved vegetatively propagated turf ber- lated inflorescences of diploid C. nlemfuensis var.
mudagrass cultivars. Over the years, equipment has nlemfuensis pollinated with pollen of tetraploid
been invented and techniques developed to facilitate C. dactylon produced 78 hybrid plants, and crosses
68 Y. Wu

of 124 inflorescences of both diploid and tetraploid C. Kenya, in 1958 (Burton 1972). PI 255445 was identi-
nlemfuensis var. robustus with tetraploid C. dactylon fied as C. nlemfuensis var. robustus by Harlan (1970)
produced 126 hybrid plants, indicating weak crossing (Taliaferro et al. 2004a). “Tifton 85” bermudagrass is
barriers between the two taxa (Harlan et al. 1969). a high yielding and highly digestible hybrid cultivar
Their study indicated that diploid interspecific hybrids derived from a cross of C. dactylon var. dactylon PI
between the two species were very vigorous and pro- 290884 by a C. nlemfuensis selection, “Tifton 68”
ductive. The substantial value of C. nlemfuensis in (Burton et al. 1993). Tifton 85 produced 26% more
interspecific hybridization with C. dactylon has been dry matter that was 11% more digestible than Coastal
clearly demonstrated in the development of several bermudagrass in two replicated small plots tests over 3
cultivars, most notably “Coastcross 1” and “Tifton years (Burton et al. 1993). Hill et al. (2001) noted
85”. Tifton 85 has largely supplanted Coastal as the lower concentrations of either-linked ferulic acid in
cultivar of choice in the southern USA (34
S lati- Tifton 85, explaining the higher digestibility of Tifton
tude) and is being extensively used in many countries 85 than of Coastal hays. Higher dry matter yield and
in mild climatic areas. greater forage digestibility of Tifton 85 than many
Released forage cultivars of bermudagrass (C. dac- other forage bermudagrasses allows increased stock-
tylon), stargrass (C. nlemfuensis, C. aethiopicus, and ing rates and better body weight gain per hectare. The
C. plectostachyus) and interspecific hybrids were forage cultivar has received increased acceptance for
derived from ecotype selection and organized breed- hay production and pastures in the southern US and
ing procedures (Taliaferro et al. 2004a). “Coastal” been grown on more than 1 million ha in Brazil (Hill
bermudagrass, the most widely used forage bermuda- et al. 2001). The same breeding protocol using crosses
grass cultivar, was developed from the bermudagrass between C. dactylon by C. nlemfuensis was and will be
breeding program by Burton at the USDA-ARS, used to develop nutritive-value-enhanced bermuda-
Coastal Plain Experiment Station in Tifton, GA. grass cultivars.
“Coastal” produced more than twice as much forage
as naturalized common bermudagrass (Myers 1951).
The performance (body weight gain) of animals
4.7 Recommendations for Future
grazing bermudagrass including Coastal and other
cultivars clearly indicated their high production poten- Actions
tial per unit area but relatively low potential on an
individual animal basis due to relatively low nutritive Plant germplasm is among the most precious assets on
value (Elder and Murphy 1961; McCormick et al. the earth and should be conserved for the future gen-
1964). Forage dry matter digestibility is significantly erations. Bermudagrass is among the major perennial
correlated with body weight gains on pasture, with warm-season forage grasses in the world. Continuing
correlation coefficient being 0.797 (McCullough and efforts in the genetic improvement of bermudagrass
Neville 1959). Burton et al. (1967) reported that ber- and related stargrass will be greatly facilitated by a
mudagrass can be improved for dry matter digestibil- much larger, better evaluated, and carefully preserved
ity by using interspecific crosses between C. dactylon germplasm collection. Individual research programs
with highly digestible C. nlemfuensis. The best F1 have collected and used Cynodon germplasm in scien-
hybrid derived from “Coastal” bermudagrass by tific studies and for breeding, but such programs are
“Kenya 56 #14” (PI 255445) released as “Coast- generally not charged with, or funded for, long-term
cross-1” had 12.3% better dry matter digestibility preservation of the germplasm. Effective collection,
than “Coastal” while both had similar forage yields evaluation, and long term preservation can best be
(Burton et al. 1967; Burton 1972). Consequently, accomplished by agencies like the USDA NPGS,
results from a 3-year grazing study indicated that which has those dedicated responsibilities. However,
average daily gain of steers on “Coastcross-1” bermu- inadequate funding has limited the ability of such
dagrass was 29% more than average daily gain of agencies to fulfill their mandate, especially for crops
steers on “Coastal” bermudagrass (Burton 1972). with lesser value than the major food crops. Forty
“Kenya 56 #14” (PI 255445) was obtained from A. years ago, Harlan noted that only a small fraction of
V. Bogdan of Grassland Research Station, Kitale, the Cynodon germplasm pool had been studied and
4 Cynodon 69

even less used in plant improvement programs (Harlan Burton GW (1951) The adaptability and breeding of suitable
1970). Taliaferro recently noted that the extent of grasses for the southeastern States. In: Norman AG (ed)
Advances in agronomy, vol 3. Academic, New York,
Cynodon germplasm collected, studied, and used in pp 197–241
breeding new cultivars remains basically the same as Burton GW (1954) Coastal Bermuda grass for pasture, hay and
many years ago (Taliaferro et al. 2004a). This clearly silage. Bull N.S. 2. Georgia Coastal Plain Experiment Sta-
indicates the need of greater efforts in collecting, tion, Tifton, GA
Burton GW (1956) Utilization of heterosis in pasture plant
evaluating, and preserving Cynodon germplasm at breeding. In: Neale GJ (ed) Proceedings of the 7th interna-
local, national, and international levels. Governments tional grassland congress, Palmerston North, NZ, 6–12 Nov
and their agencies should make larger and sustained 1956, Wellington, New Zealand, pp 3–12
investments in support of Cynodon germplasm research. Burton GW (1962) Registration of varieties of bermudagrass.
Crop Sci 2:352–353
As Cynodon forage cultivars are used in many countries Burton GW (1972) Registration of ‘Coastcross-1’ bermuda-
and Cynodon forage germplasm is not distributed grass. Crop Sci 12:125
evenly across the world, international collaborations Burton GW, Hanna WW (1995) Bermudagrass. In: Heath ME,
are necessary to facilitate collection and exchange of Barnes RF, Metcalfe DS (eds) Forages: the science of grass-
land and agriculture. Iowa State Univ Press, Ames, IA,
Cynodon germplasm, communication of research and pp 421–429
breeding information, and distribution of improved Burton GW, Monson WG (1978) Registration of ‘Tifton 44’
cultivars. bermudagrass. Crop Sci 18:911
Burton GW, Monson WG (1984) Registration of ‘Tifton 68’
bermudagrass. Crop Sci 24:1211
Burton GW, Monson WG (1988) Registration of ‘Tifton 78’
Acknowledgments The author dedicates the humble chapter to bermudagrass. Crop Sci 28:187–188
his mentor, Dr. Charles M. Taliaferro, Regents Professor Emer- Burton GW, Hart RH, Lowrey RS (1967) Improving forage
itus of the Oklahoma State University, and also thanks him for quality in bermudagrass by breeding. Crop Sci 7:329–332
his review and comments for improving the manuscript. Burton GW, Gates RN, Hill GM (1993) Registration of ‘Tifton
85’ bermudagrass. Crop Sci 33:644–645
Casler MD, Heaton E, Shinners KJ, Jung HG, Weimer PJ,
Liebig MA, Mitchell RB, Digman MF (2009) Grasses and
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.
Chapter 5
Dactylis

Alan V. Stewart and Nicholas W. Ellison

5.1 Introduction fourth most widely used grass genus with 3.3% of
the world’s temperate seed, following Lolium, Festuca
and Phleum (Bondesen 2007).
The genus Dactylis consists of a single perennial
It is almost never used for turf despite fine dense
forage species, Dactylis glomerata L. known as orch-
turf forming types being available. This may be
ardgrass or cocksfoot. The basic chromosome number
because of the limitations imposed by seed yield and
is 2n ¼ 14 and, although the species consists of
subsequent seed price as well as suitable cultivars.
diploids, tetraploids and a hexaploid, the commonly
used forage form is tetraploid.
Dactylis species have only rarely been crossed with
species outside this genus (Nakazumi et al. 1997) and 5.3 Basic Botany of the Species
the genus is not closely related to any major crop.
Therefore, the story of Dactylis wild relatives is 5.3.1 Morphology
largely one of the diversity within the genus.
Although the distribution and diversity of Dactylis
is complex, much of this can be readily explained by D. glomerata is in general a perennial bunchgrass.
its evolutionary, migrational and genomic history over Forms adapted to more humid habitats usually have
the last few hundred thousand years. large tillers and broad leaves while those adapted to
The understanding of the diversity within the wild drier conditions exhibit a more xeromorphic form with
relatives of Dactylis has considerable implications for a greater number of smaller tillers with narrower glau-
germplasm collection and conservation as well as for cous leaves. Some forms growing on coastal cliffs
the breeding techniques employed. There is consider- have branching stems, which sprawl along the ground,
able urgency to make sure that all the geographically even hanging in some situations.
diverse diploid subspecies are collected and repre-
sented in genebanks.
5.3.2 Taxonomy of D. glomerata L
5.2 Agricultural Status
As a genus Dactylis is subject to differing taxonomic
D. glomerata is an important forage grass in the tem- interpretations in different regions of its natural range
perate areas of the world. Each year approximately (Europe, Asia, North Africa and the Canary Islands).
14,000 tons of seed is harvested making it the There are no modern taxonomic treatments which
interpret all forms on the same basis. Here we interpret
D. glomerata to be monotypic consisting of one
A.V. Stewart (*)
diverse species complex (Jogan 2002). The genus
PGG Wrightson Seeds, PO Box 175, Lincoln, Christchurch
7640, New Zealand however, is clearly on the verge of speciation with
e-mail: [email protected] many diploid, tetraploid and a hexaploid subspecies

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 73
DOI 10.1007/978-3-642-14255-0_5, # Springer-Verlag Berlin Heidelberg 2011
74 A.V. Stewart and N.W. Ellison

altaica
28 ITS
1 cp
Lamarckia aurea
G 510 T C 90 T

A 613 G

C 55 T

A 425 G G 92 C judaica

A 571 G
lusitanica
A 601 G T 190 C
G 475 T
592 T insert
C 649 A
santai
C cp444 T ibizensis sinensis parthiana himalayensis
aschersoniana
castellata cpDNA
G 610 A smithii Del 287
izcoi C 627 T
juncinella mairei woronowii

Fig. 5.1 Molecular descent in diploid Dactylis glomerata using ITS and trnL intron chloroplast sequences (Numbers refer to
mutations in the ITS or chloroplast (cp) sequences)

and some reduction in fertility when different diploid hypothesis that the diploid progenitor originated in,
forms are crossed (Borrill 1977). and migrated out from Central Asia (Fig. 5.1). This
progenitor is probably very similar to subsp. altaica.
We have observed no more than 2–7 ITS mutations
5.3.2.1 Diploid D. glomerata Forms and only 0 or 1 trnL intron chloroplast mutation in any
of the diploid Dactylis lineages and this allows an
Today almost all diploid populations have restricted estimate of timing of the original divergence. The
ranges, indicative of a remnant distribution from a rate at which ITS and chloroplast sequences mutate
former widespread range. The populations are usually depends on many factors including effective popula-
described as subspecies, or sometimes even species, tion size, annual or perennial habit, and the breeding
and publications vary as to their interpretation. Here system. In annual grasses, such as barley and maize,
we will not enter into any taxonomic debate but rec- the ITS mutation rate has been calculated to be approx-
ognize them as discreet remnant geographic popula- imately one mutation every 23,000 years (Zurawski
tions. They occur from China across to Portugal and et al. 1984), while the rate in perennial grasses is
North Africa as well as on to nearby north Atlantic slower and potentially similar to that observed in
islands such as the Canary Islands and Cape Verde. perennial Phleum, where Stewart et al. (2008) observed
the most rapidly changing lineage to be one in 30,000
years. Chloroplast genome trnL intron mutation rates
5.3.2.2 Origin of Diploids are usually 3–8 times slower, with one per 200,000
years in rice and maize (Yamane et al. 2006) and one
It is very clear that Dactylis originated as a diploid and per 90,000 years for the most rapidly changing peren-
that the polyploids have developed from them. Molec- nial Phleum lineage (Stewart et al. 2008).
ular information on diploids using the nuclear internal These rates for Dactylis would suggest that the first
transcribed spacer (ITS) and trnL intron chloroplast divergence occurred in the order of 60,000–210,000
sequences, reported here for the first time, support the years BP, a date which is far more recent by orders of
5 Dactylis 75

magnitude than the 10 million years, or more, dating glacial refugia at lower altitudes or latitudes (Hewitt
back to the tertiary period postulated by Stebbins and 1999) and allowed Dactylis to expand into the exten-
Zohary (1959) who used geological events as the sive North African grasslands.
explanation. However, it is only with the recent advent Subsp. woronowii shows a very similar flavonoid
of molecular data that the powerful influence of his- composition to aschersoniana (Lumaret 1988) and
torical glaciation events on plant migration and evolu- exhibits a molecular sequence derived from it, but its
tion have become to be fully appreciated (Hewitt morphological characteristics are more similar to
1999). This relatively recent origin for Dactylis is other xeromorphic Mediterranean types. It is likely
much more consistent with the perceived interglacial that this entity originated from the forest margin
age of many of the tetraploids (Lumaret 1986) and dwelling aschersoniana to become the first xero-
there is no reason to believe that the process of tetra- morphic form adapted to drier grassland conditions
ploidy was not initiated early in the development of during the interglacial period.
Dactylis. This time period is also consistent with the Ancestors of smithii would have easily migrated the
observation that, while there is sufficient genetic dif- 100 km from North Africa to the Canary Islands, and it
ferentiation to result in reduced fertility among crosses is likely that bird migration was responsible for the
of the diploid subspecies, there are no major structural larger 1,500 km movement to the Cape Verde Islands.
chromosomal differences (Borrill 1977). The origin of smithii from aschersoniana and/or wor-
One of the closest genera to Dactylis is Lamarckia, onowii is one of the options put forward by Stebbins.
and L. aurea diverged from it with more than 28 ITS In his opinion smithii was either a very old remnant
sequences differences and 1–2 trnlL intron chloroplast equally as old as aschersoniana, or a recent migrant
differences (Fig. 5.1). As L. aurea is an annual, these via North Africa (Stebbins and Zohary 1959). Its
differences suggest a divergence time in the order of unique morphological characteristics made him favor
200,000–750,000 years BP. the former option whereas our molecular data support
The first molecular divergence within Dactylis a more recent migration via North Africa. Sahuquillo
(Fig. 5.1) divides judaica, himalayensis and the pater- and Lumaret (1999) also provide evidence of tetra-
nal side of parthiana from the remaining diploids. ploid Dactylis migrations from North Africa to the
This is consistent with an origin in Central Asia for Canary Islands, they used chloroplast molecular mar-
Dactylis and an early geographic split leaving the kers supported by morphological, allozyme and phe-
ancestors of judaica, himalayensis and parthiana nolic indicators.
spread widely over the southern regions of Western In the post-glacial period as the climate became
Asia, and the ancestors of the remaining diploids in the warmer and drier North African grasslands contracted
north-west of this region. leaving dissected remnant populations, mairei in the
The remaining diploids exhibit a second divergence Kerrata Gorge of Algeria, santai and “castellata” in
(Fig. 5.1) with the Spanish forms lusitanica, izcoi, Algeria and Morocco, and smithii in the Canary and
juncinella and ibizensis splitting off from the group. Cape Verde Islands.
This is consistent with a migration through the Eurasian As the glaciated conditions of northern Europe
temperate forest regions from the Caucasus to Portu- contracted, the northern temperate forms could
gal. This most probably occurred during a warm inter- migrate from their glacial refugia to high altitudes or
glacial period, probably 75,000–150,000 years BP, latitudes. The European progenitor of aschersoniana
when conditions allowed continuous distribution of in particular was able to recolonize northern Europe
temperate forest in Europe. from a probable refuge in the Caucasus region, even
This European form has since migrated into North extending to the east to China as sinensis.
Africa and over to China as the molecular profiles of It seems probable also that the tetraploids also
aschersoniana, santai, “castellata”, smithii and sinen- recolonized Europe to cover northern regions so suc-
sis are identical, with both woronowii and mairei cessfully that they excluded the expansion of the
derived from this group. The lack of molecular differ- remaining diploids.
ences suggests a relatively recent migration, probably The Iberian lusitanica progenitor was able to
no older than the last glacial period. Glaciation in migrate throughout Iberia so that today we have a
Europe forced the European flora south into dissected dissected distribution with lusitanica in central Portugal,
76 A.V. Stewart and N.W. Ellison

izcoi in Galicia and juncinella in the Sierra Nevada Himalayensis

Mountains. The ancestral form also migrated to the
Balearic Isle, where ibizensis occurs today. The Ibe- D. glomerata subsp. himalayensis Domin occurs in the
rian Peninsula is composed of a complex set of differ- western Himalayas at altitudes of 1,800–4,000 m in
ent glacial refugia which, if understood, may help cool temperate forest zones.
explain the relationships between the different forms
present today (Gómez and Lunt 2006). It is also appar-
ent from hybrid ITS sequences of tetraploids in North Sinensis
Africa, the Canary Islands and Spain that many of
these forms have resulted from hybridization between D. glomerata subsp. sinensis Camus occurs in Sichuan,
North African and Spanish forms. Hubei, Guizhou, Xinjiang and Yunnan in central
It is clear also that parthiana has been influenced China, at altitudes of 1,000–3,800 m in cool temperate
by genetic introgression from overlapping woronowii forest zones.
populations. Although some authors have classified these Chinese
The probable migration of the diploids determined forms into himalayensis (Stebbins and Zohary 1959;
from the molecular research is outlined in Fig. 5.2. As Borrill 1977) the molecular differences reported here
yet we have been unable to obtain samples of hyrcana, and the genetic differences reported by Lumaret
reichenbachii, metlesicsii or smithii from Cape Verde (1987) provide considerable support to maintaining
to determine their origins. sinensis as separate from himalayensis.

5.3.2.3 The Diploid Subspecies Altaica

The following are the known diploid remnant The diploid D. glomerata subsp. altaica (Bess.)
populations: Domin occurs in the Alatau Mountains of Kazakhstan
(Czerepanov 1981; Mizianty 1991). The one sample

Fig. 5.2 Probable migration routes of diploid Dactylis based on molecular results. (black lines – before glaciation, dashed lines –
North Africa during the glaciation; dotted lines – post-glacial, Northern Europe and China)
5 Dactylis 77

that we have obtained from this region had an ITS can not discount the possibility that this genebank
sequence basal to all other diploid Dactylis forms accession of parthiana was collected in the zone of
(Fig. 5.1). No collections are available in genebanks hybridization and that those higher altitude forms
and collection must be a priority. which may be collected less frequently may be more
“pure”.

Aschersoniana

D. glomerata subsp. lobata (Drejer) Lindb.  subsp. Reichenbachii

aschersoniana (Graebn.) Thell.  D. aschersoniana
Graebn.  D. polygama Horv.  subsp. polygama D. glomerata subsp. reichenbachii (Dalla Torre et
(Horv.) Domin occurs mostly in deciduous forests from Sarnth.) Stebbins et Zohary  D. reichenbachii (Dalla
the Caucasus Mountains across central Europe, into Torre et Sarnth.) Meusel occurs on south-facing mea-
Sweden in the north, western Russia in the east, south dows on dolomite soils of the European Alps and French
into Yugoslavia and northern Macedonia. This distribu- Massif Central (Speranza and Cristofolini 1987).
tion is typical of a species recolonizing Europe from a Although we were unable to obtain a sample for
glacial refuge in the Caucasus region (Hewitt 1999). molecular analysis, flavonoid phytochemistry places
The habitats of aschersoniana and himalayensis are this between aschersoniana and lusitanica (Fiasson
similar as they both occur in cool temperate forests in et al. 1987), while isozyme data places it very close
continental climates often at high altitudes and they to aschersoniana and himalayensis (Lumaret 1988).
bear a close resemblance in isozyme allelic patterns Stebbins and Zohary (1959) note that this form is
(Lumaret 1988), flavone phytochemistry (Lumaret highly variable exhibiting characteristics of both wor-
1988), DNA content (Tuna et al. 2007) and morphol- onowii and aschersoniana, and is likely to be an
ogy, so much so that the geographic origin is often edaphic remnant of the original European temperate
necessary to correctly assign them to the appropriate forest flora.
taxa (Stebbins and Zohary 1959).
This form has potentiality to be used in agriculture
(Stuczynski 1992) and there is at least one commercial Lusitanica
cultivar, Tosca, bred in the Czech Republic (Mı́ka
et al. 1999). D. glomerata subsp. lusitanica Stebbins et Zohary is
restricted to the Serra de Sintra, and a few other
localities in central-southern Portugal (Stebbins and
Parthiana Zohary 1959). It appears to be a remnant temperate
forest species of the mildest temperate forest zone.
D. glomerata subsp. parthiana Parker et Borrill.
occurs in the moist oak woodlands on the northern
slopes of the Elburz Mountains of Iran between
1,600 and 1,900 m. At lower altitudes its distribution Iscoi
overlaps with woronowii and hybrid plants occur
(Borrill and Carroll 1969). Parker and Borrill (1968) D. glomerata subsp. izcoi Ortiz et Rodriguez-Oubina,
showed that artificial hybrids of this cross are fully has also often been referred to as “Galician diploid”.
fertile. Our molecular results provide confirmation of This temperate form occurs in Galicia, Spain, persist-
this hybridization in the only genebank accession ing in forest habitats at altitudes of 400–650 m (Ortiz
available, which combines a himalayensis type ITS and Rodriguez-Oubiña 1993; Lindner et al. 2004). It
sequence with a woronowii type chloroplast sequence. shows the greatest molecular similarity with juncinella
Similarly, flavone phytochemistry has shown that both and both must have developed from populations in the
parental pathways are present (Ardouin et al. 1985), same complex glacial refugia of the Iberian Peninsula
and electrophoretic patterns are also more variable (Gómez and Lunt 2006). The UK commercial cultivar
than other subspecies (Lumaret 1988). However, we Conrad is a diploid izcoi type (Borrill 1977).
78 A.V. Stewart and N.W. Ellison

Woronowii as do morphological comparisons, so these two proba-

bly do not deserve separate subspecies status (Lumaret
D. glomerata subsp. woronowii (Ovcz.) Stebbins et 1988; Amirouche and Misset 2007). The names, how-
Zohary  D. woronowii Ovcz. is a xeromorphic ever, may remain useful as indicators of geographic
grassland taxon in the mountain steppes and steppe origin with “castellata” from Algeria and santai from
forests in northern and north-eastern Iran and adjacent Morocco.
Turkmenistan (Stebbins and Zohary 1959). It occurs The Moroccan forms of santai show characteristics
on dry base-rich limestone outcrops in open thorn of the nearby ibizensis diploid (Mousset 2000) and this
scrub up to 1,500 m in the Elburz Mountains (Borrill is also apparent in hybrid ITS sequences which suggest
and Carroll 1969). Sympatric tetraploids are also com- introgression from nearby southern Spanish forms.
mon in this region (Doroszewska 1963).

Smithii
Hyrcana
D. glomerata subsp. smithii  D. smithii Link. occurs
D. glomerata subsp. hyrcana Tzvelev is an endemic
in the wetter sides of the Canary Islands at altitudes of
subspecies of deciduous woods in the Talish plains of
100–700 m. It has a distinctive growth habit with
Azerbaijan and Iran at 200–300 m altitude (Tzvelev
branching culms with numerous nodes (Stebbins and
1983). We were unable to obtain samples so it is
Zohary 1959). Today its habitat has reduced because
unclear how this relates to the others but its habitat
of construction in this zone. The form on the Cape
appears to be an extension of the range of woronowii
Verde Islands at 1,200 m is most probably extinct
and it is likely to be closely related to it.
(Lumaret pers. comm.). Isozyme studies group smithii
near mairei, santai and “castellata” (Lumaret 1988).
In hybridization studies there was evidence for
Mairei
cytoplasmic and nuclear differentiation in smithii
(Parker and Borrill 1968). Clearly smithii is the most
D. glomerata subsp. mairei Stebbins et Zohary is
differentiated in a range of features. Its habitat is more
confined to shaded limestone cliffs of the Kerrata
subtropical and this has influenced its flavonoid chem-
Gorge in Algeria with a relatively high annual rainfall
istry and morphology greatly (Ardouin et al. 1985). Its
of 1,100–2,000 mm (Stebbins and Zohary 1959). It is
flowering habit has changed so that it requires little
highly probable that since the last glaciation its range
vernalization for flowering and its trailing habit may
has become restricted as the North African grasslands
be due to a lack of apical dominance.
have declined (Borrill and Lindner 1971). Molecular,
isozyme (Lumaret 1988) and flavone data (Ardouin
et al. 1985) show that it exhibits close phylogenetic
affinity to “castellata”, santai, smithii and woronowii. Metlesicii
This form is an isolated remnant of a diploid wide-
spread in the North African grasslands since the last Dactylis metlesicsii Schönfelder et D. Ludwig occurs
glaciation (Borrill and Lindner 1971). at a high altitude in the Canary Islands and is poten-
tially a high altitude form of smithii (Schönfelder and
Ludwig 1996) or results from a separate migration into
Santai and “castellata” the Canary Islands. We have not been able to obtain a
sample of this form for analysis. This form is listed as
D. glomerata subsp. santai Stebbins et Zohary and an endangered species in the Spanish Red List,
subsp. “castellata”. occur in western Algeria to west- although one could question its species status.
ern Morocco at 150–1,500 m in the Tell Atlas, in Analysis of phenolic compounds showed that high
humid and subhumid bioclimates (Amirouche and altitude tetraploid forms from Gran Canary Island
Misset 2007). Flavonoid phytochemistry and enzyme were well differentiated from the lower altitude
data fail to differentiate between “castellata” and santai, forms in the region (Jay and Lumaret 1995). This
5 Dactylis 79

suggests that diploid metlesicsii may well be substan- This supports a recent post-glacial change from the
tially different from lower altitude smithii. temperate form to the more summer dormant xero-
morphic Mediterranean form. Judaica is also most
probably a relic population of the Eurasian temperate
Juncinella flora that extended into the ancient forests of Lebanon
and Israel. However, the rapid post-glacial warming
D. glomerata subsp. juncinella (Bory) Boiss occurs in and drying of the climate in this region has forced it to
the Sierra Nevada range in Spain at altitudes of adopt a xeromorphic growth pattern and morphology
2,200–2,900 m in the subalpine and alpine herb belt. in response to the Mediterranean climate. This is sup-
Its flavonoid phytochemistry exhibits similarity to ported by a heterogeneous flavone phytochemistry
lusitanica, ibizensis, and less so to izcoi (Ardouin with some individuals showing characteristics of
et al.1985). Hybridization studies suggest a close rela- “temperate” phenolics (Ardouin et al. 1985), a phe-
tionship between juncinella and ibizensis and a rea- nomenon attributed to evolutionary lag between a
sonably close relationship between smithii and temperate electrophoretical pattern (Lumaret 1984,
juncinella (Wei-Lin Hu and Timothy 1971). 1986) and Mediterranean morphology (Borrill 1977).

Ibizensis 5.3.2.4 Tetraploid D. glomerata Forms

D. glomerata subsp. nestorii Rossello et L. Sáez  Tetraploids occupy almost the full range of the species
D. ibizensis Gandoger  subsp. ibizensis (Gandoger) and consist of both autotetraploids based on single
Stebbins et Zohary. Although renamed subsp. nestorii diploid subspecies, and interecotypic hybrids based
by Castro et al. (2007), it is still commonly known as on more than one diploid subspecies. These intereco-
ibizensis in almost all of the literature. It occurs on the typic tetraploids often have a selective advantage over
Balearic Isle near Spain and is derived from the origi- their diploid counterparts.
nal Spanish forms but with some molecular introgres-
sion from nearby North African mairei. Sympatric
tetraploid forms also occur on this Island (Castro 5.3.2.5 Origin of Tetraploids
et al. 2007).
It seems likely that the process of tetraploidization was
initiated early in the development of Dactylis. Unre-
Judaica duced gametes are known to occur frequently in dip-
loid populations and gene flow is largely from diploids
D. glomerata subsp. judaica Stebbins et Zohary, and to tetraploid with triploids generally being aborted at
subsp. lebanotica. Subsp. judaica occurs in the hills of an early stage (Lumaret and Barrientos 1990; Lumaret
Israel while lebonotica occurs in Lebanon (Stebbins et al. 1992). Early tetraploids were most probably
and Zohary 1959; Apiron and Zohary 1961) and autotetraploids in the sense that they were based
potentially also in Syria. Whether these deserve taxo- upon only one diploid population and as such they
nomic differentiation is doubtful but the names at least may never have had any selective advantage over
have value as indicators of geographic origin. It has a their diploid counterparts. However, with the forced
Mediterranean summer dormant growth pattern yet the retreat of both diploids and tetraploids into confined
molecular results show that it has developed from a glacial refugia during the last glaciation, some tetra-
temperate himalayensis progenitor. The heteroge- ploids were able to benefit from gene exchange with a
neous flavone phytochemistry of judaica shows some number of ecologically divergent diploid populations.
individuals having characteristics of “temperate” phe- These interecotypic hybrid tetraploids often gain con-
nolics (Ardouin et al. 1985). This heterogeneity could siderable evolutionary success by their hybrid vigor
be related to the evolutionary lag between temperate and their enhanced genetic variation (Stebbins 1971).
patterns (Lumaret 1984, 1986) and Mediterranean According to Soltis and Soltis (1993) tetraploids have
morphology (Borrill 1977). more polymorphic loci than diploids, 0.80 compared
80 A.V. Stewart and N.W. Ellison

to 0.70, higher heterozygosity, 0.43 compared to 0.17, Stebbins also notes that as reichenbachii is similar
and a greater number of alleles per locus, 2.36 com- to woronowii in many features it is possible that some
pared to 1.51. The hybrid vigor obtained from these glomerata have developed from the hybridization of
interecotypic tetraploids readily allowed them to dom- reichenbachii and aschersoniana in the Alps where
inate the post-glacial expansion to such an extent they occur together.
that we have an almost continuous distribution of Domin (1943) notes that the primary distribution of
tetraploids over the full range of the species today subsp. glomerata is in Northern Europe but that it has
(Lumaret 1986). a secondary distribution in the temperate forest
Autotetraploids frequently have a sympatric distri- regions of Algeria and Morocco. However, in this
bution with the diploid population that they are based case it is likely that the ancestral diploids of the two
upon (Borrill and Lindner 1971; Lumaret et al. 1989) forms may be different.
while interecotypic hybrid tetraploids based on multi- Although subsp. glomerata is usually considered to
ple diploid populations are more widespread. It is be summer active and hispanica summer dormant,
commonly reported that almost all the diploid entities glomerata forms with summer dormancy are known
have a sympatric association of autotetraploids or at to occur in the Languedoc-Roussillon Mediterranean
least tetraploids derived predominantly from a single climatic region of France (Mousset 1995).
diploid. Often this coexistence is dependent upon each
occupying different ecological niches (Maceira et al.
1993). Slovenica
The taxonomically distinguishable tetraploid sub-
species include the widespread glomerata, the taller D. glomerata subsp. slovenica (Domin) Domin 
calciphilous slovenica, the xeromorphic hispanica, D. slovenica Domin occurs in the mountains of Cen-
the seaside forms of marina  hackelii and oceanica tral Europe mostly at altitudes between 600 and
and the cliffside hylodes. However, it is probably fair 1,300 m and usually on calcareous or dolomitic soils
to conclude that the tetraploid entities merge into one (Fig. 5.3; Mizianty 1997). One of the most notable
complex species where characters vary along mutu- characteristics of slovenica is its plant height which
ally independent clines (Speranza and Cristofolini can be up to 1.5 m (Domin 1943). It is likely that
1986). diploid aschersoniana has contributed to this form as
they are both tall and have a hairless keel and margin
(or few hairs) (Mizianty and Cenci 1995).
5.3.2.6 The Tetraploid Subspecies The current distribution suggests that it has spread
from a glacial refuge in the Alps or Carpathians.
Five tetraploid subspecies are recognized.

Hispanica
Glomerata
D. glomerata subsp. hispanica (Roth) Nyman 
D. glomerata L. subsp. glomerata is the most wide- D. hispanica Roth is widespread in the Mediterranean
spread tetraploid entity of temperate forest regions and region from the Iberian Peninsula to the Crimea and
is the form most widely used in agriculture. Stebbins Caucasus. It is xeromorphic exhibiting a smaller plant
and Zohary (1959) suggest that it may have developed size with a compact panicle of limited branching
from the interecotypic hybridization between ascher- (Speranza and Cristofolini 1986). It usually exhibits
soniana and woronowii, as artificial hybrids are inter- early flowering and a summer dormant growth pattern
mediate between the two and indistinguishable from which provides good drought tolerance.
subsp. glomerata. These subspecies occur in close Domin (1943) notes that hispanica is a variable
geographic proximity and both could indeed have complex with one stable feature, having lemmas
provided the very broad interecotypic tetraploids nec- divided into two blunt lobes from the middle of
essary for successful colonization of Europe from the which is a short awnlet, or a mere point which some-
Caucasus glacial refugia. times does not reach the size of an awn. This lobed
5 Dactylis 81

Fig. 5.3 Distribution of Dactylis glomerata subsp. slovenica [after Mizianty (1997)]

characteristic is less pronounced in the north where it provide the plant with an advantage under coastal con-
shows a transition towards the large growing glomer- ditions. The smooth leaf margin trait is believed to
ata plants used in agriculture. enhance palatability (van Dijk 1961) while the herbage
The summer dormant forms in the Languedoc- has superior digestibility in the order of 2–6%. For this
Roussillon Mediterranean region of France represent reason marina has attracted the attention of plant bree-
hybrids, or introgression hybrids, of hispanica and ders (Tyler and Borrill 1983) but as yet this feature has
glomerata (Lumaret 1986). not resulted in any cultivar, presumably because
A number of summer dormant cultivars developed of marina crosses have not had adequate yields.
hispanica types from Mediterranean germplasm includ- Marina is believed to have arisen from diploids
ing Uplands, Sendace, Berber, Kasba, (Australia), Jana closely associated with smithii and ibizensis (Borrill
(Italy), Medly (France), Perouvia and Chrysopigi 1961). However, it is also possible that the character-
(Greece), and germplasm of this form offer consider- istic smooth leaf margins, papillose epidermis and
able potentiality for breeding cultivars suited to Medi- glaucous appearance could arise in material of differ-
terranean climates. ent origins as well.

Marina Oceanica

D. glomerata subsp. hackelii (Asch. et Graeb.) Cif. et D. glomerata subsp. oceanica G. Guignard occurs on
Giacom.  D. marina Borrill  D. glomerata L. subsp. the coast of southern France and the Atlantic and the
marina (Borrill) Greuter  D. smithii Link subsp. Channel coasts of north-western France (Guignard
marina (Borrill) Parker is localized on sea cliffs of 1985). This form could potentially be viewed as an
the Mediterranean extending along the coast of South- extension of the range of the marina type into northern
west Europe (Portugal and Galicia) and the Atlantic Europe, however, the marina types of the Mediterra-
Islands (Borrill 1961; Fig. 5.4). nean exhibit many hispanica features while oceanica
Marina is characterized by smooth leaf margins exhibits many glomerata features.
devoid of silicified teeth, papillose epidermis and a Forms similar to oceanica have been observed along
particularly glaucous appearance, all features which the coast of Cornwall and an Irish island (Lumaret 1988).
82 A.V. Stewart and N.W. Ellison

Fig. 5.4 Distribution of Dactylis glomerata subsp. marina [after Borrill (1961)]

Hylodes 5.3.3 Climatic Races

D. glomerata subsp. hylodes P.F. Parker is confined
The development of climatic races has been part of the
to inland cliffs of Madeira, Canary Islands, and
continuing process of adaptive radiation with climatic
Cape Verde whereas marina occurs on the coastal
races adapted to drier conditions and also to warmer
cliffs (Parker 1972). This subspecies is closely
subtropical conditions. The oldest diploid forms such
related to the diploid smithii with persistent, trail-
as altaica, himalayensis and aschersoniana adapted to
ing, woody stems and papillose leaves. Parker
temperate forest margins, while their derivative wor-
(1972) noted that both hybridization and polyploidy
onowii developed xeromorphic features as it expanded
have provided a physiological versatility allowing
into the drier grasslands of western and central Asia.
this subspecies to maintain a wider distribution than
These xeromorphic features are also typical of hispa-
smithii. Our molecular results show that these tetra-
nica tetraploids in the drier Mediterranean, and
ploids have profiles from both smithii and Spanish
include smaller, more glaucous leaves and tillers as
diploids.
well as smaller inflorescences. Independently, xero-
morphic forms of judaica have developed in the Med-
iterranean from temperate pre-himalayensis types.
5.3.2.7 Hexaploid D. glomerata The seaside dwelling forms marina and oceanica
have developed glaucous characteristics and in the
Hexaploid populations have been reported from a case of marina have papillate epidermises. Those on
restricted range within Libya and western Egypt coastal cliffs (hylodes, smithii) have developed a
(Jones et al. 1961; Jones 1962). Potentially these are sprawling branching habit.
also interecotypic polyploids, but little is known of
their origin.
Two hexaploid plants are reported from the coast
near Pontevedra in Galicia, Spain but these are proba- 5.3.4 Edaphic Adaptation
bly rogue individuals within the local tetraploid rather
than a self maintaining hexaploid population (Horjales The development of edaphic races has also been part
et al. 1995). of the continuing process of adaptive radiation so that
5 Dactylis 83

today many commercial glomerata forms are noted many Islands. These resources are often explored by
for their tolerance of acidic soils with aluminum, and plant breeders but they are of lesser significance
subsp. slovenica grows on calcareous soils while than the primary pool simply because adaptation to
reichenbachii is adapted to dolomitic soils. agricultural environments is usually inferior. In gen-
eral, these resources still occur in nature and in
genebank collections. Many of the wild populations
are under threat with climate warming and human-
5.3.5 Genome Size induced habitat changes, while even those in gene-
banks will only be maintained if adequate resources
Genome size in Dactylis has been reported to decrease are available.
with altitude in France and Italy, often by as much as The secondary gene pool includes tetraploid forms
30% (Reeves et al. 1998). There is also approximately sympatric with diploid populations. The Australian
a 15% reduction in the DNA mass of diploids forms, cultivar Porto and the Spanish cultivar Adac 1 are
which have developed in southern Iberia, North Africa based on natural tetraploids with lusitanica influence.
and the Canary Islands when compared to those in A number of cultivars have been based on tetraploid
northern Iberia, Europe, Middle East and central forms of izcoi from Galicia, such as Grasslands Wana
Asia (Tuna et al. 2007). Interestingly, these are proba- (New Zealand), Cambria (UK) and Artibro (Spain). It
bly more recent in origin. is also possible that other cultivars are based on tetra-
ploid forms of diploid populations. The tetraploid
forms of many of these diploid populations represent
an enormous resource for breeders and representative
5.4 Genetic Resources collections are required. Although many have been
collected, few are identified as tetraploid forms of
For plant improvement it is appropriate to divide the remnant diploid population.
genetic resources into primary, secondary, tertiary
and quaternary gene pools as follows.

5.4.3 Tertiary Gene Pool

5.4.1 Primary Gene Pool These can be defined as those populations from other
ploidy levels where introgression into commercial
These can be defined as cultivars and elite breeding breeding programs is possible. These would largely
lines adapted to the region of agricultural use. This is consist of diploids, but potentially the hexaploids
the primary activity of most effective plant breeders could also be used for tetraploid breeding.
and consists almost exclusively of tetraploids, Breeders have crossed germplasm from diploid
although the diploids izcoi and aschersoniana have lusitanica into tetraploid material with examples
been used commercially. In general, these resources being the two UK commercial cultivars Saborto and
are well used by breeders and their “working” collec- Calder, as well as the NZ cultivar Grasslands Kara.
tions largely represent this gene pool. The largest Tetraploidy can be induced from the diploids by col-
threat to this gene pool would be any decrease in the chicine treatment as in Saborto, or via unreduced
number of breeding programs in Dactylis. gametes as in Calder (Lewis 1975).
In general, these resources are poorly represented
in genebanks and are seldom used by breeders. Yet
because of the foundation upon which modern tetra-
5.4.2 Secondary Gene Pool ploids are based, the remnant diploid populations rep-
resent an enormous resource. Many are under threat
This can be defined as those tetraploid populations with climate warming and human induced habitat
outside the regions of main agricultural use of the changes. Even those in genebanks will only survive
species in Europe, Asia and North Africa and the if adequate funding is provided.
84 A.V. Stewart and N.W. Ellison

5.4.4 Quaternary Gene Pool Table 5.1 Diploid accessions of Dactylis glomerata in gene-
banks of Europe (Eurisco), North America (USDA and Canada)
and New Zealand (Margot Forde Germplasm Center) with
This can be defined as those species outside Dactylis, priority for collection
which may be a source of genes for breeding. However, Diploid population Number of Priority
accessions for
only a few species have ever been hybridized with
collection
Dactylis, such as Lolium multiflorum (Oertel et al.
altaica 0 Urgent
1996), Festuca arundinacea (Matzk 1981), and Phleum aschersoniana 26 Low
pratense (Nakazumi et al. 1997), and successful embryos “castellata” 3 High
were formed in cereals (wheat, barley, rye) pollinated himalayensis 4 High
with D. glomerata (Zenkteler and Nitzsche 1984). As hyrcana 0 Urgent
fertility is extremely low these hybrids have never been nestorii ¼ ibizensis 2 High
fully explored in breeding programs and their develop- izcoi 40 Low
judaica 10 Moderate
ment does not appear to be a priority.
juncinella 4 High
lusitanica 11 Moderate
mairei 10 Moderate
5.4.5 Conservation Recommendations metlesicsii 0 Urgent
parthiana 3 High
reichenbachii 0 High
We can conclude that genetic resources of the primary santai 9 Moderate
and secondary tetraploid gene pools are moderately sinensis 0 Urgent
well represented in genebanks and breeders’ collec- smithii (Canary Islands) 10 Moderate
smithii (Cape Verde) 0 Urgent
tion. The tertiary diploid gene pool is in urgent need of
woronowii 21 Low
collection as many of the geographic forms are not
Note: these collections are likely to include some duplication
present in international genebanks or are represented
by no more than two accessions (Table 5.1). In partic-
ular, altaica, hyrcana, metlesicsii, reichenbachii and repeats (SSRs) identified from these libraries will be
smithii from Cape Verde are absent from collections, aligned to rice chromosomes to determine predicted
and sinensis is difficult to obtain for anyone outside locations of the SSR markers. PCR primers designed
China. It is also important to collect and store the from 30 UTR regions from other species contain high
autotetraploid forms of the diploid populations such degrees of polymorphism, and the same is expected
as smithii, woronowii, izcoi, reichenbachii and mairei, for this library. Additionally, the sequence data from
and also of any others that occur. the library will be used to identify single nucleotide
Although in situ conservation should be encour- polymorphisms (SNPs). The USDA-ARS Forage and
aged, it is important to maintain ex situ collections of Range Research Laboratory have an extensive orch-
each form. These can be stored as seed collections in ardgrass improvement program, and their objective is
the international agricultural genebanks. to identify phenotypic associations, and apply the
markers to a marker-assisted selection program for
increased salt tolerance and winter hardiness.
In order that molecular resources can be applied in
5.4.6 Genomics Resources an effective and non-dominating balanced way, it is
important to ensure that pragmatic field breeding pro-
Bushman et al. (2007) and Robins et al. (2008) are grams continue in all major regions.
generating Dactylis expressed sequence tag (EST)
libraries using four tissues: cold acclimated crowns,
etiolated seedlings, salt/drought stressed shoots, and
salt/drought stressed roots. They are also using genetic 5.4.7 Karyotype
association mapping populations, and trait evaluation
field plots to help overcome winter hardiness and to The karyotypes of many diploid Dactylis forms has
provide tolerance to dry summers. Simple sequence been reviewed by Wetschnig (1991). There has been no
5 Dactylis 85

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169: 1615–6110 Sahuquillo E, Lumaret R (1999) Chloroplast DNA variation in
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Tyler B, Borrill M (1983) The use of wild species in forage grass in Dactylis. NZ J Bot 10:573–584
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Chapter 6
Dichanthium

Vishnu Bhat, C. Mahalakshmi, Shashi, Sunil Saran, and Soom Nath Raina

6.1 Introduction have been reported in the D. annulatum complex

(Mehra 1961). Diploid species such as D. armatum,
D. maccannii, and D. panchaganiense are endemic to
The genus Dichanthium Willemet belongs to the tribe
India (Harlan 1963), whereas other diploid species
Andropogoneae of the family Poaceae, having wide
such as D. humilis, D. setosum, D. sericeum, and
distribution across the tropics and the subtropics of the
D. superciliatum are of Australian origin but widely
Old World. It is one of the major components of Sehima-
distributed (de Wet and Harlan 1962; Fig. 6.1). The
Dichanthium and Dichanthium–Cenchrus–Lasiurus
tetraploid agamospecies such as D. annulatum,
grass covers of India (Dabadghao and Shankarnarayan
D. aristatum, and D. caricosum have diploids, which
1973). Various species of Dichanthium such as D. annu-
are sexually reproducing and are well adapted poly-
latum, D. sericeum, D. caricosum and D. aristatum are
haploids (de Wet 1968). D. caricosum produce hybrids
of economic importance because of their high palatabil-
with both D. annulatum and D. aristatum naturally
ity and nutritional value. D. aristatum is useful for
because of which these species are grouped as one
stabilization of waterway and banks and for suppressing
large agamospecies.
invasive weeds such as Phyla canescens. Most of the
Development of efficient genetic improvement
Dichanthium species are used as pastures for grazing
methods requires comprehensive analysis of various
and are suitable for silage and hay. Recent studies have
information on the taxonomy, cytology, reproductive
shown that D. annulatum has the highest relative prefer-
biology, and molecular genetics of cultivated
ence value than any other grasses of northern grasslands
Dichanthium species and their wild relatives. The
of Pakistan (Sultan et al. 2008). Dichanthium forms an
information provided in this chapter can be utilized
agamic complex with Bothriochloa Kuntze and Capilli-
for systematic exploitation of the gene pool for com-
pedium Stapf. (de Wet and Harlan 1968, 1970a). It has
mercial breeding.
sexually reproducing diploid representatives and apo-
mictic polyploids with pseudogamous type of apomixis
(Brown and Emery 1957; Harlan et al. 1958; Harlan and
de Wet 1963a; de Wet and Harlan 1970a; D’Cruz and 6.2 Taxonomy and Distribution
Reddy 1971).
Around 20 species have been reported in this genus
Dichanthium first described by Willemet (1976)
from tropical to the subtropical region. The basic
belongs to the family Poaceae, Subfamily Panicoi-
chromosome number is 10 and different ploidy levels
deae, Group Andropogonodae, Tribe Andropogoneae.
The genus Dichanthium Willemet is closely asso-
ciated with the genera Bothriochloa Kuntze and
Capillipedium Stapf. because of which they were all
V. Bhat (*)
included under Andropogon Linn. by Hackel (1889),
Department of Botany, University of Delhi, Delhi 110 007, India forming Dichanthium–Bothriochloa–Capillipedium
e-mail: [email protected] agamic complex (de Wet and Harlan 1968, 1970a;

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 89
DOI 10.1007/978-3-642-14255-0_6, # Springer-Verlag Berlin Heidelberg 2011
90 V. Bhat et al.

Fig. 6.1 Diagram showing distribution of four important spe- end shows the suitable areas (dark red), i.e., the locations where
cies, namely D. annulatum (a), D. aristatum (b), D. caricosum a species is highly adapted and marginal areas (orange), where
(c), and D. sericeum (d), major forage crop species of the species occurrence is uncertain but can survive in that
Dichanthium across the world [From: Cook et al. (2005)]. Leg- location

Clayton 1977). It is sometimes very difficult to treat are recognized taxonomically as B. glabra and B.
agamic complex taxonomically as every adaptive grahamii, respectively (de Wet and Stalker 1974).
genotype is maintained as a distinct morphological Majority of the species of Dichanthium are perennial,
unit (Baker 1959; Löve 1960; Valentine 1960). The herbaceous, branched or simple, erect, sometimes
hybrids and their derivatives are usually included decumbent or geniculate; rhizomatous or stoloniferous,
with a species that bridges the gap between the mem- tufted grasses, forming upright tussocks, sometimes
bers of agamic complex. Such a species is termed as with extensive creeping stolons; leaves usually glau-
“compilospecies” (Harlan and de Wet 1963b). In case cous, sometimes aromatic; hairy or bearded culm
of Dichanthium–Bothriochloa–Capillipedium, de Wet nodes; auricles absent; ligule membranous and usually
and Harlan (1970b) suggested Bothriochloa interme- ciliate or fringed; plants bisexual; inflorescence terminal
dia as a compilospecies. For example, hybrids of B. or also axillary, solitary or compound racemes digitately
intermedia and either Capillipedium parviflorum or or subdigitately arranged at the summit of the culm;
D. annulatum behave as apomictic allotetraploids and homogamous pairs and obtuse, sessile spikelets, lower
6 Dichanthium 91

or basal spikelets either male or sterile; sessile spike- Queensland, Northern Territory, and New South
lets: bisexual and dorsally compressed or imbricate, Wales and was introduced to Pakistan as a fodder
callus obtuse; fragile rachis, 2 glumes more or less grass (eFloras). It is a perennial, erect, and genic-
equal or subequal, lower glume boat-shaped and char- ulate grass with tall and slender clumps, bluish
taceous to cartilaginous, upper lemma entire and green leaves, nodes hairy or bearded, racemes
awned, glabrous awn bent about the middle; pedicel- silky greenish and purplish, very similar to D.
late spikelets: male or sterile and unawned, palea sericeum (R. Br.) A. Camus. It is distinguished
absent or present, 2 lodicules free and fleshy, 1–3 from D. annulatum by its sessile or subsessile
stamens, ovary glabrous, 2 stigmas; and fruits com- racemes and a subapical ciliate fringe on the
pressed, ornamental, and attractive (Jacobsen 1981; lower glume of the sessile spikelet.
Shouliang and Phillips 2006; Quattrocchi 2008; 2. D. annulatum (Forssk.) Stapf, commonly known as
http://www.efloras.org/florataxon.aspx?flora_id¼2& Marvel grass, is native to Southeast Asia, India,
taxon_id¼109887). The diversity in the type of spike- China, and tropical Africa and also was introduced
lets and the ornamentations of the lower glume of to Australia and America. It has been introduced in
various Dichanthium species has been shown in Texas, South and Southwest America, where it
Fig. 6.2. The taxonomic features of various species persists as casual weed (Gould 1967). It is very
based on Quattrocchi (2008) are described as follows: well adapted to most textures of soil, i.e. from
coralline sands to heavy black clay soil preferably
1. D. affine (R. Br.) A. Camus, commonly known with neutral to alkaline pH. They show moderate to
as bluegrass, is native to Australia with its poor frost tolerance but moderate intolerance to
distribution in southern and western Australia, shade (Cook et al. 2005). Plant has twisting and

Fig. 6.2 Different types of inflorescences of few Dichanthium (m–n) D. panchaganiense and (o) D. sericeum (Courtesy: Dr.
species. (a–d) D. annulatum; (e) D. aristatum; (f) D. armatum; Anil Kumar and Prof. S.R.Yadav)
(g–i) D. caricosum; (j–k) D. foveolatum; (l) D. oliganthum;
92 V. Bhat et al.

ascending to semi-decumbent to prostrate, creep- spikelets elliptic to obovate, pitless, lower glume
ing habits; paniculate inflorescence of compound pilose and fringed.
and several racemes subdigitate and short and gla- 4. D. armatum (Hook.f.) Blatter and McCann has its
brous peduncle, racemes densely villous, spikelets roots of origin in India and is endemic to Western
subimbricate or closely overlapping, sessile spike- Ghats, Maharashtra (Bor 1960). It is an annual
let bisexual, pedicelled spikelets male or neuter, species with characteristic lower glume of pedi-
lower floret barren, upper floret hermaphrodite, celled spikelet armed with marginal bulbous-
pedicellate spikelet without a pit, sessile spikelets based bristles and not pitted, lower glume of sessile
narrowly oblong and awned, lower glume obtuse or spikelet pitted (Bor 1960; de Wet and Harlan 1961).
truncate, keel not winged median nerve present, 5. D. caricosum (L.) A. Camus Hochst. ex Miq.,
sheath terete, larger ligule, upper lemma awned, 3 commonly known as Nadi bluegrass, was first
stamens, ovary glabrous (Bor 1960; Celarier et al. collected at Suva, Fiji Islands in 1936 (Bisset
1962; Shouliang and Phillips 2006; http://www. and Sillar 1984). It shows wide distribution in
efloras.org/florataxon.aspx?flora_id¼2&taxon_id¼ India, Burma, and Ceylon, extending to
200025242). Figure 6.3 represents a diagram- southern China, Indonesia, and Malaya (Bor
matic sketch of D. annulatum showing different 1960; Celarier et al. 1962), where it grows on
taxonomic characters. slightly acidic to alkaline soil with heavy black
(a) D. annulatum (Forssk.) Stapf var. annulatum clay soil texture, intolerant to frost and salinity
is found mostly in tropical Africa, Southwest and moderately tolerant to shade (Cook et al.
Asia, India, and Indonesia. Racemes subdigi- 2005). It forms small clumps, prostrate habit,
tate, sessile spikelet with lower glume pubes- peduncle of raceme glabrous, sessile spikelets
cent to villous and not pitted. hermaphrodite, lower floret empty, upper floret
(b) D. annulatum (Forssk.) Stapf var. papillosum bisexual, pedicelled spikelets male, lower glume
(Hochst. Ex A. Rich.) de Wet and J.R. Harlan of the sessile spikelet obovate or oblong-
is mostly distributed in southern Africa, truncated, without a median nerve, sheath com-
Ethiopia, Tanzania, and Kenya. Rhizomatous, pressed, ligule short ciliate membrane, and mar-
ring of hairs around the nodes, leaf blade gins with long and short hairs interspersed, 3
flattened, leaf sheath glabrous, spikelets stamens, ovary glabrous (Bor 1960; Celarier
paired, sessile spikelet with lower glume not et al. 1962; Clayton and Renvoize 1982;
pitted villous and fringed, pedicelled spikelet Shouliang and Phillips 2006; http://www.
awnless (Mehra 1964a). efloras.org/florataxon.aspx?flora_id¼2&taxon_id¼
3. D. aristatum (Poir.) C.E. Hubb., commonly known 242317870). Figure 6.4 shows the diagrammatic
as Angleton grass, is indigenous from tropical India representation of various taxonomic features of
to Indonesia. It has been introduced to many tropi- this species.
cal regions and has been naturalized in Australia 6. D. compressum (Hook.f.) Jain and Deshpande
(Bisset and Sillar 1984), America (Gould 1967), is a species endemic to India, especially to
South Africa (Bor 1960; Celarier et al. 1962), and Western Ghats in Maharashtra, Lonavala, and
Taiwan and Yunnan region of China (Shouliang Khandala, Poona District (Jain and Deshpande
and Phillips 2006; http://www.efloras.org/flora- 1978). It is a rare and threatened species, with
taxon.aspx?flora_id¼2&taxon_id¼200025243). It leaf blades linear-lanceolate and finely acumi-
is mainly adapted to dark or red soils with neutral nate, aromatic, and lower glume of sessile spi-
to alkaline pH, moderately tolerant to drought, kelets hairy.
shade, and grazing, poor salt tolerance, and moder- 7. D. fecundum S.T. Blake, commonly called as Gulf
ate shade tolerance (Cook et al. 2005). It is weakly bluegrass, has Australian origin with its distribu-
tufted, characterized by hairy peduncle, culm tion in northeastern (Queensland) and western
nodes bearded by very short spreading hairs or Australia (Jacobsen 1981). It has erect or genicu-
glabrous when old, compound racemes sparsely late habit, forming erect tussocks, glaucous leaves,
villous, spikelets imbricate and paired, sessile green or purplish racemes solitary or in group.
6 Dichanthium 93

Fig. 6.3 Dichanthium annulatum Stapf. (a) Part of the plant (i–l) Pedicellate spikelet: (i) Pedicellate spikelet; (j) Lower
with inflorescence; (b) Sessile and Pedicellate spikelet; (c–h) glume; (k) Upper glume; (l) Lemma; (m) Collar (Courtesy:
Sessile spikelet: (c) Lower glume; (d) Upper glume; (e) Lower Dr. Anil Kumar and Prof. S.R. Yadav)
lemma; (f) Upper lemma; (g) Stamens; (h) Pistil and lodicules;
94 V. Bhat et al.

Fig. 6.4 Dichanthium caricosum (L.) A. Camus (a) Habit; (b) spikelet: (j) Lower glume; (k) Upper glume; (l) Lower Lemma;
Sessile and Pedicellate spikelet; (c–i) Sessile spikelet: (c) Lower (m) Upper Lemma; (n) Palea; (o) Stamens; (p) Collar (Cour-
glume; (d) Upper glume; (e) Lower lemma; (f) Upper lemma; tesy: Dr. Anil Kumar and Prof. S.R. Yadav)
(g) Palea; (h) Stamens; (i) Pistil and lodicules; (j–o) Pedicellate
6 Dichanthium 95

8. D. foveolatum (Delile) Roberty, commonly and with marginal bulbous-based bristles (Bor
known as Gandhel is a native species of Africa 1960; de Wet and Harlan 1961).
and tropical and temperate Asia including India, 15. D. queenslandicum B. Simon, commonly known
Ceylon, and Pakistan. It is a wiry, very slender, and as king bluegrass, is a native of Queensland,
much-branched plant, with spathiform leaf sheaths, Australia. It is a vulnerable species (Kew).
basal sheaths silky, ligule membrane-like, leaves 16. D. sericeum (R. Br.) A. Camus, called as Queens-
narrow almost filiform, terminal and axillary land bluegrass, has a wide distribution in tropical
racemes solitary and long-exserted, awn on one Asia, Papua New Guinea, western Australia,
side of spike, lower glume of sessile spikelet pitted, Queensland, Northern Territory, Victoria, and
pedicelled spikelet pitted or not, and upper lemma New South Wales and has been introduced to
sometimes bidentate (Clayton and Renvoize 1982; US (Gould 1967). It is usually prevalent in neutral
GRIN, eFloras). to alkaline soil conditions and very well adapted
9. D. humilius (Benth.) J.M. Black is widely to heavy black clay soil moderate to poor toler-
distributed in Central Australia. The plant is char- ance to shade and grazing but intolerant to shade
acterized by sessile or subsessile raceme, hairs on (Cook et al. 2005). It is identifiable with its tus-
lower glume of sessile spikelets not strictly socky nature, bluish green and purplish, culm
arranged in a subapical transverse fringe, racemes nodes bearded or with long white hairs, racemes
2–7 per inflorescence (de Wet and Harlan 1962; 2–7 with white and bluish silky hairs, pedicels
GRIN). usually bilaterally ciliate, spikelets paired and
10. D. maccannii Blatter is a vulnerable and crowded, lowest pairs of spikelets sterile and
threatened species, endemic to India, especially reduced to glumes, callus shortly bearded, pedi-
to Panchgani Plateau in Western Ghats, Mahar- cellate spikelets sterile, hairs on lower glume of
ashtra. It has racemes solitary or two and charac- sessile spikelet arranged in distinct subapical
teristic not pitted lower glume of sessile spikelet transverse fringe (de Wet and Harlan 1962, Kew).
and marginal bulbous-based bristles on lower (a) D. sericeum (R. Br.) A. Camus subsp. humi-
glume of pedicellate spikelet (Bor 1960; de Wet lius New South Wales.
and Harlan 1961; Naik 1982). (b) D. sericeum (R. Br.) A. Camus subsp. poly-
11. D. mucronulatum Jansen has its distribution in the stachyum Australia, Asia.
tropical Asia mainly Indo-China and Malaysia. It (c) D. sericeum (R. Br.) A. Camus subsp. seri-
is a rare species, ligule eciliate membrane, hairy ceum New South Wales.
awn, upper lemma sometimes bidentate (Kew). 17. D. setosum S.T. Blake is an Australian species,
12. D. oliganthum (Hochst. ex Steud.) Cope is mainly mainly distributed in Queensland, New South
found in India. It is an annual species (Kew). Wales, and western Australia. It differs from
13. D. pallidum (Hook.f.) Stapf ex C.E. is reported to other species by the presence of few, never
be endemic to India, especially Nilgiris, Tamil strictly digitate racemes, awned pedicellate
Nadu, by Bor (1960), and its occurrence was spikelet, hairs on the lower glumes arranged in a
reported in other parts of India namely, Mirzapur, distinct subapical fringe, lower glume of sessile
24 Parganas in West Bengal and Nellore by Bhat- spikelet 5–6 nerved (de Wet and Harlan 1961,
tacharyya and Uniyal (1973). It is characterized 1962; Jacobsen 1981).
by its small creeping nature, solitary racemes of 18. D. tenue (R. Br.) A. Camus, commonly called as
tightly overlapped spikelets, peduncle glabrous small bluestem, is a species from Australia now
and lower glume of sessile spikelet with shallow confined to Malaya, Polynesia, Queensland, and
longitudinal furrow (GRIN; Bor 1960; Celarier New South Wales. It has prostrate habit, culm
et al. 1962). nodes essentially glabrous, glabrous peduncle of
14. D. panchaganiense Blatter and McCann is a spe- raceme, racemes 1–3, lower glume of sessile spike-
cies endemic to India particularly to Panchgani let with short hairy margins (Celarier et al. 1962).
Plateau, Maharashtra. It is a rare and threatened, 19. D. tenuiculum (Steudel) S.T. Blake is an Austra-
annual species, with racemes solitary or digitate, lian species, commonly known as tassel bluegrass.
lower glume of pedicelled spikelet always pitted It is widely spread around western Australia,
96 V. Bhat et al.

Queensland, Northern Territory, Philippines, D. acutiusculum, D. humilius, D. panchganiense,

Timor and New Guinea. It is a robust grass, leaves D. maccannii, and D. tenuiculum, and diploid races
bluish green to blue and very scabrous, culms of D. annulatum, D. aristatum, and D. caricosum
robust and branched, racemes in a terminal and have chromosome number 2n ¼ 20. The tetraploid
digitate or tasseled inflorescence (Jacobsen 1981). races of facultative apomicts such as Dichanthium
20. D. woodrowii (Hook.f.) S.K. Jain and Deshpande annulatum, D. aristatum, D. caricosum, and obligate
is a rare grass endemic to India, confined to apomict D. fecundum have 2n ¼ 40, whereas the hexa-
Mawal and Paud of Pune District, Maharashtra. ploid obligate apomict D. papillosum and hexaploid
It is a vulnerable and threatened species with race of D. aristatum have 2n ¼ 60. D. armatum is
aromatic, tufted, woody rootstock, sessile spike- cytologically unknown (Oke 1950; Gould 1956; Celar-
lets oblong-lanceolate (Jain and Deshpande 1978; ier and Harlan 1957; Bor 1960; Bogdan 1977).
Mehrotra 1979). Morphological data have suggested hybridization
and chromosome doubling as the major phenomena in
the evolution of Dichanthium Willemet. The meiotic
6.3 Cytology and Hybridization behavior is regular in case of diploids, whereas poly-
ploids show irregularities, which suggest meiotic irre-
gularities in addition to morphological variations in
Cytogenetic diversity and chromosomal behavior play
these species. de Wet and Borgaonkar (1963) found
an important component of any breeding program. The
that naturally occurring hybrids were euploids and
occurrence of natural interspecific and intergeneric
facultative apomicts, but artificial hybrids were either
hybrids has been reported in Gramineae, and a thor-
euploid or aneuploid, and very few were strictly sex-
ough study of these hybrids enrich our understanding
ual. Depending on these observations, it was proposed
on the fields of genetics, cytogenetics, biosystematics,
that apomixis is a dominant trait over sexuality and
and phylogenetics. Many workers including Myers
also that euploids have selective advantage over aneu-
(1947) and Carnahan and Hill (1961) have reported
ploids in nature (de Wet and Borgaonkar 1963; Harlan
many interspecific and intergeneric hybrids, especially
et al. 1964).
in forage grasses. Stebbins (1947) and Löve (1964)
By cytoecological studies, it was hypothesized that
have proposed that natural polyploids, which arise
the tetraploids have arisen from doubling of the related
usually, fall between autopolyploids and allopoly-
diploid species, which, over a period of time, evolved
ploids. Polyploidy played an important role in the
and differentiated cytologically, whereas hexaploids
evolution of species, especially the grasses. Allopoly-
originated through hybridization of tetraploids and
ploids played significant role in speciation than autop-
diploids. These theories have been postulated by
olyploids.
Celarier and his coworkers in 1958, especially in
case of D. annulatum. In D. annulatum, four different
morphological types can be distinguished, viz. Tropi-
6.3.1 Cytological Characters cal, Mediterranean, Senegal, and South African types
(Harlan et al. 1958, 1961; Mehra 1964b). Tropical
Dichanthium, like many other members of agamic ones include prostrate or decumbent, diploid as well
complex, has autotetraploidy along with haploidy as a as tetraploid types. The Mediterranean types are gen-
mechanism to bridge gap and ensure contact with sex- erally erect with longer inflorescences, tetraploid, and
uality (de Wet 1965, 1968). Mehra (1961), de Wet and more adapted to desert conditions. The Senegal type,
Harlan (1968) and many others have reported chromo- represented by single collection from northwest
some numbers in the sexually reproducing and aga- Africa, is considered to be extreme form of Mediterra-
mospecies of Dichanthium. The basic number for the nean type. The erect-growing South African types are
tribe Andropogoneae is 5 or 10 and Dichanthium spe- all hexaploids and obligate apomicts (Mehra and
cies show multiples of 10 in sexual and apomictic types Celarier 1958; Mehra 1962, 1964b; de Wet 1968).
(Celarier 1956, 1957; Celarier and Harlan 1957). The Morphological and cytological studies of these races
sexually reproducing diploid species of Dichanthium, and their hybrids show that they are not genetically
namely D. sericeum, D. affine, D. setosum, D. tenue, isolated. It was, therefore, suggested to group three
6 Dichanthium 97

races, namely the Tropical, Mediterranean, and Sene- 6.3.3 Hybridization

gal types under D. annulatum var. annulatum.
Crossing experiments in Dichanthium were started ear-
lier by many workers in order to study the phenomenon
6.3.2 Ploidy Cycles in Dichanthium of apomixis in this genera. Several combinations of
crossings between different species of Dichanthium,
Various embryological, cytogenetical, and hybridiza- Capillipedium, and Bothriochloa and analyses of dif-
tion studies indicated that diploid species of all the ferent characters of the progenies from such crosses
three genera of the agamic complex reproduce sexu- revealed that they were morphologically same as mater-
ally, while the polyploids (2x, 4x, 6x) are facultative nal types, which were due to apomictic mode of repro-
apomicts, although there are a few exceptions (de Wet duction and not due to maternal inheritance. Celarier
and Borgaonkar 1963; de Wet 1965, 1968). Diploid and Harlan (1957) were able to establish this when they
Dichanthium spp. are fully sexual and fertile, but crossed among diploid and polyploid species of
occasionally produce autotetraploids as a result of D. annulatum (2n ¼ 20, 40, 60) and D. caricosum
fertilization of the unreduced male and female (2n ¼ 40); D. annulatum (2n ¼ 40) with diploid and
gametes. The present day diploid races of tetraploid polyploid Bothriochloa sp. [B. intermedia (2n ¼ 40,
agamospecies of D. annulatum, D. aristatum, and 60), B. ischaemum (2n ¼ 50, 60), B.venusta
D. caricosum have been found to represent well (2n ¼ 40)] and Capillipedium sp. [C. parviflorum
adapted polyhaploids as they resemble few sexually (2n ¼ 40), C. spicigerum (2n ¼ 40)]. Cytologically
reproducing artificial polyhaploids (de Wet and Singh reduced or unreduced female gametes can function
1964; de Wet 1965; de Wet and Harlan 1970a; D’Cruz sexually or develop parthenogenetically in this genera
and Reddy 1971). Polyhaploidy is rare in nature as (Harlan and de Wet 1963a).
they are usually completely sterile and poorly adapted. Introgressive hybridization between different spe-
But a few polyhaploids produced from natural faculta- cies of Dichanthium, for example, D. annulatum with
tive apomicts or from natural or artificial hybrids other species such as D. aristatum, D. caricosum,
between D. caricosum and D. annulatum, or D. aris- D. papillosum, and D. fecundum (Celarier et al.
tatum resemble natural diploids at morphological 1961; Chheda and Harlan 1963; Borgaonkar and de
and chromosomal level (de Wet 1965). Haploids Wet 1964; Mehra 1966) or within different ecotypes as
obtained from Mediterranean ecotype (Mehra 1964b) available in case of D. annulatum (Mehra and Celarier
of D. annulatum do not survive and are completely 1958; Mehra 1962), or even across genera, for exam-
sterile, whereas those from Tropical ecotypes were ple, Bothriochloa and Capillipedium (Harlan et al.
found to be vigorous, reproduce sexually but sterile, 1958; Harlan 1963; de Wet and Harlan 1968, 1970a)
indicating that apomixis is not functional in haploids. provide a wide platform of variability in the genus
Presence of such fertile, sexually reproducing haploids Dichanthium Willemet.
have phylogenetic significance as it helps in maintain- Studies in this direction were carried out by Harlan
ing contact with sexuality in predominantly polyploid et al. (1964), wherein crosses between Bothriochloa
agamospecies like Dichanthium (de Wet 1968; de Wet grahmii and D. annulatum yielded hybrids, which
and Harlan 1970a). Triploids derived from hybridiza- were apomictic when both parents were apomicts
tion within species of sexually reproducing diploid and sexual when both were sexual. Similar results
races of tetraploid D. annulatum, D. aristatum, and were also obtained by Harlan et al. (1961) and Singh
D. caricosum and between sexual tetraploid D. annu- (1968) when they crossed tetraploid D. annulatum
latum and other diploids or from fertilization of dip- with tetraploid B. grahmii; and diploid and tetraploid
loid female gametes with haploid male gametes have D. caricosum, respectively. In crosses between apo-
been reported. These triploids were all sterile but quite mictic  sexual and vice versa, hybrids were mostly
vigorous and cytologically irregular (de Wet and apomicts, but frequency of sexuals was high compared
Singh 1964). Hexaploid D. papillosum, which has to apomict  apomict crosses. Harlan and de Wet
hybrid origin from two tetraploid D. annulatum (1975) were even able to obtain sexual types by cross-
(Mehra 1962), is an obligate apomict. ing facultative apomicts.
98 V. Bhat et al.

Hybrids with different ploidy levels were obtained et al. (1962) along with previous works by de Wet
in crosses between facultative apomictic tetraploid (1954) and Celarier et al. (1961) proposed that spe-
D. annulatum and diploid sexuals of D. aristatum cies such as D. aristatum, D. caricosum, D. pallidum,
and D. caricosum, and their progenies were categor- and D. tenue form an interrelated agamic complex
ized as sexual and apomictic types (D’Cruz and wherein cytologically reduced and unreduced female
Reddy 1971). In another study, D. caricosum was gametes function sexually or parthenogenetically to
found to resemble polyhaploids from F1 hybrids of form a series of hybrids with chromosome numbers
D. caricosum and D. annulatum, even though diploid ranging from 2n ¼ 20 to 2n ¼ 60.
D. annulatum and D. caricosum are reproductively The natural tetraploids of D. annulatum are regarded
isolated. D. aristatum was also indistinguishable as segmental allopolyploids (Stebbins 1947) and prob-
from polyhaploid derived from F1 hybrids of ably originated from hybridization between closely
D. aristatum and D. caricosum. Thus, D. caricosum related species or partially isolated races of same spe-
acts as a genetic bridge between D. annulatum and cies. de Wet et al. (1961) studied the chromosomal
D. caricosum at tetraploid level. association in hybrids of Dichanthium. Hybridizations
In a tetraploid cross, when an unreduced female egg between two plants of D. annulatum with chromosome
cell is fertilized by a reduced male gamete, the resul- complement AAA1A1 and A2A2A3A3 result into
tant progeny is a hexaploid. Best example of this being hybrids (AA1A2A3), which associate cytologically
D. papillosum, which is the result of fertilization of into 20 bivalents, sometimes as trivalents or tetrava-
unreduced egg cell of Tropical type with reduced male lents. Hexaploids have AAA1A1A2A3, thus bringing
gamete of Mediterranean type of D. annulatum together whole genome of one parent with half of the
(Borgaonkar and Singh 1962; Mehra 1964b). It was, other parent. Hexaploid hybrids from D. aristatum
therefore, suggested to classify D. papillosum as (DDD1D1) and D. caricosum (CCC1C1) have chromo-
D. annulatum var. papillosum (Pilger 1954). Simi- some compliment as CCC1C1DD1 and DDD1D1CC1
larly, tetraploid D. fecundum show morphological and behave cytologically as D. annulatum. Hybrids
and cytological characters same as D. annulatum and (AA1FF1) between D. annulatum and D. fecundum
is thus classified as D. annulatum var. fecundum (FFF1F1) behave like one of the parents. Therefore,
(Hackel 1889). These three species together form an de Wet et al. (1961) were able to prove that the hybrids
interrelated agamospecies, i.e., the D. annulatum com- show autosyndetic pairing of the chromosomes in
plex (Singh and Mehra 1965). Dichanthium Willemet and also that chromosome pair-
Celarier et al. (1961, 1962) also studied morpho- ing can take place within basic genome (n ¼ 10), indi-
logical and cytological characteristics of the hybrids cating the original basic number as 5, as earlier
between D. tenue (2n ¼ 40) and D. aristatum suggested by Celarier et al. (1960).
(2n ¼ 40), which differ in their morphology by the
presence of glabrous peduncles and prostrate habit in
the former. Three of the hybrids with 2n ¼ 40 chro-
mosomes were decumbent with wooly peduncles,
6.4 Embryology
whereas two of them had 2n ¼ 60, which resembled
D. tenue when it was used as female parent. The The embryological studies and apomixis in Dichanthium
hexaploid was found to be the result of cross between have been conducted in detail by many workers (Brown
cytologically unreduced female gamete from D. tenue and Emery 1957; Celarier and Harlan 1957; Reddy
and reduced male gamete from D. aristatum. The and D’Cruz 1969a) using microtomy and ovule squash
reciprocal crosses also yielded a hexaploid with the method. Microsporogenesis was found to be slightly
unreduced gamete coming from D. aristatum and irregular with the formation of a few univalents and
resembled D. aristatum morphologically. Hybrids multivalents at metaphase I and bridges and laggards
from such crosses also yielded morphologically inter- at anaphase I. Second meiosis was comparatively
mediate types with semi-decumbent habit and slightly more regular and forms 3-nucleate pollen grain with
pubescent peduncles. These morphological variations thick wall and single germ pore.
in the hybrids indicate hybridization with limited The female gametophyte development initiates with
genetic segregation in facultative apomicts. Celarier the differentiation of megaspore mother cell from the
6 Dichanthium 99

nucellus, which later degenerates (Bhanwra 1988). One function of the opening was to provide the least resis-
or more cells of the nucellus show enlargement and tance to the growing pollen tube and therefore enhance
vacuolation and form the aposporous embryo sacs. the chances of fertilization. The presence of such an
D. annulatum, being a facultative apomict, has sexual opening in the facultative apomicts favors the chances
and apomictic embryo sac occurring together in the of sexuality. Harlan et al. (1964) extensively studied
same ovule or different ovules of same inflorescence. the embryology of six accessions representing differ-
The frequency of apomictic embryo sacs are generally ent ploidy levels and four species belonging to two
higher than the sexual ones (Brooks 1958). The embryo genera, viz. Bothriochloa and Dichanthium. They also
sacs can be of sexual, reduced type with 8-nuclei or studied three diploid species of Bothriochloa and 15
aposporous, unreduced with 4-nuclei. collections representing four diploid species of
Apart from genotype, environment also plays a role Dichanthium. They found sexuality at the diploid
in regulating the type of embryo sac development. level and apomixis at higher levels and concluded
According to Heslop-Harrison (1961), photo- and sexuality and asexuality to be dependent on synchro-
thermo-periods have considerable role in embryo sac nization of different embryological events.
development. In D. aristatum, photoperiodic condi- With respect to fertilization and post-fertilization
tions during inflorescence development can be corre- events, in case of 4-nucleate embryo sac, only one of
lated to apomixis and pollen fertility. Knox (1967) the male gametes fuses with the polar nuclei and the
correlated the presence of apomictic embryo sacs and other one either enters egg or remain outside. In most
day-length in six localities in Australia, which was of the ovules, single aposporous embryo sac is func-
earlier shown by Knox and Heslop-Harrison (1963) tional and therefore embryo and endosperm develop-
under controlled conditions. The pollen fertility was ment was observed in only one embryo sac. In case of
also found to be affected under short day conditions, apomicts, there was precocious development of
which favor apomictic embryo sac production (Knox embryos in the ovules, and embryo formation was
and Heslop-Harrison 1966). Similarly, in case of observed even before anthesis. Reddy and D’Cruz
D. annulatum, the frequencies of sexual and apomictic (1969a) also described the apomictic mode of devel-
embryo sacs fluctuate with season (Gupta et al. 1969). opment of embryo by various mechanisms for elimi-
Saran and his coworkers made worldwide collections nation of syngamy mechanisms are gamtes either
of different Dichanthium species and studied the vari- getting degenerated inside the egg cell or fail to enter
ation in these species. They found that all the species it or are restricted to the suspensor cell. Other mechan-
were uniform in their characters when collected along isms include fertilization of both polar nuclei by two-
the equatorial region, whereas the species showed male gametes or development of adventitive embryos
differences when moved away from equator. They from nucellus. Polyembryony in Dichanthium has
reasoned the kind of uniformity shown among the also been reported in the apomictic species, D. annu-
species along the equator due to the photoperiod, i.e., latum. The development of twin embryos occurs in
equal day-length. In this way, the breeders can induce two ways. Either the two embryos develop indepen-
the type of embryo sac development by manipulating dently in two different embryo sacs or in the same
the environmental conditions. Experimentally, the embryo sac. The former one is more prevalent than the
shift from sexual to apomictic embryo sac has been latter. In the latter case, one of the embryos enlarges
tried. For example, Asker (1966) was able to increase and pushes the other, which remains as a rudimentary
the sexual embryo sacs in facultative apomict Poten- structure (Reddy and D’Cruz 1969b).
tilla. Apart from the 8-nucleate and 4-nucleate embryo
sac distinction for detecting sexuality and apomixis,
respectively, Saran and de Wet (1969) found a pecu- 6.5 Nature and Inheritance of
liar opening, which was consistently found in 8-nucle-
ate embryo sacs of D. intermedium [equivalent to
Apomixis in Dichanthium
Bothriochloa bladhii (Retz.) S.T. Blake according to
Quattrocchi 2008] and never in the aposporous Apomixis is generally defined as asexual reproduction
embryo sacs. The opening was the result of unequal through seeds (apo ¼ away from, and mixis ¼ act of
thickening in the wall layers of the embryo sac. The mixing; Asker and Jerling 1992). The increase in
100 V. Bhat et al.

number of chromosomes due to polyploidy many a 6.5.1 Bothriochloa–Dichanthium–

times induces meiotic abnormalities, resulting in steril- Capillipedium Agamic Complex
ity. To overcome sterility, species have evolved an
alternate mode of reproduction, namely apomixis. The
phenomenon of apomixis in Poaceae is quite frequent as Grass species have a typical agamic complex, which
reported by Brown and Emery (1957, 1958) and Pull- consists of several basic diploid sexuals and many
aiah and Febulaus (2000). Apomixis in Dichanthium sp. apomictic polyploids (Harlan and de Wet 1963a, b).
along with its two closely related members of the tribe The genus Dichanthium forms an interrelated agamic
Andropogoneae, viz. Bothriochloa O. Kuntze and complex (Celarier and Harlan 1957; de Wet and
Capillipedium Stapf., was first studied by Celarier Harlan 1970a; Saran and de Wet 1970; de Wet and
and Harlan (1957). Using hybridization and cytolo- Stalker 1974) with two other genera of the tribe
gical studies, they were able to establish apomictic Andropogoneae, viz. Bothriochloa and Capillipedium
reproduction in 12 species of these three genera, exclud- (Fig. 6.5). Gene flow within this agamic complex
ing the possibilities of maternal inheritance and self- occurs in different directions but sometimes there
fertilization (Celarier and Harlan 1957; Celarier et al. are a few incompatibility barriers. Srivastava and
1958). The type of apomixis in Dichanthium Willemet is Purnima (1990) studied seven polyploid taxa involv-
of aposporous pseudogamous type (Gustafsson 1947a, ing Bothriochloa–Dichanthium complex and Hetero-
b; Brown and Emery 1957; Celarier and Harlan 1957; pogon contortus and Paspalum paspaloides to
Harlan et al. 1958; de Wet and Harlan 1970a; D’Cruz establish different aspects of apomixis in these spe-
and Reddy 1971). In aposporous type of apomixis, all cies. Species of the genus Bothriochloa, especially B.
the four megaspores developed from megaspore mother intermedia (R.Br.) A. Camus [equivalent to B. blad-
cell degenerate and functional embryo sac is formed hii (Retz.) S.T. Blake according to Quattrocchi
from another nucellar cell (called aposporous initial), 2008], acts as a bridging species for crosses between
which is unreduced; pseudogamy is the development of Dichanthium and Capillipedium spp. (de Wet and
endosperm after fertilization of male gamete with the Harlan 1970b; Berthaud 2001). B. intermedia
polar nuclei or the secondary nucleus. (R.Br.) A. Camus introgresses with B. ewartiana
Apomixis was reported to be dominant over sexu- (Domin) C.E. Hubbard, B. ischaemum (Linn.) Keng,
ality and two pairs of genes were found to be involved Capillipedium parvifolium (R.Br.) Stapf, and
at tetraploid level, which are heterozygous in case of D. annulatum (Forssk.) Stapf. Therefore, it was sug-
apomicts and homozygous in sexuals. The tetraploid gested that B. intermedia acts as a compilospecies
apomicts were assigned the genotype AaAa and sexual and is carrying genes from several other species of
ones aaaa, in the sense that the “A” allele induces Bothriochloa O. Kuntze, as well as species of Capil-
formation of 4-nucleate apomictic embryo sac and lipedium Stapf. and Dichanthium Willemet (Harlan
“a” allele does not (Harlan et al. 1964; de Wet and and de Wet 1963a, b; de Wet and Harlan 1966,
Harlan 1970a). D’Cruz and Reddy (1971) also con- 1970a, b). It was also suggested by de Wet and
cluded sexuality and apomixis to be independent phe- Harlan (1968) to combine all the three taxa under
nomena, associated with ploidy but found its Dichanthium genera.
inheritance as complex compared to earlier report by
Harlan et al. (1964). They, like many other workers
(Harlan et al. 1958 in Dichanthium sp., Chheda et al. 6.6 Breeding and Genomics
1961 and Borgaonkar and de Wet 1961 in Bothrio-
chloa sp.), also emphasized on delicate nature of
genetic balance required for the expression of apo- 6.6.1 Germplasm Collections
mixis in Dichanthium spp. due to sexual reversion of
hybrids in F2 generation. Therefore, agamospecies, The online software project namely Germplasm
which were once considered to be genetic dead ends, Resources Information Network (or GRIN) of the
are now characterized by dynamic genetic system, National Genetic Resources Program of USDA pro-
several mechanisms to facilitate recombination, and vides germplasm information about plants, animals,
capable of high degree of adaptive polymorphism and microbes, and invertebrates important for food and
progressive evolution (de Wet and Harlan 1970a). agricultural production. This project lists different
6 Dichanthium 101

CAPILLIPEDIUM DICHANTHIUM

caricosum papillosum
2x, 4x 6x

parviflorum assimile aristatum annulatum

2x, 4x 2x, 4x 2x, 4x 2x, 4x

glabra grahamii
4x, 6x 4x, 6x

ewartiana intermedia ischaemum

5x, 6x 4x 4x, 6x

concanensis insculpta woodrowii

5x 5x, 6x 5x

kuntzeana pertusa radicans compressa

2x 4x 4x 2x

BOTHRIOCHLOA

Fig. 6.5 Diagrammatic representation showing relationship among the members of Bothriochloa–Dichanthium–Capillipedium
agamic complex [Redrawn from Berthaud (2001)]

species and varieties of genus Dichanthium, and its basic methods for cultivar development focuses on
available accessions are in the National Plant Germ- selection of useful commercial genotypes from a col-
plasm System (NPGS). It shows that there are around lection of apomictic genotypes or breeding of syn-
80 accessions of D. annulatum with majority of the thetic tetraploids with natural tetraploid apomicts.
accessions with Pakistan (29 accessions) and India With the available germplasm collections, the major
(28 accessions). There are seven accessions of job is to identify the best genotype among the
D. aristatum with majority in US and 14 of D. sericeum candidate genotypes. The current and previous propo-
with majority available in Australia. There are only sals for breeding involved the perpetuation of hetero-
two accessions available for D. foveolatum and one zygous genotypes but failed to include means to
each of D. caricosum and D. maccannii. exploit heterosis (Miles 2007). The success of any
breeding program depends on the range of variability
present in the germplasm collection, the proportion of
sexual and apomictic seed production, and variability
6.6.2 Development of Cultivars from crossbred progenies. Like many other grass
genera, Dichanthium lacks a collection of different
Breeding of apomictic species was a serious challenge genotypes, and there is a need to look for mechanisms
due to difficulty in obtaining recombination. With the that could create variability.
increasing advances in understanding of the genetics A wide platform of variability is available in case of
of apomixis and availability of various tools, this task Dichanthium among its different species such as
now seems to be achievable (Bashaw 1975, 1980). The D. aristatum, D. caricosum, D. papillosum, and
102 V. Bhat et al.

D. fecundum and among different ecotypes (as avail- diversity can be evaluated on the basis of morphologi-
able in case of D. annulatum) or even across related cal, phenotypic, and genotypic characters. The last is
genera, for example, Bothriochloa intermedia, B. more convenient and authentic than the first two, which
ischaemum, B. pertusa, and C. parviflorum (Harlan are usually time-consuming and under the influence of
et al. 1958; Harlan 1963; de Wet and Harlan 1968, environmental changes (Tanksley et al. 1989; Powell
1970a). Also, through genetic segregation and recom- et al. 1996).
bination, new types can be developed by the break-
down of genomic relationships among the
chromosomes introgressed from the species of Bothrio- 6.6.3.1 DNA Extraction Methodology
chloa and Dichanthium (Mehra and Singh 1968).
Variability can be introduced by the use of mutagenic The preliminary requirement of genetic screening
agents as well. For example, conversion of pedicellate studies is good quality DNA. The common problems
spikelet to bisexual flower in D. fecundum, a character observed in tropical grasses are high content of poly-
governed by a single gene is an outcome of mutation of saccharides and lignin, which act inhibitory to poly-
the gene governing it (Borgaonkar and de Wet 1960). merase chain reaction (PCR) analysis and other
Dichanthium species are usually the freely available molecular techniques (Pandey et al. 1996). Chandra
rangeland grasses with high nutritive value. Rai et al. and Saxena (2007) were able to optimize rapid proto-
(1981) undertook a study to evaluate the available mate- col for isolating genomic DNA from five tropical grass
rial and to select the best variety for forage production. species, viz., D. annulatum, Heteropogon contortus,
They identified two cultivars, S-32 and S-65, of Sehima nervosum, Chrysopogon fulvus, and Cenchrus
D. annulatum for maximum forage production, whereas glaucus. The method involved modified CTAB proce-
the cultivars S-123 and S-128 were found to be suitable dure based on Murray and Thompson (1980).
for soil conservation purposes. Seeds of native grass like
D. sericeum are in great demand for the rehabilitation of
landscapes affected by mining or urban development.
6.6.3.2 Protein or Isozyme Markers
Many cultivars have also been released in many
countries for these forage species of Dichanthium. Fol-
Although a facultative apomict, D. annulatum shows
lowing are the promising accessions reported by Cook
high level of polymorphism at molecular level.
et al. (2005) that have been released for cultivation:
Gupta et al. (2003) studied the phylogenetic relation-
1. D. annulatum accessions IGFRI-S-495-1 and ships in four tetraploid species of Dichanthium–
IGFRI-S-495-5 from Indian Grassland and Fodder Bothriochloa complex based on isozyme phenotypes.
Research Institute IGFRI, Jhansi, India, and CPI They found D. annulatum to be evolving continu-
84146B from Queensland, Australia. ously and D. aristatum and D. caricosum to be
2. D. aristatum accession CPI 104839 from Australia. isolated from D. annulatum over a period of time.
3. D. caricosum accessions from Paraguay and north- D. caricosum was found to be more closely related to
ern Argentina D. annulatum than D. aristatum according to protein
4. D. sericeum accessions ES-100 and ES-200 from marker studies.
Australia. The cultivar names and countries of ori-
gin, along with their features, have been tabulated
in Table 6.1.
6.6.3.3 DNA Markers

A high level of polymorphism in the agromorpholo-

gical characters has been reported in D.annulatum,
6.6.3 Genomics independent of its geographical distribution (Chandra
et al., 2004, 2006; Agrawal et al., 1999). Random
Little is known about the genome of Dichanthium, and amplified length polymorphism (RAPD) marker-
it is difficult due to the occurrence of Dichanthium– based studies in different accessions of marvel grass
Bothriochloa complex and their hybrids. The genetic collected from two contrasting regions including the
6 Dichanthium 103

Table 6.1 Detailed list of released cultivars of four major forage species of Dichanthium [reported by Cook et al. (2005)]
S. no. Name of the Name of the cultivar Country of origin Features
species
1. Dichanthium “Kleberg” USA (1944) Excellent drought tolerance, good seed
annulatum producer, moderate salinity tolerance,
relished by cattle.
“Pretoria 90” USA (1954) Selected for seedling vigor, rapid growth
(PI 188926, and yield. Good drought tolerance,
BN-6730, T-20090) leafy; used for pasture, hay and silage.
“T 587”, “PMT-587” USA (1981) Derived from a composite of 80
accessions. Suitable for pasture, hay,
revegetation of disturbed areas and salt
scalds, erosion control, and range
reseeding. More cold tolerant than
“Gordo”, “Medio”, and “Kleberg”.
High forage producer.
“Marvel 8” India 40–100% better dry matter yields than
(CPI 106073) local strain.
2. Dichanthium “Medio” (NSL 20670, USA (1954) Selected from naturalized ecotype in
aristatum NSL 22695) Texas. Best adapted to heavy soils
“Gordo” (PI 190302, USA (1957) Good seed producer.
BN-6851, T-20062,
NSL 22694)
“Alabang X” Philippines Slow to establish and susceptible to weed
(PI 297430) competition when young. Best on
poorer soils Erroneously referred to as
var. heteropogonoides, but may in fact
be an ecotype of D. caricosum.
“T 587”, “PMT-587” USA (1981) Derived from a composite of 50
Dichanthium accessions established in
1962. Suitable for pasture, hay,
revegetation of disturbed areas and salt
scalds, erosion control, and range
reseeding. More cold tolerant than
“Gordo”, “Medio” and “Kleberg”.
Resistant to leaf rust. High forage
producer.
“Floren” (CPI 106374) Australia (1995) Leafy, very palatable ecotype.
3. Dichanthium “Marvel 40” India (1971) Produce 40–100% more dry matter than
caricosum (CPI 106073) the naturally occurring strain.
“Marvel 93” India (1971) Produce 40–100% more dry matter than
(CPI 106075) the naturally occurring strain.
“Alabang X” Philippines Not available
4. Dichanthium “Scatta” Pending release in Not available
sericeum Australia

North Indian plateau and South India revealed South 6.6.3.4 Phylogenetic Studies
Indian accessions to be more polymorphic, whereas
those from central plateau were found to be more Andropogoneae is a member of monophyletic
suitable for drought stress. Thus, Saxena and subfamily Panicoideae and forms a part of large
Chandra (2006) were able to identify three acces- and well-supported panicoid–arundinoid–chloridoid–
sions of D. annulatum suitable for genetic resource centothecoid (PACC) clade of the family Poaceae.
management and for isolating genes controlling Andropogoneae is monophyletic based on data from
drought tolerance. morphology (Kellogg and Watson 1993), chloroplast
104 V. Bhat et al.

100 Andropogon spp. - AN

Core Andropogoneae 78
Hyparrhenia hirta - AT
Schizachyrium scoparium - AN
64
100 Bothriochloa odorata - SO
98
Capiltlpedium parviflorum - SO
Dichanthium aristatum - SO
99 Cymbopogon flexuosus - AN

Cymbopogon jwarancuss - AN
Heteropogon contortus - AT
Cleistachne sorghoides - SO

Microstegium nudum - SA
72
Miscanthus japonicus - SA
Origin of Awns Saccharum officinarum - SA
100 Chrysopogon futvus - SO
Chrysopogon gryllus - SO
Ischaemum spp. - IS
Apluda mutica IS
Sorghum bicolor - SO
Coelorachis selloana - RO
Elionurus muticus - RO
Origin of Disarticulating 95 Tripsacum dactyloides - TR
Rachis
95 Zea mays - TR
Chionachne koenigii - CH
100 Phacelurus digitatus - RO
Coix aqua - CO
52 100 Arundinella hirta
Arundinella nepalensis
Paspalum spp.
Panicum spp.
71
100 Tristachya superba
100
Urochloa mutica
Pennisetum alopecuroides

Danthoniopsis dinteri
10 changes

Fig. 6.6 Single-most parsimonious tree based on combined Anthistiriinae; SO: Sorghinae; SA: Saccharinae; IS: Ischaemi-
sequences of PHYB, ndhF, and GBSSI. Abbreviations follow- nae; RO: Rottboelliinae; TR: Tripsacinae; CH: Chionachninae;
ing taxon names indicate respective subtribes according to CO: Coicinae. (Mathews et al. 2002)
Clayton and Renvoize (1986). AN: Andropogoninae; AT:

gene ndhF (Spangler et al. 1999), and nuclear genes, set of different members of Andropogoneae (Fig. 6.6).
viz., granule-bound starch synthase I (GBSSI) (Mason- Among the awned taxa, they identified a poorly
Gamer et al. 1998) and phytochrome B (Mathews et al. supported clade named as “core Andropogoneae,”
2002). Mathews et al. (2002) combined the three matri- which included Andropogon, Bothriochloa, Capillipe-
ces namely PHYB, GBSSI, and ndhF to generate a data dium, Cymbopogon, Dichanthium, Heteropogon,
6 Dichanthium 105

Hyparrhenia, Schizachyrium, and Sorghastrum. In case of Dichanthium could be for lowering the lignin content
of individual trees, they were able to resolve a clade of while increasing the protein quality. The high lignin
Bothriochloa, Dichanthium, and Capillipedium, but in content of tropical forage plants reduces in vitro digest-
PHYB and GBSSI trees, Bothriochloa was found to be ibility and voluntary feed intake by ruminants. Dalton
united with Capillipedium, while in ndhF tree, it et al. (2003) reported three transgenic plants of
grouped with Dichanthium. This confirms their rela- D. annulatum containing hygromycin resistance gene,
tionships and interfertilities as suggested by Harlan while two of them contained b-glucuronidase gene
and de Wet (1963a) and Kellogg (2000). Giussani (GUS) after embryogenic calli were co-transformed
et al. (2001) studied the molecular phylogeny of the with two plasmids encoding either of these genes.
subfamily Panicoideae and also found Dichanthium– Hygromycin resistance varied from 68 to 100% in the
Bothriochloa–Capillipedium to represent together in progeny of these three transformants (Fig. 6.7).
cladogram when they mapped base chromosome num- Although DNA gun-mediated genetic transforma-
ber, anatomical characters including chloroplast struc- tion is the method of choice for range grasses (Dalton
ture, position, and number of bundle sheaths, and et al. 2003), which are recalcitrant to Agrobacterium
physiological characters including type of decarboxy- tumefaciens, it generally results in transgenics with
lating enzyme, viz. NADP-maleic enzyme. Rondeau multiple copies of a gene that may cause gene silencing,
et al. (2005) studied phylogenetic relationship among whereas the Agrobacterium-mediated transformation
the members of this tribe using an enzyme, NADP- method is cheaper, easier, more reliable, while only a
dependent malate dehydrogenase (NMDH). This single copy of a gene is delivered. Hence, a comparison
enzyme belongs to multigene family and is highly con- between these two methods in terms of their transfor-
served among plants. They found two distinct genes mation efficiencies would help in improving the level of
(NMDH-I and NMDH-II) encoding this enzyme, Agrobacterium-mediated transformation efficiency to
which would have arisen due to gene duplication asso- that of the gene gun-mediated transformation. Towards
ciated with selection pressure. They also speculated this, Kumar et al. (2005) compared frequency of GUS
presence of both genes in few species analyzed, e.g., expression between both methods of transformation
D. aristatum, H. contortus, etc. but could not detect the using binary vectors pCAMBIA1305 and pCAM-
transcripts of the latter because of its scarcity. BIA1301. Among two binary vectors used for Agro-
bacterium-mediated transformation, pCAMBIA1301
showed higher frequency of GUS expression while
6.6.4 Biotechnological Studies pCAMBIA1305 recorded more of the GUS spots per
in Dichanthium callus. Particle inflow gun-mediated transformation
resulted in higher GUS expression compared to the
Agrobacterium method using pCAMBIA1305. Further
Tissue culture and genetic transformation tools could optimization is necessary before using Agrobacterium-
be effectively used to generate somaclonal variation mediated transformation of Dichanthium.
and targeted gene transfers, respectively. Due to pre-
dominant apomictic mode of reproduction, variability
in this taxon can be heritably fixed in the subsequent
6.7 Primary Productivity and Biomass
generations. Hence, optimization of tissue culture pro-
tocol was a very essential initial step toward genetic Production
improvement of D. annulatum. In vitro plant regener-
ation has been reported from immature inflorescence Production, compartment transfer of dry matter, and
(Gupta et al. 1997), nodal explant (Gupta et al. 1998), aboveground production are higher in case of tropical
shoot tip (Dalton et al. 2003), and mature seeds grasslands compared to their temperate counterpart
(Kumar et al. 2005) of D. annulatum. (Misra and Mall 1975). Studies on dry matter, organic
Genetic transformation is a means to alter the matter, and crude protein content for rangeland grasses in
genome by introgression of genes from distant or unre- Pakistan show the highest relative preference for
lated grass species or by introduction of alien genes. D. annulatum than other rangeland grasses (Sultan
The desired genes for improving the nutritional quality et al. 2008). D. annulatum has been shown to produce
106 V. Bhat et al.

Fig. 6.7 Tissue culture, gene-gun-mediated transformation, pACT1-F. (f) Bombarded tissue surviving after 6 weeks of
regeneration, and progeny screening of transgenic Dichanthium selection on callus medium containing 75 mg/l hygromycin. (g)
plants. (a) Shoot tip-derived embryogenic calli (8 weeks old) on Root growth in transformed plants established in soil: tillers of
maltose-based medium (W1). (b) Shoot regeneration in 47 R1 and R10 (A) and non-transformed tillers (B) grown hydro-
weeks old callus on regeneration medium. (c) Callus after 5 ponically in a solution containing 50 mg/l hygromycin. (h)
weeks of growth on Hygromycin (0, 50 and 100 mg/l) containing Flowering in transformed Dichanthium plants. (i) Floral head
callus medium (top to bottom). (d) Transient GUS expression in of transformed plant. (j) Survival of seedlings of T1 progeny of
callus 48 h after bombardment with plasmid pACT1-F. (e) GUS plant R2 germinated on medium containing 50 mg/l hygromycin
expression 11 days after bombardment of callus with plasmid (right) compared with non-transformed control (left)

maximum organic matter at Magan grassland at Dhakar- sp. of D. annulatum range from 2–6 t/ha and even upto
wara (Trivedi and Mishra 1979). 17 t/ha, whereas for D. aristatum and D. caricosum, it
With reference to dry matter yields, data has been is in the order of 10–12 t/ha. In case of D. sericeum,
collected for the major forage crops of Dichanthium the yield is dependent upon the soil and climatic
6 Dichanthium 107

condition as D. sericeum has a wide distribution range as it is less desirable than the native species such as
(Cook et al. 2005). A good D. annulatum stand yields Cynodon nlemfuencis, Panicum maximum, Brachiaria
3 t/ha hay compared to an average of 9 t/ha in case of decumbens, and Digitaria decumbens.
D. aristatum (Cook et al. 2005). The growth performance like seed weight of
With regard to the animal production or carrying D. annulatum has also been found to decrease with
capacity, the four major cultivated crop species of increase in biotic disturbances, whereas the number of
Dichanthium have been studied thoroughly. D. annu- vegetative buds on rhizomes is found to increase, which
latum stand can support up to 7 sheeps/ha in semiarid results in higher biomass and productivity. It has also
environment. In case of D. aristatum and D. carico- been found that the habit of the plant changes from
sum, the live-weight gain of an animal is 270 kg/ha/yr erect to prostrate forms in case of protected and grazed
and 150 kg/ha/yr, respectively, when grown in combi- grounds, respectively (Ambasht and Maurya 1970).
nation with legumes. On the other hand, D. sericeum
has a comparatively low carrying capacity compared
to these species (Cook et al. 2005). Diseases and Pests
The next section discusses various factors, which
affect the productivity and biomass production in Many fungi have been reported to attack Dichanthium
Dichanthium sp. sp., but major damage has not been reported. Balansia
sclerotica, Cerebella andropogonis, Cochliobolus
cymbopogonis, Curvularia andropogonis, C. lunata,
C. robusta, C. gudauskasii, Phyllachora ischaemi (tar
6.7.1 Various Factors Affecting spot), Pithomyces graminicola, Puccinia cesatii,
Productivity and Biomass P. duthiae, P. propinqua, P. kenmorensis (rust), Scler-
ospora dichanthicola, Sphacelotheca annulata,
Production in Dichanthium
S. andropogonis-annulati, Striga lutea, Tolypospor-
ella obesa, Uredo susica, Uromyces andropogonis-
6.7.1.1 Biotic Factors annulati, and U. clignyi to name a few. Downy mildew
fungus, namely Sclerospora sorghi, has been identi-
Grazing and Herbage Removal fied as pathogens of D. caricosum in Thailand (GRIN).
In 1966, an undescribed species of Curvularia from
The effect of increase in the density or crowding of Texas (later named as C. robusta due to the large size
D. annulatum and little or no grazing showed initial of conidia) was identified to be pathogenic on
increase in yield but subsequent decrease in above- D. annulatum (Kilpatrick and Luttrell 1967). A list
ground herbage, biomass, and higher mortality of various smut fungi including Sporisorium
(Tripathi and Gupta 1980). dichanthii affecting D. aristatum; S. dichathicola
Studies on D. annulatum show high degree of pal- pathogenic on D. caricosum; and S. sahahyai (Wang
atability at all stages of its growth, and therefore, it is and Piepenbring 2002), Ustilago duthiei and
considered to be grazed preferentially compared to U. sabourieana on D. annulatum has been reported
other species. This study on relative palatability is Vánky (2004). Claviceps sp., which causes ergot dis-
useful for studying the impact of grazing on natural ease pose a significant hindrance in seed production in
grasslands and also in evolving suitable practices D. annulatum and D. aristatum. D. caricosum has been
required for their management (Dabadghao and Marwaha found to be susceptible to root-knot nematodes
1962). However, recent studies have shown that whereas D. aristatum acts as an alternate host for
grazing in rangelands causes weakening and death of sheath blight of rice caused by Rhizoctonia solani
native and introduced desirable species and allow less (Cook et al., 2005).
desirable invasive plants. For example, D. annulatum Apart from pathogenic fungi, grasslands support a
competes with native desirable species as it can persist rich fungal population, which exists in the mantle
grazing pressure, high temperature, and less water. around the roots. These rhizosphere fungi were found
Ortega-Santos et al. (2007) found that D. annulatum to effect seed germination and root growth of the
affects primary productivity of grazing lands for cattle seedlings (Leelavathy 1969a, b).
108 V. Bhat et al.

6.7.1.2 Abiotic Factors growth of D. annulatum seedlings have been found to

decrease with increase in salinity level (Akhtar and
Seasonal Burning Hussain 2008; Khan et al. 2009). It was found that the
moisture content in seedlings grown under saline con-
Fire is considered to be a powerful factor to ditions was more than control seedlings, which indi-
design community structure in rangelands and cating adaptive mechanism in grasses by becoming
to manipulate plant population. Burning has shown succulents in saline soils.
to stimulate the shoot–root growth of D. annulatum
but decrease in belowground biomass. It has been
found that burnt plots show higher productivity as
Soil Moisture
a result of improved nutrient status, less shading, and
presence of young tillers (Pandey 1988). Fire thus
Soil moisture stress along with clipping intensity has
influences the uptake and release of nutrients like
shown to increase the water use efficiency (WUE) in
NPK and in turn increases the mineral pool (Paulsamy
D. annulatum. As soil moisture stress increases, the
et al. 1995, 2005) encouraging higher productivity.
transpiration rate drops down. The drop in transpira-
tion rate is more pronounced in winters than in sum-
mers, which indicates that a strong moisture
Factors affecting Seed Germination
conservation mechanism operates in D. annulatum
(Singh and Misra 1985). The root:shoot ratio was
Various physical factors such as light, temperature,
found to be higher with increase in soil moisture stress,
and habitat condition affect seed germination of
and studies have indicated that dry matter production
D. annulatum. Seeds of D. annulatum are photosensi-
was maximum when water potential is between field
tive and show high germination rate during rainy sea-
capacity and saturation percentage (Singh and Sinha
son. Seed cover and increasing depth of sown seeds
1983). Except for nitrogen, concentration of all other
affects germination capability, whereas continuous
nutrients decrease with increase in soil moisture stress.
light and optimum temperature of 30 C shows maxi-
Nitrogen, potassium, and phosphorus use efficiencies
mum germination percentage in D. annulatum
increase with high soil moisture stress (Misra and
(Maurya 1972). On the other hand, the warm season
Singh 1982).
native perennial grass D. sericeum germinates usually
during mid-summer to early autumn period.

Shade
Seed Dormancy
Shade affects chlorophyll accumulation in D. annu-
In Dichanthium, seed dormancy is imposed by cover- latum, i.e., chlorophyll b increases but chlorophyll a
ing of the embryo (i.e., the seed coat) or within the decreases. An increase in chlorophyll (a þ b) shows
embryo. Smoke-stimulated germination and scarifica- adaptability of this grass under shade. Shade also
tion help to release dormancy due to seed coat, increases the accumulation of carotenoids, which
whereas exposure of seeds to fluctuating temperatures increases the quality of fodder (Baig et al. 2005).
releases dormancy imposed by the embryo in Shade has no effect on C and N influxes in D.
D. sericeum (Read and Bellaris 1999; Adkins et al. aristatum, but there is a preferential allocation of C
2002). and N under low irradiance, which changes shoot:
root ratio in turn affecting its photosynthetic capac-
ity. K and P accumulation also increases under
Salinity shade. D. aristatum was not able to use radiation
efficiently, which indicated that it is not adapted to
The germination percentage, plumule–radicle length, silvopastoral system where shade is more (Cruz
shoot–root dry weight, and in turn establishment and 1997a, b).
6 Dichanthium 109

6.8 Conclusion bution in Marvel grass [Dichanthium annulatum L. (Stapf)].

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.
Chapter 7
Eleusine

Susana S. Neves

7.1 Introduction economically important herbicide-resistant weeds

(Basu et al. 2004; Beckie 2007).
In spite of the importance of finger millet for so
Eleusine Gaertn. is a small genus (eight species) of the
many human populations, the development of genetic
grass family (Poaceae, subfamily Chloridoideae) that
resources and breeding efforts in this crop or its allied
includes the finger millet or ragi (Eleusine coracana
species have been very limited until recent years. This
subsp. coracana), a highly valuable crop that over the
chapter gives an overview on current knowledge and
centuries has guaranteed the survival of millions of
resources thus far developed in Eleusine, including
people, particularly in eastern and southern Africa and
species characterization, details on research achieve-
India (National Research Council 1996). In sub-
ments, and some limitations associated with some of
Saharan Africa and South Asia, millets are survival
the species, and a discussion on future prospects in
crops for poor people, and nearly 4 million hectares
conservation and utilization of these valuable plants.
(ha) of land are devoted to the cultivation of finger
millet: 3 million ha in India and 1 million ha in Africa
(Naylor et al. 2004). The grain of finger millet is used
for preparing diverse types of food (porridge, soup, 7.2 Taxonomy and Species
bread, cakes, malt, alcoholic beverages), is highly Characterization
nutritious and tasty, and has excellent storage qualities
(Hilu and De Wet 1976a; Barbeau and Hilu 1993;
The genus Eleusine includes eight species of annual
National Research Council 1996; Nout 2009). Finger and perennial herbs, mostly of African origin, with
millet and some other Eleusine species are also useful
only one species being native to South America
as forage grasses. Two species, E. floccifolia and
(E. tristachya). Eleusine is essentially a tropical and
E. jaegeri, are also used in Ethiopia for making bas- subtropical genus, with several of its species (E. floc-
kets and other household items (Phillips 1995;
cifolia, E. intermedia, E. jaegeri, E. kigeziensis,
Grassland Index 2009). However, the genus also
E. multiflora) being adapted to upland habitats (grass-
includes a very problematic and cosmopolitan weed, land and open forest or bushland), growing in alti-
commonly known as goosegrass (E. indica), consid-
tudes well above 1,000 m (Phillips 1972, 1995). Other
ered one of the worst weeds in the world (Holm et al.
species of Eleusine (E. coracana, E. indica, and
1977; Radosevich et al. 2007) and one of the ten most E. tristachya) grow in a wider range of open habitats,
from sea level to the highlands. The adaptation
to highland habitats is also evident in the crop
(E. coracana subsp. coracana), which grows better
S.S. Neves in higher altitudes than other tropical cereals – finger
Plant Cell Biotechnology Laboratory, ITQB, Instituto de millet is cultivated in Africa at altitudes between
Tecnologia Quı́mica e Biológica, Universidade Nova de Lisboa,
Apartado 127, 2781-901 Oeiras, Portugal 1,000 and 2,000 m and in Nepal up to at least
e-mail: [email protected] 2,400 m (National Research Council 1996).

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 113
DOI 10.1007/978-3-642-14255-0_7, # Springer-Verlag Berlin Heidelberg 2011
114 S.S. Neves

Eleusine species are herbaceous plants with flat- and after the thin pericarp that encloses the grain has
tened culms (stems) and a digitate or subdigitate inflo- been removed.
rescence (more rarely shortly racemose), which is Keys for identification of Eleusine species are
formed by (1–)2–10(–20) spikes, arranged into a ter- provided by Phillips (1972, 1974, 1995). The online
minal whorl or clustered at the top of the flowering database and software available through GrassBase
stem, sometimes with one or two spikes placed a little (Clayton et al. 2006-onwards) are also useful for inter-
lower in the axis (the finger-like appearance of the active identification of the world’s grass species.
terminal inflorescence is the reason why the crop is General descriptions of each Eleusine species are given
known as “finger millet”). Each spike has many biseri- below and in Table 7.1; emphasis is given to characters that
ate and overlapping (imbricate) spikelets that are help to discriminate species. Table 7.1 summarizes infor-
laterally compressed and usually disarticulate at matu- mation on species names, chromosome numbers, morpho-
rity, except in the crop where spikelets are persistent. logical characters, geographical distribution, uses and
The fruit (grain) of Eleusine is ornamented and is economic importance. Authors of species/taxon names
enclosed by a thin pericarp that is easily removed given in Table 7.1 are not repeated in the text.
from the grain when soaked in a drop of water (Phillips
1972, 1995).
The grain of grasses is usually smooth and has a
pericarp that is fused to the seed surface (a true cary-
opsis). Several grass genera have smooth grains with
7.2.1 E. coracana
free pericarp (e.g., Brachychloa S.M.Phillips, Chloris
Sw., Ochtochloa Edgew., Sclerodactylon Stapf, Spor- E. coracana is an annual allotetraploid (2n ¼ 4x
obolus R.Br.). However, the character combination of ¼ 36) that includes two distinct subspecies: subsp.
an ornamented grain with free pericarp is highly coracana (finger millet) and subsp. africana (wild
unusual in grasses (Phillips 1972), and it is found finger millet) (Hilu and De Wet 1976a; Hilu 1994).
only in Eleusine and four other chloridoid genera, Morphological, cytogenetic, and molecular evidence
Acrachne Chiov., Dactyloctenium Willd., Coela- suggests that cultivated finger millet (subsp. coracana)
chyrum Hochst. and Nees, and a few species of Spor- was domesticated from wild populations of E. cora-
obolus (for descriptions of grass genera, see Clayton cana subsp. africana (Chennaveeraiah and Hiremath
et al. 2006-onwards). These genera, however, can be 1974; Hilu and De Wet 1976a; Hilu and Johnson 1992;
easily distinguished from Eleusine based on inflores- Dida et al. 2008). The crop, with its stout spikes and
cence characters. Eleusine has spikes that typically large and globose grain, is easily distinguished from
terminate in fertile spikelets, and glumes and lemmas wild taxa of Eleusine, and, for that reason, some
(bracts) that are always awnless (i.e., do not have any authors prefer to treat cultivated and wild finger millet
awns or bristle-like prolongations). In contrast, the as separate species: E. coracana (the crop) and
spikes of Acrachne terminate in an abortive spikelet, E. africana (e.g., Hiremath and Salimath 1991; Phil-
and those of Dactyloctenium terminate in a bare lips 1995; Bisht and Mukai 2002).
pointed extension of the rhachis (axis of the spike); However, hybridization between wild (subsp. afri-
these two genera also have awned bracts. Coela- cana) and cultivated (subsp. coracana) populations
chyrum has a paniculate or racemose inflorescence occurs naturally and frequently, giving rise to many
(not digitate) and a grain that is concave-convex in morphological intermediates wherever the two popu-
cross-section; the grain in Eleusine is trigonous in lations meet (Kennedy-O’Byrne 1957; Mehra 1962;
section or globose (Phillips 1972, 1995). Sporobolus De Wet et al. 1984; Phillips 1995). Dida et al. (2008)
has paniculate inflorescences with 1-flowered spike- have also recently demonstrated by genotypic analysis
lets, while Eleusine always have several-flowered spi- of microsatellites that there is clear evidence of gene
kelets. As it is so uncommon among grasses, the flow between subsp. africana and subsp. coracana.
ornamentation of the Eleusine fruit is thus a very Therefore, as previously discussed (Neves et al.
useful character for identification of the genus, but 2005), the rank of subspecies more adequately reflects
the ornate grain surface (finely granular or striate) is the biological relationship between the cultivated
visible only with some magnification (10
or more) “E. coracana” and the wild “E. africana”.
Table 7.1 The species of Eleusine: names, chromosome numbers, morphological characters, geography, uses, and economic impact
Taxon Vernacular names Chromosome Diagnostic characters Geographical Uses and economic impact
number distribution
7 Eleusine

Eleusine coracana (L.) Finger millet, ragi, 2n (4x) ¼ 36 Annual; culms robust; leaves Mainly East and Cereal crop. Grain used to prepare
Gaertn. subsp. nachni, koddo, herbaceous (soft), glabrous, southern Africa and porridge, soup, bread, cakes, malt,
coracana dagussa, dagusha, sometimes with pilose margins; India, also in Nepal, and alcoholic beverages (beer,
tocussa, barancia, inflorescence digitate or China, and other liquors). Also used for forage, soil-
baranika, uembe, subdigitate; spikes 3–10(–20), thick South Asian retention, papermaking, and in
ulezi, wimbi, rapoko, and stout, incurved or straight, countries traditional medicine
etc 4–14 cm long, (7–)9–15 mm wide; (cultivated)
grain globose, black, brown,
reddish, or whitish
E. coracana (L.) Gaertn. Wild finger millet 2n (4x) ¼ 36 Annual; culms moderately robust; Tropical and southern Forage grass. Also a weed that when
subsp. africana leaves  soft, usually glabrous; Africa, mainly associated to finger millet fields
(Kenn.-O’Byrne) inflorescence digitate or eastern and southern may seriously reduce the yields of
Hilu and de Wet subdigitate; spikes (2–)4–17, uplands the crop
[Synonym: slender, straight, 4–17 cm long,
E. africana Kenn.- 4–8 mm wide; grain ovate-oblong,
O’Byrne] trigonous in section, black or
brownish
E. floccifolia (Forssk.) Akirma, akrma, dagoo, 2n (2x) ¼ 18 Perennial; culms moderately robust; Ethiopia, Eritrea, Used for making baskets (Ethiopia).
Spreng. garrgorr leaves  tough, with small tufts of Yemen, Somalia, Frequent in heavily grazed pastures
white hairs scattered along the Kenya of mid-altitude in Ethiopia, but
margins; inflorescence subdigitate; (1,500–3,200 m unpalatable to cattle
spikes 2–8(–10), slender, straight, altitude)
5–12 cm long, 3.5–6 mm wide;
grain elliptic to oblong, trigonous,
blackish
E. indica (L.) Gaertn. Goosegrass, crowsfoot 2n (2x) ¼ 18 Annual; culms slender; leaves soft, Cosmopolitan weed of Very problematic weed, difficult to
grass, silver glabrous to sparsely pilose; African origin; eradicate and resistant to multiple
crabgrass, wiregrass, inflorescence digitate or mostly tropics and herbicides.
paragis, grama de subdigitate; spikes (1–)2–10(–17), subtropics On the positive side: used as forage and
caballo, pata de slender, straight, 3.5–15.5 cm long, in traditional medicine in Africa and
gallina, etc. 3–6 mm wide; grain elliptic, Asia
trigonous, blackish
E. intermedia (Chiov.) – 2n (2x) ¼ 18 Perennial; culms moderately robust; Ethiopia, Kenya Unknown uses but possibly of value as
S.M. Phillips leaves herbaceous (not tough), (1,100–1,800 m forage grass
glabrous to pilose (no hair tufts); altitude)
inflorecence subdigitate or shortly
racemose; spikes 4–15, slender,
straight, 5–12 cm long, 4–8 mm
wide; grain broadly elliptic,
trigonous, blackish
(continued)
115
Table 7.1 (continued)
116

Taxon Vernacular names Chromosome Diagnostic characters Geographical Uses and economic impact
number distribution
E. jaegeri Pilg. Manyata grass, 2n (2x) ¼ 20 Perennial; culms robust; leaves tough Kenya, Tanzania, Used for making baskets. Frequent
mafutiana, akirma, (like leather) with rough (scabrid) Uganda, Ethiopia invader of highland pastures in East
dagoo, titima margins, glabrous; inflorescence (1,800–3,300 m Africa but avoided by livestock
subdigitate or shortly racemose; altitude) (unpalatable)
spikes 2–10(–13), slender, straight,
4–17 cm long, 3–7 mm wide; grain
elliptic-oblong, trigonous, blackish
E. kigeziensis S.M. – 2n (4x) ¼ 38 Perennial; culms moderately robust; Uganda, Congo, Possibly of value as forage grass
Phillips leaves  soft, glabrous beneath, Rwanda, Burundi,
sparsely pilose on the upper surface; Ethiopia
inflorescence digitate; spikes 2–7, (2,000–2,700 m
slender, straight, 7.5–14 cm long, altitude)
4.5–5.5 mm wide; grain elliptic,
trigonous, blackish
E. multiflora Hochst. ex – 2n (2x) ¼ 16 Annual; culms slender; leaves soft, Ethiopia, Eritrea, Potentially valuable as forage grass.
A.Rich. sparsely pilose (no hair tufts); Kenya, Tanzania Occasional weed
inflorescence racemose; spikes (1,500–3,000 m
(2–)3–8, more or less clustered at altitude)
the top of the axis, oblong to ovate,
straight or slightly curved, 1–4 cm
long, 8–16 mm wide; grain oblong
in profile, laterally compressed,
blackish
E. tristachya (Lam.) Three-spike goosegrass 2n (2x) ¼ 18 Annual; culms slender; leaves soft, South America Forage grass (Argentina). Also a weed
Lam. glabrous; inflorescence digitate; (introduced in other with increasing occurrence in many
spikes (1–)2–3, oblong, straight, regions) regions of the world
1–6(–8) cm long, 5–16 mm wide;
grain oblong-globose, trigonous in
section, blackish
Notes: Species/taxon information was obtained from Kennedy-O’Byrne (1957), Phillips (1972, 1974, 1995), Hilu and De Wet (1976a), Hansen (1980), Hilu (1980, 2003), National
Research Council (1996), Sisay and Baars (2002), Agnew (2006), Clayton et al. (2006-onwards), Chen and Phillips (2006), Lovisolo and Galati (2007), Agyare et al. (2009),
Grassland Index (2009), USDA NRCS (2009), and herbarium specimens (S. Neves, pers. observ.). Chromosome numbers were obtained from Hiremath and Chennaveeraiah (1982)
and Hiremath and Salimath (1991)
S.S. Neves
7 Eleusine 117

7.2.1.1 E. coracana subsp. coracana 7.2.1.2 E. coracana subsp. africana

The taxon E. coracana subsp. coracana corresponds This taxon was first recognized as a distinct species:
to cultivated finger millet and, in the literature and E. africana (Kennedy-O’Byrne 1957). Later,
current databases, is more frequently referred simply “E. africana” was considered a subspecies of E.
as E. coracana. Finger millet is easily recognized by its indica in view of morphological similarities that
typical digitate or subdigitate inflorescences, with thick were thought to result from hybridization between
and stout spikes (Fig. 7.1a), which are often incurved, these two taxa [see: “E. indica subsp. africana” in
and its globose grain that may be blackish, brown, Phillips (1972, 1974) and Lye (1999)]. However, this
reddish, or even whitish; the grain of other Eleusine information was not confirmed as attempts to produce
species/taxa is typically blackish (visible after pericarp hybrids between the tetraploid “E. africana” and the
removal) (Phillips 1972, 1974, 1995). Other diagnostic diploid E. indica resulted in sterile plants (Chenna-
characters of cultivated finger millet are the persistent veeraiah and Hiremath 1974; Hiremath and Salimath
(non-shattering) spikelets and the exposed grains (visi- 1992). In contrast, as mentioned above, “E. africana”
ble in the gaping floret when ripe); in other Eleusine readily hybridizes with finger millet (subsp. coracana)
species/taxa (including subsp. africana), spikelets dis- and, for that reason, is regarded here as a subspecies
articulate (between the florets) at maturity and grains of E. coracana.
remain enclosed (Phillips 1972, 1995; Chennaveeraiah The tetraploid E. coracana subsp. africana
and Hiremath 1973; Chen and Phillips 2006). Morpho- (2n ¼ 36) may be confused with the diploid E. indica
logical variation of the inflorescence in finger millet is (2n ¼ 18). As Kennedy-O’Byrne (1957) pointed out,
considerable and, for that reason, several races have most of the morphological differences between these
been described in the literature (Hilu and De Wet two closely related species are differences in size, with
1976b; De Wet et al. 1984; Dida and Devos 2006; the tetraploid (E. coracana subsp. africana) usually
Upadhyaya et al. 2007). The five races more frequently being a more robust plant, with longer leaves, thicker
recognized are: “coracana,” “elongata,” “plana,” “com- and longer spikes, larger bracts, etc., than its diploid
pacta,” and “vulgaris”; for descriptions and illustrations ally (E. indica). However, identification can be diffi-
of these races, see De Wet et al. (1984) and Dida and cult when these two taxa present similar sizes. Chro-
Devos (2006). A review on the origin and evolution of mosome numbers and genome size clearly distinguish
the crop is presented in Sect. 7.5 of this chapter. E. coracana subsp. africana from E. indica, but

Fig. 7.1 Types of inflorescence in Eleusine: (a) digitate: E. coracana subsp. coracana (finger millet). (b) racemose: E. multiflora.
Photos: (a) S. Neves; (b) G. Swire-Clark and V. Baird
118 S.S. Neves

cytogenetic analysis is usually not an option for regular 7.2.4 E. intermedia

identification. Phillips (1972) suggested that the ligule
and grain characters may help to differentiate the two
E. intermedia is a perennial diploid (2n ¼ 2x ¼ 18)
species (the ligule is a leaf structure in the shape of a
with very restricted distribution in the uplands of
membrane or fringe of hairs that is located at the base
northern Kenya and southern Ethiopia (Phillips
of the leaf blade). In E. coracana subsp. africana, the
1972). This species was first described as a variety of
ligule has a clear ciliate fringe and the grain surface is
E. indica but later recognized by Phillips (1972) as a
finely granular with low ridges (barely visible),
distinct species. E. intermedia can be easily distin-
whereas in E. indica, the ligule is a truncate, scarcely
guished from E. indica because of its perennial habit
ciliolate membrane and the grain has well-marked,
(it has a stout rhizome), more laxly arranged spikelets,
oblique ridges/striae (magnification of 50
or more is
and 3-nerved lemmas with 1-nerved keel, whereas
needed to observe the fine details of grain surface in
E. indica is an annual, with tightly arranged spikelets,
any Eleusine species) – drawings of the ligules and
and 3-nerved lemmas with a 3-nerved keel (Phillips
SEM (scanning electron microscope) photos of the
1972). E. intermedia may be confused with other
grains of these species are shown in Phillips (1972)
perennials of the genus, particularly E. jaegeri, but
and Hilu et al. (1979).
the latter has tough, glabrous leaves with rough mar-
gins, whereas E. intermedia has softer leaves, slightly
7.2.2 E. floccifolia pilose, with generally smooth margins (Phillips 1972).
Uses of E. intermedia are unknown, but the species
may have some value as a forage grass when other
E. floccifolia is a perennial diploid (2n ¼ 2x ¼ 18)
grazing is scarce (Grassland Index 2009).
that can be easily recognized within genus by its
unique small tuffs of white hairs that are scattered
along the smooth margins of the leaves (Phillips
1972, 1974, 1995). The species is found in mid and
high altitude in NE Africa and Arabia, and is 7.2.5 E. jaegeri
reported as dominant in heavily grazed pastures of
mid-altitude in Ethiopia (Sisay and Baars 2002), pos- E. jaegeri is a perennial diploid (2n ¼ 2x ¼ 20) and the
sibly a consequence of being unpalatable and most robust species of Eleusine. It can be easily distin-
avoided by livestock. The species, however, is guished from other species in the genus by its robust
much used by local populations, particularly in habit and stiff, pale green leaves, with rough (saw-
Ethiopia, for making baskets and other household edged) margins; the plants of the species form dense
craft articles (Phillips 1995). and coarse tussocks (clumps), with its culms (stems)
branching to form thick bunches of whitish, overlapping
leaf-sheaths (Phillips 1972, 1974, 1995; Grassland
Index 2009). E. jaegeri is common in the grasslands of
7.2.3 E. indica
the East African highlands and is used by locals for
basket making; however, the species is unpalatable to
E. indica (goosegrass) is an annual diploid (2n ¼ livestock and is invasive in heavily grazed pastures of
2x ¼ 18), which is a major weed problem in many high altitude (Grassland Index 2009).
countries in the world, mostly in the tropics, but now
affecting temperate countries as well (see Sect. 7.8.2
for further discussion on this topic). E. indica may
sometimes be confused with the closely related
7.2.6 E. kigeziensis
E. coracana subsp. africana, an annual tetraploid
that is usually a more robust plant; see Sect. 7.2.1.2 E. kigeziensis is a perennial tetraploid (2n ¼ 2x ¼ 38)
above for details on how to differentiate these two with a restricted distribution in the East African high-
species. lands (Phillips 1972). The inflorescence of E.
7 Eleusine 119

kigeziensis resembles that of the annuals E. coracana other species in the genus with similar short and
subsp. africana and E. indica, with which sometimes broad spikes (Phillips 1972, 1974, 1995). However,
is confused; but it is distinguished from these taxa by E. tristachya has a digitate inflorescence with (1–)2–3
its perennial habit (E. kigeziensis has a short ascending spikes that are tightly clustered to each other at the top
rhizome that is absent in the annuals). E. kigeziensis is of the axis, whereas E. multiflora usually has three or
separated from other perennials by its slender rhizome more spikes that clearly alternate in the axis, the lower
(not stout), fairly soft leaves, which are sometimes ones often separated from each other by a distance of
pilose, but with no tuffs of hairs in the leaf margins, 1 cm or more.
and by its lemmas with a central 3-nerved keel (i.e., Phillips (1972) and Clayton and Renvoize (1986)
the lemma has a central nerve with two lateral, incon- considered that E. multiflora represented a link
spicuous nerves); the midnerve of the lemmas is between the genus Acrachne and Eleusine, because
always simple (1-nerved) in other perennial species the species presents some morphological features
(E. floccifolia, E. intermedia, E. jaegeri, and E. jaegeri) that appear intermediate between the two genera.
(Phillips 1972, 1974, 1995). The placement of E. multiflora in the genus was also
questioned by other authors based on: flavonoid con-
tent (Hilu et al. 1978), restriction fragment length
polymorphisms (RFLPs) of ribosomal DNA (Hilu
7.2.7 E. multiflora and Johnson 1992), genome size when compared to
that of other diploids (Mysore and Baird 1997; see also
E. multiflora is an annual diploid (2n ¼ 2x ¼ 16) that Sect. 7.3), and cytogenetic pattern of ribosomal DNA
is easily recognizable within the genus because of its sites (Bisht and Mukai 2000). On the other hand,
short, oblong-ovate spikes that alternate at the top of isozymes (Werth et al. 1994) and chloroplast DNA
the inflorescence axis (Fig. 7.1b). E. multiflora has restriction site data (Hilu and Johnson 1997) supported
been sometimes confused with E. tristachya, the only the inclusion of E. multiflora in the genus. If any

Fig. 7.2 Phylogeny of the genus Eleusine. Simplified phyloge- (2005). The B genome sequences of E. coracana refer only to
netic tree of the combined sequences of the nuclear ITS ribo- ITS data. Eleusine kigeziensis (1) corresponds to the accession
somal DNA and plastid trnT-trnF regions; data from Neves et al. “Kew 12560”; E. kigeziensis (2) corresponds to “K. Hilu 2505”
120 S.S. Neves

doubts remained, recent sequence analyses of plastid Neves et al. 2005). Therefore, there is currently no
and nuclear DNA have unequivocally demonstrated evidence that this species exists in the wild.
that E. multiflora is a member of the Eleusine lineage Some species that were originally described in
(Neves et al. 2005); see the placement of E. multiflora Eleusine are now classified in other genera, such as
in the phylogeny of the genus in Fig. 7.2. Acrachne and Ochthochloa. For example, Eleusine
racemosa B.Heyne ex Roem. and Schult. and Eleusine
verticillata Roxb. are presently recognized as
Acrachne racemosa (B.Heyne ex Roem. and Schult.)
7.2.8 E. tristachya Ohwi; whereas Eleusine compressa (Forssk.) Asch.
and Schweinf. ex C.Chr. and Eleusine flagellifera
E. tristachya is an annual diploid (2n ¼ 2x ¼ 18) that Nees currently correspond to Ochthochloa compressa
is easily recognizable in the genus by its digitate inflo- (Forssk.) Hilu (Hilu 1981; Phillips 1974, 1995).
rescence with (1–)2–3 oblong spikes (see spike mea-
surements in Table 7.1), which are tightly clustered at
the top of the axis, and its neatly arranged spikelets that
are perpendicular to the spike axis (Hilu 1980). The 7.3 Morphological, Classical Genetic
species may occasionally be confused with E. multi- and Cytogenetic Studies in the
flora (see Sect. 7.2.7, above). E. tristachya is the only Genus Eleusine
non-African member of the genus, being native to
South America, where it is important as a forage
grass, particularly in Argentina (Lovisolo and Galati From the late nineteenth century until the 1950s,
2007). However, this species has now a much larger researchers supported the notion that E. indica was
distribution and is naturalized in many regions of the the direct ancestor of cultivated finger millet, and that
world, including parts of North America, Europe, domestication had likely occurred in India, with the
Africa, and Australia (see e.g., Hansen 1980; Hilu crop being later introduced in Africa (e.g.,. Greenway
1980, 2003; Sanz Elorza et al. 2001; Ellis et al. 2004; 1944). However, the finding of a new African tetra-
Clayton et al. 2006-onwards; USDA NRCS 2009). ploid species (“E. africana”), with close resemblance
to the crop, led to the review of the theories of origin of
finger millet. Kennedy-O’Byrne (1957) was the first to
suggest that domestication may have occurred in North
East Tropical Africa, with the diploid E. indica given
7.2.9 Species of Uncertain Status or Now
origin to the wild tetraploid “E. africana” by chromo-
Placed in Other Genera some doubling and, subsequently, to the development
of the crop by selection and cultivation of a large grain
The list of Eleusine species addressed in this chapter mutant of “E. africana.” Supporting the close relation-
does not include E. semisterilis S.M.Phillips, a species ship between “E. africana” and E. coracana sensu
name published based on a single herbarium specimen stricto (s.str.) were the large number of morphological
(holotype) located at the Kew herbarium, in England intermediates between these taxa that were found in
(Phillips 1972). This specimen presents a number of cultivated fields in Africa (Kennedy-O’Byrne 1957;
unusual features for an Eleusine species (abortive Mehra 1962). Initially, E. indica was thought as the
spikelet at the end of the spikes and laxly arranged only contributor to the genome of finger millet (e.g.,
spikelets), and even Phillips (1972) suggested that the Greenway 1944; Kennedy-O’Byrne 1957). Later,
atypical inflorescence of the specimen may be the result Mehra (1963a, b), in a comparative morphological
of anomalous development. All subsequent references analysis of E. coracana (s.str.), “E. africana,” and
to this species (e.g., Clayton et al. 2006-onwards and E. indica, supported the view that E. coracana had
Grassland Index 2009) continue to be based on that originated in Africa from “E. africana” and that the
single specimen. Furthermore, attempts by K.W. Hilu latter had evolved by hybridization of E. indica and
to find more material in the Kenyan locality where the another taxon. Subsequent morphological and archeo-
type was collected have failed (Werth et al. 1994; logical studies further confirmed the origin of finger
7 Eleusine 121

millet in East Africa, and that E. coracana subsp. E. indica
E. floccifolia and E. tristachya
E.
africana is wild finger millet (Harland 1971; Hilu and indica. The resulting hybrids (F1) were analyzed cyto-
De Wet 1976a; Hilu et al. 1979). genetically, with demonstration of strong homology
Until the early 1970s, most cytogenetic work in among the genomes of E. indica, E. tristachya, and
Eleusine concerned chromosome counts [reviewed by E. floccifolia. However, in spite of the genomic simi-
Hiremath and Chennaveeraiah (1982)]. Chennaveer- larity of the parental species, all the hybrids obtained
aiah and Hiremath (1973, 1974) started a series of were once again completely seed sterile (Salimath et al.
comparative cytogenetic studies intended to elucidate 1995b). More recently, a similar study of hybridization
the genomic origin and relationships of finger millet between diploid species was performed by Devaru-
with its wild relatives. Chennaveeraiah and Hiremath math et al. (2005), the main difference being the inclu-
(1973) analyzed chromosome pairing during meiosis sion of E. intermedia and the production of F1 hybrids
of a hybrid between the annual E. tristachya and the between this species and E. indica, E. tristachya, and
perennial E. floccifolia, two diploid species with the E. floccifolia. This study reported high degree of
same chromosome number (2n ¼ 18). These authors homology among the four species but, as in earlier
observed good pairing between the chromosomes of studies, reported a lack of fertility in the F1 hybrids.
these species, but the resulting hybrid was completely
sterile. Chennaveeraiah and Hiremath (1974) used sim-
ilar cytogenetic methods to analyze genome homology
in two sets of hybrids, E. coracana s.str.
“E. afri- 7.3.1 FISH and GISH Analyses
cana” and E. coracana s.str.
E. indica; these authors
demonstrated that the genomes of “E. africana” and E. Bisht and Mukai (2000) used for the first time fluores-
coracana (s.str.) are basically the same and that these cence in situ hybridization (FISH) in Eleusine species
two taxa are allotetraploids with AABB genomes. to map and compare the distribution of ribosomal
Chennaveeraiah and Hiremath (1974) also suggested DNA (rDNA) sites in their genomes. This work further
that, due to lack of homology, E. indica may not be a confirmed the similarity of the genomes of cultivated
genome donor to finger millet, but these authors were E. coracana (s.str.) and wild “E. africana” as the two
cautious about their own results, which were based on taxa presented similar location and number of 18S-
analyses of a single hybrid and a small number of cells. 5.8S-26S and 5S rDNA sites on their chromosomes.
After chloroplast DNA evidence (see Sect. 7.4.1) Bisht and Mukai (2000) also found some similarities
demonstrated that E. indica is the “A” (maternal) in the patterns of rDNA sites of E. indica and
genome donor to the crop (Hilu 1988), new cytogenetic E. floccifolia when compared to those seen in E. cor-
analyses (chromosome pairing in hybrids) were carried acana (s.str.) and “E. africana” and suggested that
out by Hiremath and Salimath (1992). These authors E. indica and E. floccifolia may be the two genome
performed several crosses between tetraploids (E. cor- donors to the crop. In another study, Bisht and Mukai
acana s.str. or “E. africana”) and diploids (E. indica, E. (2001a) used genomic in situ hybridization (GISH)
floccifolia, E. intermedia, E. multiflora, and E. trista- techniques to hybridize genomic DNA of six diploid
chya) to obtain triploid hybrids (found to be sterile). species of Eleusine (E. indica, E. floccifolia, E. trista-
Hiremath and Salimath (1992) then confirmed that the chya, E. internedia, E. multiflora, and E. jaegeri) to
genome of E. indica has strong homology to (half of) the chromosomes of the tetraploid E. coracana s.str.
the chromosomes of E. coracana (s.str.), contradicting (finger millet). The genomic DNA of E. multiflora and
the initial finding of lack homology between these E. jaegeri did not produce any hybridization signals in
species (Chennaveeraiah and Hiremath 1974). Hire- the chromosomes of E. coracana s.str., suggesting that
math and Salimath (1992) thus confirmed that the two species are not closely related to finger millet
E. indica is the “A” genome donor of the crop but and may be ruled out as genome donors (Bisht and
found no evidence for a possible “B” genome donor Mukai 2001a). This study also indicated a close geno-
among the species they analyzed. In a subsequent mic relationship among four diploids (E. indica,
study, Salimath et al. (1995b) attempted to produce E. floccifolia, E. tristachya, E. internedia) and that
hybrids between all the diploid species of Eleusine, the genomes of E. indica and E. tristachya are very
but crosses were only successful in two combinations: similar, with nearly identical patterns of hybridization
122 S.S. Neves

to the chromosomes of E. coracana s.str. (Bisht and (1991) using Feulgen microspectrophotometry. Later,
Mukai 2001a). In a separate publication, Bisht and Mysore and Baird (1997), using laser flow cytometry,
Mukai (2001b) reported the results of a similar GISH determined that there had been an overestimation on
study using the four diploids they previously found to nuclear DNA contents for most species analyzed by
be closer to finger millet but whose genomic DNA was Hiremath and Salimath (1991). The new genomic sizes
now hybridized to the chromosomes of “E. africana” determined by Mysore and Baird (1997) for the Eleu-
(wild finger millet). The two studies (Bisht and Mukai sine species/taxa were as follows (1 pg ¼ 980 Mbp ):
2001a, b) reported similar hybridization patterns E. coracana subsp. coracana, 2C ¼ 3.36–3.87 pg
between the genome of the diploids and the chromo- (2n ¼ 36); E. coracana subsp. africana, 2C ¼ 3.34 pg
somes of E. coracana s.str. and “E. africana,” support- (2n ¼ 36); E. indica, 2C ¼ 1.61–1.76 pg (2n ¼ 18);
ing the view that these two tetraploids are highly E. tristachya, 2C ¼ 1.51 pg (2n ¼ 18); E. floccifolia,
similar at the genomic level. Taking into account the 2C ¼ 2.0 pg (2n ¼ 18); E. multiflora, 2C ¼ 2.65 pg
results of double GISH on chromosomes of E. cora- (2n ¼ 16); and E. jaegeri, 2C ¼ 1.90 pg (2n ¼ 20).
cana (of the two subspecies) using the genomic DNA Mysore and Baird (1997) highlighted the fact that, in
of E. indica and E. floccifolia, Bisht and Mukai spite of having the smallest chromosome number in the
(2001a, b) suggested that these two species are the genus, E. multiflora has a considerably larger genome
genome donors to finger millet. However, as discussed than any of the other diploids.
in Sect. 7.4.1, recent sequence data contradicts the Although the genome size of the Eleusine species is
hypothesis that E. floccifolia is a genome donor to small when compared with that of many other plants,
finger millet (Neves et al. 2005). The contradiction including other grasses and important crops (see e.g.,
between the results of GISH and sequence analyses Bennett and Leitch 2005), it is nonetheless considered
is only apparent, because GISH results cannot be reli- relatively large from a genetics or genomics point of
ably used for phylogenetic inference (Neves et al. view (Dida et al. 2007). Nevertheless, a genetic map
2005). GISH techniques are certainly useful for for finger millet has already been produced (Dida et al.
assessment of chromosome genetic similarity, but 2007), demonstrating that genome size is not an
genomic comparisons can only be performed using impediment for detailed understanding of the genome
two or three samples (species) at a time, and as the of the Eleusine species.
results cannot be expressed quantitatively (no mea-
surements of genetic distance), they are problematic
to interpret at the phylogenetic level, particularly
among closely related species that naturally have 7.4 Evolution of the Genus Eleusine:
some degree of genomic similarity (Neves et al. Molecular Evidence
2005). Bisht and Mukai (2001a) obtained almost iden-
tical results of double GISH with E. coracana and the
species pairs, E. indica–E. floccifolia and E. trista- 7.4.1 Eleusine Evolution: The DNA Data
chya–E. floccifolia. If only the GISH results were
taken into account, E. tristachya could as easily be Previous work in Eleusine using morphology (Mehra
considered one of the genome donors as E. indica 1962, 1963a; Hilu and De Wet 1976a), phytochemistry
(Neves et al. 2005). But, of course, this was not sug- (Hilu et al. 1978), and cytogenetics (Chennaveeraiah
gested by Bisht and Mukai (2001a) who relied on the and Hiremath 1974) had demonstrated high similarity
results of earlier studies (e.g., Hilu 1988) to exclude between cultivated finger millet (E. coracana s.str.)
E. tristachya as a potential genome donor. and wild “E. africana,” that these two taxa hybridize
in nature, and that the crop was probably an allotetra-
ploid, with the diploid E. indica a possible parental
species. However, Chennaveeraiah and Hiremath
7.3.2 Genome Size (1974) had raised some questions about the status of
E. indica as a genome donor to the crop. With the
Analyses of nuclear DNA content in the species of advent of molecular biology techniques that could be
Eleusine were first carried by Hiremath and Salimath easily used by plant scientists, it became possible to
7 Eleusine 123

use data directly from DNA to address questions on (subsp. coracana and subsp. africana), with the diploid
relationships and genome homology that had remained E. indica sharing an ITS locus “A” sequence closer to
unsolved by more classical approaches. that found in both wild and cultivated finger millet. The
The first DNA analyses in the genus were carried ITS locus “B” sequence of E. coracana was not found in
out by Hilu (1988), who used chloroplast DNA any other Eleusine species. As sequences of the two ITS
(cpDNA) restriction fragment length polymorphism loci are clearly nested within the Eleusine lineage (see
(RFLP) to examine the relationships among cultivated Fig. 7.2), the two genome donors must be members of the
and wild finger millet and two other species in the genus; this leads to the conclusion that the “B” genome
genus: E. indica and E. tristachya. Hilu (1988) con- donor may now be extinct (Neves et al. 2005). E. indica
firmed the close genetic relationship between E. cor- shares with E. coracana highly similar chloroplast
acana subsp. coracana and its putative wild sequences, confirming that the former is the maternal
progenitor E. coracana subsp. africana and demon- genome donor of the crop; the high sequence similarity
strated that E. indica is the maternal (“A”) genome of these taxa also suggests a relatively recent allopoly-
donor of finger millet, with the three taxa sharing a ploidization event (Neves et al. 2005). However, this
common chloroplast genome (maternally inherited in study contradicts the hypothesis of Bisht and Mukai
most plants). Later, Hilu and Johnson (1992) used (2000, 2001a, b) that E. floccifolia is the second genome
RFLPs to study the variation of ribosomal DNA donor. The ITS sequences obtained by Neves et al.
(rDNA) among six species of Eleusine; the rDNA (2005) for E. floccifolia are very distinct from those of
data confirmed the close relationship among the two the putative “B” genome donor (see Fig. 7.2). Earlier,
subspecies of E. coracana and E. indica, with E. Salimath et al. (1995a) had also considered E. floccifolia,
tristachya being the next closest species to the crop an unlikely “B” genome donor based on DNA finger-
and E. multiflora the most distant. Similar results were printing analyses. The results of Neves et al. (2005) also
obtained by Hilu (1995) using random amplified poly- show good support for a “CAIK” group (E. coracana
morphic DN (RAPDs), although the samples of wild subsp. coracana and subsp. africana, E. indica, and
finger millet (E. coracana subsp. africana) grouped E. kigeziensis), with the South American E. tristachya
closer to E. indica than to the cultivated cereal. Sali- as its sister lineage. The perennials E. jaegeri and
math et al. (1995a) used three different DNA marker E. multiflora are shown as the earliest diverging lineages
techniques, RFLP, RAPD, and inter-simple sequence in the genus and the most distant to the crop (Neves et al.
repeats (ISSR), to analyze five species of Eleusine; 2005). In this study, several Eleusine species (for which
their results supported the close affinity of E. cora- more than one accession was sequenced) were shown as
cana, E. indica, and E. tristachya, with E. floccifolia monophyletic taxa, with the exception of E. kigeziensis,
being a more distant relative. Hilu and Johnson (1997) where an accession from Uganda (Kew 12560; see
extended earlier restriction site analyses of cpDNA in Fig. 7.2) did not group with the others (KH2505,
the genus to demonstrate the monophyly of Eleusine, KH2506, and KH2507) (see Neves et al. 2005).
with the inclusion of E. multiflora, a species whose Hiremath and Salimath (1991) and Bisht and Mukai
placement in the genus had been previously ques- (2002) suggested an allotetraploid origin for E. kigezien-
tioned (see Sect. 7.2.7 for more details). sis (2n ¼ 38), possibly from hybridization of the
Monophyly of Eleusine has been conclusively diploids E. indica (2n ¼ 18, x ¼ 9) and E. jaegeri
demonstrated by analyses of DNA sequences of the (2n ¼ 20, x ¼ 10). However, in spite of extensive
trnT–trnF (chloroplast genome) and the internal tran- clone sequencing, Neves et al. (2005) only found one
scribed spacer (ITS) region of nuclear rDNA in the type of ITS sequence in E. kigeziensis, raising the possi-
eight species that constitute the genus (Neves et al. bility of this species being an autopolyploid.
2005). A simplified molecular phylogenetic tree repre- E. kigeziensis belongs to a clade (lineage) of species
senting data and results of Neves et al. (2005) is shown in that has x ¼ 9 as basic chromosome number (E. flocci-
Fig. 7.2. This study further confirmed the allotetraploid folia, E. intermedia, E. tristachya, E. indica, and
origin of finger millet, since two putative ITS home- E. coracana; see Fig. 7.2). If E. kigeziensis is an auto-
ologs, corresponding to the “A” and “B” genomes, polyploid, it may have originated from polyploidization
were detected in all the accessions of E. coracana of an ancestral species with x ¼ 9 that subsequently
124 S.S. Neves

underwent chromosome rearrangement and/or aneu- 7.5 The Origin of Finger Millet:
ploidy, with a slight increase on chromosome number. Current Understanding

Cultivated finger millet (E. coracana subsp. coracana)

7.4.2 Phylogenetic Placement of Eleusine is believed to have been domesticated in East Africa,
Among Other Grasses probably in the highland region that extends from
Ethiopia to Uganda, where populations of wild finger
The placement of Eleusine in subfamily Chloridoideae millet (subsp. africana) are particularly abundant (Har-
is undisputed (e.g., Clayton and Renvoize 1986; Hilu land 1971; Hilu and De Wet 1976a, b; Hilu et al. 1979;
and Alice 2001), but its exact position in the chloridoid De Wet et al. 1984); E. coracana subsp. africana rarely
clade or its relationships to other chloridoid genera extends to the lowlands of East Africa, and hence the
remain uncertain (Hilu and Alice 2001; Columbus suggestion of an upland origin for the crop (Hilu and
et al. 2007; Bouchenak-Khelladi et al. 2008). Due to De Wet 1976b). The oldest archeological record for
some morphological similarities, Eleusine was previ- finger millet (well preserved grains obtained from a
ously considered a close ally of Acrachne and Dacty- prehistoric site in Ethiopia) has been tentatively dated
loctenium (Phillips 1972, 1982, 1995; Clayton and to the third millennium BC (Hilu et al. 1979), and this
Renvoize 1986). However, current molecular data suggests that domestication of the crop may have
indicates that Dactyloctenium (Hilu and Alice 2001; occurred in East Africa at about 5,000 years ago or
Roodt-Wilding and Spies 2006; Columbus et al. 2007) earlier (De Wet et al. 1984). The crop was subsequently
and Acrachne (Neves et al. 2005) are not closely introduced to India, possibly during the second or first
related to Eleusine. As mentioned in Sect. 7.2.9, millennium BC, where a secondary center of diversity
the genus Ochthochloa now includes a species was developed as a consequence of prolonged artificial
(O. compressa ¼ “E. compressa”) previously placed and natural selection (Hilu and De Wet 1976a, b;
in Eleusine. But Ochthochloa and Eleusine are mor- De Wet et al. 1984).
phologically quite distinct (Hilu 1981; Phillips 1982, Conclusive data from morphology, hybridization
1995); Hilu et al. (1978) also reported a very distinct studies, cytogenetics, and molecular biology indicate
flavonoid composition for “E. compressa”. Neves that the wild E. coracana subsp. africana (native in
et al. (2005) attempted to obtain seeds for O. com- Africa and originally absent in India) is the direct pro-
pressa for cultivation and DNA sequencing, but all genitor of cultivated finger millet (Chennaveeraiah and
accessions received had been incorrectly identified Hiremath 1974; Hilu and De Wet 1976a; Hilu and
(examples like this highlight the importance of con- Johnson 1992; Dida et al. 2008). Both cultivated and
firming the identification of plants before analyses are wild E. coracana have been confirmed as allotetra-
carried out). As there is no sequence data yet for ploids, with the genome notation given as AABB (Chen-
Ochthochloa, its relationship to Eleusine still needs naveeraiah and Hiremath 1974; Hilu 1988;
be assessed. In a numerical analysis of morphological Hiremath and Salimath 1992; Werth et al. 1994; Bisht
data, Phillips (1982) identified a group comprised by and Mukai 2001a, b; Neves et al. 2005; Dida et al. 2007).
Eleusine, Acrachne, Dactyloctenium, Coelachyrum, E. coracana subsp. africana (2n ¼ 36) is an allotetra-
and Sclerodactylon. Coelachyrum is another grass ploid that resulted from natural hybridization between
genus with ornamented grain and free pericarp (see the diploid E. indica (2n ¼ 18), considered the maternal
also introductory text in Sect. 7.2). However, sequence (“A”) genome donor (Hilu 1988; Hiremath and Sali-
data indicates that Coelachyrum is not closely related math 1992; Bisht and Mukai 2001a, b), and an unknown
to Eleusine (Ingram and Doyle 2004). Currently, there diploid (probably also 2n ¼ 18), followed by polyploi-
is no sequence data available in GenBank for Scler- dization. Sequences of both “A” and “B” genomes have
odactylon, but this genus is morphologically quite been identified in finger millet (see Sect. 7.4.1); the “A”
distinct from Eleusine. In summary, current evidence genome sequences are highly similar to those of E.
suggests that Eleusine is a monophyletic group that is indica, but the “B” genome sequences have not been
relatively isolated from other chloridoid grass genera identified in any of the known Eleusine species (Neves
(Hilu and Johnson 1997; Neves et al. 2005). et al. 2005); see also Fig. 7.2. For that reason, the “B”
7 Eleusine 125

genome progenitor of E. coracana may correspond to a Salimath et al. (1995a) used ISSRs in the genus
species that is now extinct (Neves et al. 2005). and found the technique to be particularly promising
for analysis of plant diversity, including differentia-
tion of accessions within the same species of Eleu-
sine. The ISSR technique produces dominant markers
7.6 Genome Analysis in Eleusine: and is considered useful due to its low cost, good
Molecular Tools and Genomic reproducibility, and its ability to detect high levels of
Resources variation at the population level (e.g., Salimath et al.
1995a; Wolfe et al. 1998). ISSRs may be particularly
useful for a first assessment of genetic diversity in
7.6.1 Molecular Tools: SSRs, AFLPs,
species where microsatellites have not yet been
ESTs, and Others developed.
Breviario et al. (2007) have recently used a mod-
Several fingerprinting techniques have been used to ified version of the TBP (tubulin based polymorphism)
analyze the species of Eleusine. RFLPs were the first method for analyses of diversity and species relation-
DNA markers to be used in the genus (Hilu 1988; Hilu ships in Eleusine. The new “combinatorial TBP”
and Johnson 1992; Salimath et al. 1995a) and are still (cTBP) technique was evaluated as reliable, reproduc-
used in current studies, namely for genetic mapping ible, fast, and easy to use, and able to produce enough
(Dida et al. 2007). The RFLP technique (e.g., Jones polymorphisms to discriminate species and varieties
et al. 1997) is a reliable technique that produces (Breviario et al. 2007). The cTBP technique was able
codominant markers but is time-consuming and may to detect variability in individual accessions of finger
sometimes show little or no variation among closely millet but failed to detect variation in other crops,
related individuals (accessions), highly inbred lines, or suggesting that the number of polymorphisms gener-
in germplasm collections with low genetic diversity. ated by the technique may be insufficient for analyses
This limitation was evident on analyses of different of highly inbred species (Breviario et al. 2007) or
accessions of E. coracana subsp. coracana, where closely related lines, limiting their applicability for
RFLPs revealed very low levels of polymorphism general germplasm characterization.
(Muza et al. 1995; 23 lines identical out of 26) or no The amplified fragment length polymorphism
variation at all (Salimath et al. 1995a; Dida et al. (AFLP) technique, which also produces dominantly
2007). RFLPs are potentially more useful for interspe- expressed markers, has been extensively used in
cific analyses, but, in those cases, DNA sequencing of plants; it is particularly useful in the absence of geno-
variable regions of the plastid or nuclear genome will mic sequence information, when no specific DNA
most likely be preferred, mainly due to the relative markers (like microsatellites) have been developed,
simplicity of DNA sequencing when compared to and for the production of very large numbers of poly-
RFLP analyses. morphic markers, suitable for discrimination and
RAPDs have also been used to study the genetic unambiguous identification in populations and germ-
diversity and species relationships in the genus (Hilu plasm collections, even when genetic variability is low
1995; Salimath et al. 1995a; Babu et al. 2007). (Meudt and Clarke 2007). AFLPs were recently used
RAPD analysis produces dominant markers and is in Eleusine as one of the sources of markers for the
considered inexpensive and fast but has poor repro- construction of the genetic map of E. coracana (Dida
ducibility and is sensitive to experimental conditions; et al. 2007).
for those reasons, RAPDs are generally considered an SSRs (simple sequence repeats) or microsatellites
unreliable fingerprinting technique (e.g., Salimath (see e.g., Goldstein and Schlötterer 1999) are currently
et al. 1995a; Jones et al. 1997; Harris 1999). the most useful DNA markers for studies of genetic
RAPDs are rarely used nowadays, especially consid- diversity and structure of natural and cultivated popu-
ering that far more reliable markers, such as ISSRs or lations, mainly due to their high variability, abundance
microsatellites (see below), have become relatively in the genomes, and the reliability, ease of use, and
inexpensive and easy to use for analyses of diversity affordability of the technique. The only limiting factor
in plant populations. of the use of simple sequence repeats (SSRs) is the
126 S.S. Neves

need for development of specific primers for each new construction of the genetic map of E. coracana
species or group intended to be studied. However, (Dida et al. 2007).
numerous SSRs are already available for E. coracana
(subsp. coracana and africana), 45 SSRs of which
have been successfully used for the construction of
7.6.2 The Genetic Map of E. coracana
the genetic map of the species (Dida et al. 2007) and
analysis of genetic diversity and population structure and Comparative Genomics
(Dida et al. 2008). Additional SSRs markers are avail-
able from the work of Wang et al. (2005), who effec- Comparative genetic mapping has revealed conserved
tively transferred more than 100 SSRs from major gene order (colinearity) among many grass species, but
cereal crops (wheat, rice, maize, and sorghum) to studies at the gene level have demonstrated that micro-
other “minor” grass species, including E. coracana, colinearity of genes is less conserved and that small-
with an average of 58% success rate of cross-genus scale rearrangements and deletions have disturbed the
amplification in the case of finger millet. Wang et al. microcolinearity that was initially predicted, even
(2005) have also analyzed the level of polymorphism between relatively close species, such as sorghum and
detected with the new SSRs and, in the case of finger maize (Keller and Feuillet 2000) – Sorghum and Zea
millet, polymorphism was detected within the species are both members of the Andropogoneae in subfamily
in 21% of the transferred SSRs. These results suggest Panicoideae, see e.g., Bouchenak-Khelladi et al.
that many of the SSRs obtained from major grasses (2008). After it was realized that the level of synteny
with well studied genomes can be successfully ampli- between grass genomes was lower than anticipated, it
fied in other grass genera and that a significant portion became clear that genomic platforms needed to be
of these markers will show adequate variation for established for each crop or species of interest (Powell
characterization and evaluation of diversity of popula- and Langridge 2004).
tions in new grass species (Wang et al. 2005). The high A first step in the development of molecular tools
success rate of cross-genus amplification detected in that could assist the breeding (marker-assisted selec-
the work of Wang et al. (2005) also suggests that tion) of any plant crop is the construction of a genetic
transferring SSRs between closely related grass spe- linkage map. Fortunately, the molecular knowledge of
cies (for instance, from those available for E. coracana the Eleusine genome has progressed considerably in
to other Eleusine species) should be relatively easy recent years, and the first genetic map of E. coracana
with expected high levels of success. Dida et al. (2008) (subsp. coracana and subsp. africana) is already avail-
have already used the SSRs of E. coracana in samples able (Dida et al. 2007). This map was generated using
of E. indica and E. kigeziensis, which are the two a cross between E. coracana subsp. coracana (a culti-
closest species to E. coracana (see Sect. 7.4.1 and var from Nepal: Okhale-1) and its wild progenitor
Fig. 7.2). Direct use of the available E. coracana E. coracana subsp. africana (an accession from
SSRs in more distantly related species, like E. multi- Kenya: MD-20). An intersubspecific cross had to be
flora or E. jaegeri, may not be as straightforward as used for development of this linkage map due to the
with closely related species/taxa of the so-called low levels of variation detected in lines of cultivated
“CAIK” group (“coracana”, “africana”, “indica,” and finger millet (Dida et al. 2007); see also Sect. 7.6.3,
“kigeziensis”). Nevertheless, it is expected that cross- below.
species amplification will be possible for a number of Comparative analysis of colinearity between the
those markers. finger millet and rice genomes have been recently
There has also been great progress in the devel- completed (Srinivasachary et al. 2007). These
opment of expressed sequenced tags (ESTs) in Eleu- authors reported that, other than the expected rear-
sine. Buell (2009) reports that 1,749 ESTs are rangements to explain the difference on chromosome
already available for E. coracana. ESTs represent numbers between finger millet (2n ¼ 36) and rice
transcribed fragments of the genome, which are (2n ¼ 2x ¼ 24), only 10% of the markers were
particularly useful for designing new genetic mar- found in non-syntenic positions and that the finger
kers (e.g., SSRs) and for studies of functional geno- millet and rice genomes have remained relatively
mics (Buell 2009). ESTs were also used in the conserved since the divergence of the two lineages
7 Eleusine 127

from a common ancestor around 60 million years ago Johnson 1992; Werth et al. 1994; Salimath et al.
(Srinivasachary et al. 2007). 1995a; Dida et al. 2007).
In contrast with the low genetic diversity detected
in cultivated finger millet, large morphological varia-
tion has been described in the crop (Hilu and De Wet
7.6.3 Genetic Diversity and Structure
1976b; De Wet et al. 1984; Upadhyaya et al. 2007;
of Populations Dida et al. 2008). High diversity has also been
reported in protein and calcium contents in the screen-
Several molecular and chemical studies that included ing of 36 genotypes of finger millet (Vadivoo et al.
multiple accessions or lines of cultivated finger millet 1998). The morphological and chemical data indicate
(E. coracana subsp. coracana) have consistently that genetic variation exists in the crop and that it is
reported low levels of diversity in the crop. Analyses necessary to considerably increase the number of
of flavonoid content presented no qualitative differences DNA markers and accessions to detect the existing
in the cultivated races of finger millet (Hilu et al. 1978). variation in the genome of the crop. This is exactly
This uniformity in “coracana” accessions was also evi- what Dida et al. (2008) have done with the multiple
dent in restriction fragment analyses of rDNA (Hilu and SSRs that had been previously designed for the con-
Johnson 1992). Werth et al. (1994) reported an isozyme struction of the genetic map of E. coracana (Dida et al.
genotype that was identical in nearly all accessions of 2007). Dida et al. (2008) carried out a comprehensive
finger millet (19 were examined). Similar results were genotypic analysis using 45 SSRs and 96 accessions of
obtained by Muza et al. (1995) in RFLPs analyses (23 Eleusine: 79 accessions of E. coracana subsp. cora-
lines out of 26 were identical). In contrast, much greater cana; 14 of E. coracana subsp. africana; two of
variation was found in both the rDNA pattern (Hilu and E. indica and one accession of E. kigeziensis, the latter
Johnson 1992) and isozyme genotypes (Werth et al. used as an outgroup for analyses. Dida et al. (2008)
1994) of accessions of wild finger millet (E. coracana described three fairly distinct subpopulations in
subsp. africana). Salimath et al. (1995a), using multiple E. coracana that essentially corresponded to: (1)
DNA markers (RFLPs, RAPDs, and ISSRs), also subsp. africana, (2) subsp. coracana from Africa,
detected very low levels of genetic variation in the and (3) subsp. coracana from India. The limits of
crop (17 accessions from India, Nepal, and several Afri- these subgroups are not clear, as it would be expected
can countries), but they were still able to discriminate in populations that naturally interbreed. Dida et al.
each of the lines with the few polymorphisms found. (2008) also observed clear evidence of gene flow
Salimath et al. (1995a) did not include any samples of between wild and cultivated African subpopulations.
wild finger millet, but they found more variation in two Further genetic diversity analyses are underway in
accessions of E. floccifolia (another wild relative) than both cultivated and wild accessions (lines) of finger
in all the 17 lines analyzed for finger millet. The pattern millet, and the new study will include screening of
of low genetic diversity in domesticated finger millet important agronomic traits, such as grain chemical
was further confirmed by Dida et al. (2007). However, composition and resistance to blast disease (Dida and
Dida et al. (2007) highlighted the fact that traditional Devos 2006; Dida et al. 2008).
finger millet breeding have been mostly carried out by There has been no genetic screening of populations
selection of pure lines from local landraces, suggesting in any other Eleusine species. But E. coracana SSRs
that the material that have been examined for genetic can be successfully used in E. indica and E. kigeziensis
diversity may represent only a fraction of the total (Dida et al. 2008).
variation of the landraces. Various authors have also
suggested that the narrow genetic pool of cultivated
finger millet may be a natural consequence of domesti-
cation: the crop resulted from selection of a small subset
7.7 Conservation of Genetic Resources
of the diversity of wild populations and,
subsequently, the gene pool remained restricted by lim- In the past few decades, national breeding programs
ited introgression with wild relatives due to the highly in India and Africa and several international initia-
inbred (self-pollinating) nature of the crop (Hilu and tives have contributed to a considerable expansion
128 S.S. Neves

of plant genetic collections (both ex situ and in 7.8 Problems and Limitations of the
situ). As a result, a very large number of germplasm Eleusine Species
accessions of finger millet are currently preserved
and are being characterized for future use in breed-
ing programs, namely at International Crops 7.8.1 Diseases and Pests
Research Institute for the Semi-Arid Tropics (ICRI-
SAT) (Upadhyaya et al. 2008). The ICRISAT gen- Data on diseases and pests is available mainly for
ebank at Patancheru, India, holds more than 5,000 cultivated E. coracana. However, finger millet seems
accessions of finger millet (mean seed viability of to be little affected by diseases and insects, with the
96.2%), and a safety back-up of 4,580 of these exception of blast disease (National Research Council
accessions are also preserved at the ICRISAT 1996). Blast disease is caused by the fungus Magna-
Regional Genebank at Niamey, Niger (Upadhyaya porthe grisea (T.T. Hebert) M.E. Barr, a species
et al. 2008). ICRISAT has also entered an agree- closely related to M. oryzae B.C. Couch (Couch and
ment with the Royal Norwegian Ministry of Agri- Kohn 2002), the causal agent of rice blast disease,
culture and Food to deposit many thousands of responsible for crop losses in rice of up to 30% a
germplasm seed samples, including those of finger year (Skamnioti and Gurr 2009) – the name Pyricu-
millet, at the Svalbard Global Seed Vault, Norway laria grisea (Cooke) Sacc., commonly used in the
(Upadhyaya et al. 2008). literature, corresponds to an “anamorph” (different
Funding provided by the McKnight Foundation stage in the life cycle) of M. grisea (see e.g., Couch
(Minnesota, USA) (see also Sect. 7.9) has also and Kohn 2002).
helped to increase germplasm collections of finger Naylor et al. (2004) highlighted that the main pro-
millet in Africa, namely of local landraces in blems of the finger millet are its susceptibility to blast
Ethiopia, Uganda, and Kenya. In a recent project disease and drought and that blast disease may reduce
report, it is referred that, during more than three the yields of the crop by 35% or more. For that reason,
decades, National Semi-Arid Resources Research Naylor et al. (2004) recommends considerable invest-
Institute (NaSARRI), in Uganda, maintained the ment in the development of molecular markers and
largest collection of finger millet in Eastern and genetic maps, and the use of marker-assisted selection
Southern Africa, with over 2,500 accessions. How- (MAS) tools, which could facilitate the selection of
ever, this collection has diminished considerably blast-resistant varieties of finger millet. A successful
due to poor conditions of preservation, diseases, breeding program may increase yields in finger millet
pests and civil unrest, with only 965 accessions of by at least 15% (on half of the area of cultivation),
finger millet remaining at the NaSARRI genebank representing more than US$ 38 million of gross annual
(McKnight Foundation Project No. 06-448, Annual benefits (Naylor et al. 2004). Dida and Devos (2006)
Progress Report 2008). A more recent report of the have already reported that, in a small-scale study of
same project indicates that those 965 accessions diversity of E. coracana populations (in progress),
have now been planted and are being evaluated they have already detected accessions of E. coracana
for agronomic traits such as resistance to blast subsp. africana with good levels of resistance to the
and drought and that nearly 120 new finger millet blast fungus. Such blast-resistant accessions could be
accessions have been collected in Uganda, includ- very useful in breeding programs.
ing 19 local landraces (McKnight Grant No. 06- It is likely that other diseases affect Eleusine spe-
448, Annual Progress Report 2009). cies, but information is lacking. A recent report indi-
It is clear that valuable accessions of cultivated cates that the wheat streak mosaic virus (WSMS),
finger millet are being preserved, but attention should which can cause severe crop losses in several cereal
also be given to collections of wild relatives of the crops, has been identified in weed populations of
crop, including wild finger millet (E. coracana subsp. E. tristachya in Australia (Ellis et al. 2004). The
africana) and the close ally E. kigeziensis; accessions WSMS is transmitted by a mite species, but the pres-
of the latter continue to be very rare in germplasm ence of the disease in weeds such as E. tristachya,
collections. which is common in pastures in Australia, is a cause of
7 Eleusine 129

concern as it may facilitate viral infection of neighbor- lizes a critical step in the metabolism of plastids, being
ing crops. thus toxic to plants (and some microorganisms) but not
animals (Baerson et al. 2002, and references therein).
Since the introduction in 1996 of “Round-up Ready”
transgenic crops from the Monsanto company, these
7.8.2 E. indica: An “Intractable” Weed crops have been widely used and grown under single
usage or repeated applications of glyphosate (Sander-
E. indica or goosegrass (see Table 7.1, for other com- mann 2006). Historical use and initial tests suggested
mon names) is considered one of the worst weeds in the that glyphosate was a low risk herbicide for the evolu-
world, with large distribution in the tropics, but now tion of weed resistance (Baerson et al. 2002). However,
affecting temperate countries as well, and reported as a the repeated use of the herbicide led to the appearance
problematic weed for 46 different crop species in more (selection) of new glyphosate-resistant biotypes in two
than 60 countries (Holm et al. 1977); it is also one of weed grasses, Lolium rigidum Gaudin (rigid ryegrass)
the ten most important herbicide-resistant weeds (Basu and E. indica (Lee and Ngim 2000). The goosegrass
et al. 2004; Beckie 2007). One of the reasons of the glyphosate-resistant biotype, initially found in Malaysia,
reproductive success of goosegrass is its capability of was determined to be 8- to 12-fold more resistant to the
producing large quantities of seeds; a single plant has herbicide than susceptible goosegrass, rendering glyph-
been reported to produce up to 140,000 seeds (Lee and osate completely ineffective in the control of the weed
Ngim 2000). As a result, farmers often rely on inten- (Lee and Ngim 2000). The source of resistance to
sive use of herbicides to control this weed (Lee and glyphosate in the Malaysian biotypes was later identi-
Ngim 2000). fied as a single nucleotide substitution in the gene
Goosegrass has evolved or had pre-existing resistance encoding the EPSPS enzyme (target for the herbicide)
against a number of important herbicides, such as dini- that resulted in an amino acid substitution (proline for
troanilines and inhibitors of acetohydroxyacid synthase, either serine or threonine at position 106), with the
acetyl-CoA carboxylase and acetolactate synthase, altered enzyme being no longer affected by the presence
which have been used to control the spreading of this of glyphosate (Baerson et al. 2002; Ng et al. 2003).
and other major weeds among crops (Baerson et al. 2002; Resistance of E. indica to the herbicide glyphosate was
Ng et al. 2004a). The genetics and molecular mechan- determined to be inherited as a single, nuclear and
isms involved in dinitroaniline resistance in E. indica incompletely dominant gene (Ng et al. 2004a). How-
have been extensively studied (e.g., Yamamoto and ever, Owen and Zelaya (2005) and Ng et al. (2004b)
Baird 1999; Zeng and Baird 1999) and are reviewed by have evidence that indicates that there is a second resis-
Anthony and Hussey (1999). tance mechanism non-targeted to EPSPS, meaning that
Goosegrass has also been very problematic as a E. indica has developed alternative resistant mechan-
turfgrass weed but controllable, for many years, by isms to glyphosate. This is a cause of major concern
applications of the herbicides metribuzin plus MSMA given that glyphosate resistance is the dominant trans-
(monosodium methanearsonate) (Brosnan et al. 2008). genic trait being used, corresponding to 114 million ha
However, new metribuzin-resistant goosegrasss bio- of transgenic crops in 23 countries (Green 2009). The
types have now been documented in bermudagrass widespread use and often sole reliance on glyphosate
turf (Cynodon spp.), which were also resistant to has stimulated the evolution by selection of resistant
another herbicide combination: simazine plus MSMA weeds (Green 2009). Long-term sustainability of glyph-
(Brosnan et al. 2008). Nevertheless, the metribuzin- osate is thus at risk and, for that reason, companies are
resistant biotypes were susceptible to herbicides such currently developing crops that combine glyphosate
as glyphosate and foramsulfuron, which can still be resistance with resistance to herbicides with other
used to control goosegrass in bermudagrass turf (Bros- modes of action; the new transgenic crops will give
nan et al. 2008). farmers more options in the fight against weeds (Green
Glyphosate is the active ingredient in the broad spec- 2009).
trum and widely used herbicide known as “Roundup.” Herbicide-resistant weeds will continue to evolve,
Glyphosate acts by inhibition of the enzyme EPSPS (5- and only judicious use of herbicides, including the
enolpyruvylshikimate-3-phosphate synthase) that cata- alternation of herbicides with different modes of
130 S.S. Neves

action, integration of cultural and mechanical weed wild populations of finger millet and other Eleusine
management methods, as well as rotation of transgenic species. Special attention should be given to the col-
crops with conventional cultivars, will need to be lections of E. kigeziensis, which appear to be scarce.
followed if widespread selection of glyphosate- or Variability in this species is likely to be high as shown
other type of herbicide-resistant weeds is to be pre- from sequencing work on just a few accessions.
vented (Lee and Ngim 2000). In addition to the more extensive genetic screening
E. indica will likely continue to be a major weed of populations and lines of wild finger millet
problem for many years. But its remarkable capability (E. coracana subsp. africana), it will be important to
of resistance to adverse environmental conditions may carry out a comprehensive genetic diversity assess-
have some positive applications. The use of E. indica in ment in E. kigeziensis, a wild tetraploid species that
phytoremediation of petroleum-polluted soil is currently is very closely related to E. coracana (Neves et al.
under investigation (Merkl et al. 2005; Wang et al. 2008). 2005; see also Fig. 7.2) and, potentially, a source of
useful traits for the breeding of cultivated finger millet
(Dida and Devos 2006).
Hybridization between the crop and wild finger
7.9 Future Prospects and
millet could also significantly improve the nutritional
Recommendations value of the domesticated plants (Barbeau and Hilu
1993).
There has been recently great progress in the geno- For many decades, the funds directed to research on
mic knowledge of Eleusine, and the construction of “orphan crops,” such as finger millet, had been meager,
a genetic map for E. coracana (Dida et al. 2007) particularly when compared with those invested on
was a crucial step for future breeding. Comparative major cereal crops, such as maize, rice, and wheat
analysis of colinearity between the finger millet and (Naylor et al. 2004). Even recently, Naylor et al.
rice genomes determined that 90% of markers were (2004) considered that aid agencies and science fund-
found in syntenic positions (Srinivasachary et al. ing organizations should increase the investment on
2007), which suggests that other better known crops such as finger millet.
grass genomes can provide invaluable information For nearly 15 years now, the McKnight Foundation
for future breeding in finger millet. In the case of and its Collaborative Crop Research Program (CCRP)
this crop, yields are limited mainly by blast disease (http://mcknight.ccrp.cornell.edu/) has funded research
and drought (Naylor et al. 2004). The mechanisms in finger millet. In 2006, this foundation awarded a new
of blast resistance and the location of resistance grant of $920,000 to the project: “Genetic improve-
genes are better understood in rice (see e.g., Hittal- ment, technology dissemination and seed system
mani et al. 2000; Skamnioti and Gurr 2009; Yang development in African Chloridoid cereals” (Grant
et al. 2009). Therefore, the synteny between finger No. 06-448 Tef/finger millet). The project is directed
millet and rice genomes can be exploited through by Dr. Getachew Belay (Ethiopian Institute of Agricul-
marker-assisted selection to breed blast-resistant tural Research) and involves a network of scientists
varieties of finger millet (Naylor et al. 2004; Daw- from Ethiopia, Uganda, Kenya, and the USA. The
son et al. 2009). Future research in finger millet main goal of this project is to increase the productivity
could also be directed to identify ESTs linked to in tef (Eragrostis tef (Zucc.) Trotter) and finger millet
drought stress or salt tolerance; these markers could crops and eventually benefit farmers in underprivileged
then be used for selection of drought- or salt-toler- regions of Africa.
ant crop varieties that will result in increased pro- In December 2008, the Bill and Melinda Gates
duction in degraded soils and areas normally Foundation awarded $26.7 million over 5 years
affected by droughts (Dawson et al. 2009). (2009–2013) to the McKnight Foundation for research
Many new molecular markers are now available, on crops such as sorghum and finger millet (Star
such as SSRs, which can be easily used for genetic Tribune, Dec. 8, 2008). The ultimate goal of the
screening and complete characterization of existing grant is to help agricultural research and development
germplasm collections. But the effort to expand these in poor regions of the world by increasing yields and
collections should continue, particularly to sample income of small farmers.
7 Eleusine 131

Acknowledgments I extend my thanks to the herbaria and Bouchenak-Khelladi Y, Salamin N, Savolainen V, Forest F,
staff of Virginia Tech (USA), Kew Royal Botanic Gardens van der Bank M, Chase MW, Hodkinson TR (2008) Large
(UK), Sweden Museum of Natural History, Botanical multi-gene phylogenetic trees of the grasses (Poaceae):
Museum Berlin-Dahlem (Germany), Field Museum, Chicago, progress towards complete tribal and generic level sampling.
and INRB-INIA, Estação Agronómica Nacional, Oeiras (Por- Mol Phylogenet Evol 47:488–505
tugal), for granting me access to their collections during Breviario D, Baird WV, Sangoi S, Hilu K, Blumetti P, Gianı̀ S
visits, or for publishing in the web images of herbarium (2007) High polymorphism and resolution in targeted finger-
specimens (especially types), which I was then able to exam- printing with combined b-tubulin introns. Mol Breed
ine. Ginger Swire-Clark and Vance Baird (Clemson Univer- 20:249–259
sity) kindly provided photos of Eleusine for this publication. Brosnan JT, Nishimoto RK, DeFrank J (2008) Metribuzin-
My work is currently supported by a postdoctoral fellowship resistant goosegrass (Eleusine indica) in bermudagrass
(SFRH/BPD/26669/2006) from Fundação para a Ciência e a turf. Weed Technol 22:675–678
Tecnologia, Portugal. Buell CR (2009) Poaceae genomics: going from unattainable to
becoming a model clade for comparative plant genomics.
Plant Physiol 149:111–116
Chen S, Phillips SM (2006) Eleusine. In: Wu ZY, Raven PH,
Hong DY (eds) Flora of China, vol 22, Poaceae. Science
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.
Chapter 8
Eragrostis

Mahmoud Zeid, Vivana Echenique, Marina Dı́az, Silvina Pessino, and Mark E. Sorrells

8.1 Introduction by botanists, ranchers, and even military personnel

from Australia, England, South Africa, and the United
States. After the evaluation of thousands of worldwide
Eragrostis is the largest genus within the subfamily
trials, the “winners” were four African grass species
Chloridoideae of the Poaceae. The first description and
that were characterized by the ease of establishment
naming of the genus was made by von Wolf (1776) on
relative to native grasses, persistence, and high pro-
a specimen of Eragrostis minor. Although it is not
ductivity as forage species. These grasses were weep-
clear why Wolf selected this name, “lovegrass” is the
ing lovegrass [Eragrostis curvula (Schrad.) Nees],
accepted name for the genus (from eros, love, and
Lehmann lovegrass (Eragrostis lehmanniana Nees),
agrostis grass). Today, about 350 species have been
buffelgrass (Cenchrus ciliaris L.), and kleingrass
described in the genus. Most species occupy open habi-
(Panicum coloratum L.). E. curvula and E. lehmanni-
tats with poor soils and many occur in ruderal sites (Van
ana have been heavily utilized mainly in the USA and
den Borre and Watson 1994). Their distribution extends
Argentina for soil conservation and forage production
across wide altitudinal and moisture gradients from
purposes. Another well known species is E. pilosa, a
pluvial to xeric habitats. Most of the species (50%)
common weed in many parts of the world and the
appear to be native of Africa (Cufodontis 1974). Fifty
most closely related to the cultivated species E. tef.
five species are native in Australia (Lazarides 1997), 25
Because this species is considered the progenitor of
in United States and Canada, and 36 in Mexico (Peter-
the cultivated tef, many studies were initiated on this
son and Valdes-Reyna 2005).
weed, and it is now part of the breeding programs
Despite the large number of Eragrostis species,
for tef improvement in Ethiopia. The three species
they are mostly unknown weedy grasses, and only a
E. curvula, E. lehmanniana, and E. pilosa will be the
few became known as forage grasses. The adverse
focus of this chapter.
effects of the frequent droughts that hit Africa,
Australia, and North America in the period between
1880 and 1930 led to a decrease in livestock produc-
tion on grassland. The need for new grass species that
could produce good quality forage under limited pre- 8.2 Basic Botany of the Species
cipitation and heavy grazing was realized. This moti-
vated a worldwide search for the “miracle grass” [see 8.2.1 Taxonomic Position
Cox et al. (1988) for more details]. The search was led
Eragrostis Wolf, consisting of more than 350 species,
is the largest genus in the Eragrostideae, a tribe of 80
genera and about 1,000 species (Peterson et al. 1997)
M.E. Sorrells (*) mainly distributed in tropical and warm-temperate
Department of Plant Breeding and Genetics, Cornell University,
Ithaca, NY 14853, USA regions all around the world, where the centers
e-mail: [email protected] of diversity are assumed (Hartley and Slater 1960).

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 135
DOI 10.1007/978-3-642-14255-0_8, # Springer-Verlag Berlin Heidelberg 2011
136 M. Zeid et al.

The tribe is regarded as an unnatural grouping of E. pilosa is a very widely distributed species with
convenience, and Eragrostis itself is a large polyphy- extensive morphological diversity. It is an annual,
letic assemblage (Voigt et al. 2004). Members of Era- reproduces by seed that is a caryopsis, ellipsoid, later-
grostis are generally characterized by paniculate ally compressed, 0.6–1 mm long (Fig. 8.1). Culms are
inflorescence, multifloreted spikelets, glabrous three- erect or geniculately ascending, 8–70 cm long. The
nerved lemmas and unawned, longitudinally bowed- ligule is a fringe of hairs, and leaf-blades are 2–20 cm
out paleae with ciliolate keels, and leaves with ciliate long, 1–4 mm wide. The inflorescence is an open
ligules (Peterson et al. 1997). Attempts at within genus panicle, elliptic or ovate, 4–25 cm long with spikelets
classification have focused largely on characters such comprising 4–14 fertile florets. Glumes are deciduous
as the manner of spikelet disarticulation or C4 photo- with the lower glume 0.5–0.7  the length of the upper
synthesis type. However, a cladistic analysis of mor- glume. On the other hand, both E. curvula and
phological and anatomical characters has suggested E. lehmanniana are perennials, reproducing by seeds,
that the mode of spikelet disarticulation has little to although in the former, reproduction is mostly through
do with natural relationships in the group (Van den apomixis (facultative). Seed is an ellipsoid caryopsis
Borre and Watson 1994). Furthermore, relationships with the adaxial surface flattened, 1.4 mm long, 0.6 mm
may have been obscured by allopolyploidy (Ingram wide, and translucent reddish brown in E. curvula, dark
and Doyle 2004). The polyphyletic nature of Eragros- brown or black and smaller (0.6–0.8 mm long, 0.4 mm
tis based on the analysis of conserved sequences has wide) in E. lehmanniana. Culms are tufted, 40–100 cm
been a point of debate (Hilu and Alice 2000; Ingram tall, erect or ascending, and in E. lehmanniana, some
and Doyle 2004; Roodt-Wilding and Spies 2006). culms are decumbent or geniculate at lower nodes,
Ingram and Doyle (2004), using the plastid locus often stolon-like. The leaf blades are longer in
rps16 and the nuclear gene waxy, argue that Eragrostis E. curvula (20–30 cm), 1–1.4 mm wide, with ligules
is monophyletic with the inclusion of several segre- densely ciliate, shorter (5–15 cm), 1–3 mm wide, with
gates, including Acamptoclados, Diandrochloa, and truncate ciliate ligules in E. lehmanniana. The inflores-
Neeragrostis. Roodt-Wilding and Spies (2006) on the cence is an open panicle, oblong, 20–40 cm long in
other hand, using data from chloroplast trnL (UAA) 50 E. curvula while oblong to lanceolate and much shorter
exon-trnF (GAA) region and the nuclear ribosomal (7.5–16 cm) in E. lehmanniana. Spikelets are grayish-
internal transcribed spacer (ITS) regions, see the green with multiflorets (6–15), glumes deciduous, and
genus as polyphyletic. Peterson and Sanchez Vega the lower glume shorter than the upper one.
(2007) have noted that the debate is not yet over and E. curvula was initially described by Schrader like
what is lacking is a definitive treatment of the infra- Poa curvula being included in the genus Eragrostis by
generic classification of the entire genus.

8.2.2 Morphology of Eragrostis

There are many detailed reports on the morphology of

Eragrostis upon which plants could be identified from
various parts of the world, from Africa (Cope 1998),
Australia (Lazarides 1997), Bolivia (Renvoize 1998),
Brazil (Boechat and Longhi-Wagner 2000), Ecuador
(Laegaard and Peterson 2001), Malesia (Veldkamp
2002), Mexico (Peterson and Valdes-Reyna 2005),
Peru (Peterson and Sanchez Vega 2007), and the
United States and Canada (Peterson 2003). Here, we
will give a very brief description of the three species
with which this chapter is concerned, based mainly on Fig. 8.1 The small size of Eragrostis pilosa grains in compari-
Clayton et al. (2006). son to a wheat grain
8 Eragrostis 137

Nees. Weeping lovegrass, E. curvula (Schrad.) Nees, (E. chloromelas). He proposed a classification based
is a vigorous grass native of South Africa that is well on seeds, leaf blades, and ear characteristics for mate-
adapted for forage production and soil conservation in rials grown in Argentina. The new classification and
different countries. This species constitutes a poly- the most important Argentinean cultivars are the
morphic group that is morphologically diverse, poorly following: curvula (cvs. Tanganyika, Ermelo, Don
understood, and not well circumscribed. For those Arturo INTA and Morpa), pilosa (cv. Don Juan
reasons, it is frequently referred to as the E. curvula INTA), conferta (cv. Don Walter INTA). and robusta
complex or E. curvula sensu lato (Voigt et al. 2004). (cvs. Don Carlos INTA, Don Pablo INTA and Don
The difficulty in defining the structure of this group is Eduardo INTA).
caused by the lack of taxonomic knowledge. Although The reproductive mode of weeping lovegrass
there are marked morphological differences between (apomixis) seems to have played a fundamental role
varieties, the existence of ecotypes with intermediate in producing a complex superstructure of polyploid
characteristics hinders the classification. In addition, forms, as has happened in other grasses. While sexu-
the apomictic reproduction can limit any new varia- ality would allow the emergence of polyploid types
tion. This has led to considerable confusion among that are progressively more complex, apomixis would
ecotypes and cultivars that are currently available to ensure the survival of these meiotically unstable new
breeders and producers (Poverene 1988; Covas 1991). lines. These genotypes could, in turn, lead to other
The E. curvula complex includes intermediate forms genotypes through the fusion of unreduced gametes
that overlap with E. chloromelas and E. lehmanniana; and facultative apomixis. The E. curvula complex
so some authors do not consider these species to be consists of a set of populations, most of which are
separate from E. curvula sensu lato. Several publica- clonal and a smaller proportion that is sexual. The
tions in the United States have classified E. chlorome- contact between both populations continuously adds
las as Boer grass, but now it is identified as a botanical new genotypes to the complex, which in this way con-
variety E. curvula var. conferta. Lehmann grass, not stitutes a true collective species (Poverene 1988). The
always included in the complex E. curvula, is more intermediate types encountered by hybridization are
robust and resistant to cold and has morphological expressed in the cultivars as a combination of morpho-
characteristics that differentiate it from weeping love- logical and agronomic characteristics of different vari-
grass (Voigt et al. 2004). The morphological diversity eties. There is currently no precise description of each
of this complex leads to the creation of morphological cultivar or a suitable method for characterization
types that are used to describe the germplasm of of the seed. The seed lacks a sufficient number of
the species but with severe limitations. Leigh (1960) distinctive characters to identify cultivars belonging
and Leigh and Davidson (1968) recognized within to the same botanical variety (Poverene 1988). That is
E. curvula five agronomic types: curvula, robusta to say, the color, shape, and size of the seeds of various
blue, robusta green, robusta intermediate, and chloro- cultivars belonging to the same morphological type
melas. Later, another group was recognized, conferta are remarkably constant, but there are significant dif-
(Jacobs 1982). This classification was based primarily ferences between types.
on morphological characteristics of leaves and inflor-
escences, plant size, and growth habit. Such character-
istics are influenced by growth conditions and
fertilization, and although they are generally distin- 8.2.3 Cytology and Karyotype
guishable as mature plants, they are difficult to recog-
nize as plantlets. The high morphologic diversity The small size of the chromosomes, characteristic of
existing in the complex is accompanied by a similar the Eragrostoideae, and the difficulty in obtaining
change in physiological characteristics and agronomic sufficiently clear preparations contribute to the lack
traits such as cold resistance, photoperiod response, of information about the lovegrass karyotype. The
adaptation to various soil types, forage quality, chromosome sizes of E. pilosa, E. aethiopiaca, and
and response to grazing. Covas (1991) did not E. bicolor, as an example, range between 0.8 and
recognize all types defined above and discarded chlor- 2.2 mm (Tavassoli 1986). The basic chromosome num-
omelas, which was seen as a different species ber characteristic of the genus is 10 (x ¼ 10) with
138 M. Zeid et al.

various ploidy levels (2n ¼ 20, 40, 60, 80, 120), [see frequencies of ring-shaped quadrivalents), hypothe-
Voigt et al. (2004) for more details]. E. pilosa is a sized that there was a high homology of the four
tetraploid (2n ¼ 40) similar to its cultivated relative chromosomes of the association so that it would be
E. tef (Jones et al. 1978), while the ploidy level in an autopolyploid. Recently, Cardone et al. (2006) ana-
E. lehmanniana varies, i.e., 2n ¼ 40 and 2n ¼ 60 lyzed the meiosis of a tetraploid plant obtained by
(Halvorsen and Guertin 2003b). Ploidy levels within chromosome doubling of an artificial diploid-derived
the E. curvula complex, on the other hand, vary from form Tanganyika by in vitro culture from inflores-
diploid to octoploid (Spies and Gibbs Russell 1988; cences with colchicine and obtained an average of
Poverene and Voigt 1997). Although a few sexual 12.2 bivalents, 3.1 quadrivalents, 0.8 trivalents, and
diploid (2n ¼ 2x ¼ 20) plants were identified, they 0.8 univalents. These results and those from Poverene
are very infrequent (so far only four have been col- (1988) would support the hypothesis of the autopoly-
lected) and do not occur in all forms of E. curvula ploid origin of cv. Tanganyika. In species of Paspa-
(Poverene and Voigt 1997). Diploids are always sex- lum, a low frequency of tetravalents in tetraploid
ual and self-incompatible (need cross-pollination), cytotypes has been considered as the result of segmen-
while polyploids reproduce by diplosporous apomixis tal allopolyploidy (Quarin et al. 1996). The structural
and are self-compatible. Although many of the South alterations found in Eragrostis are also considered to
African collections are fully apomictic, a few of the be an indication of segmental allopolyploidy (Vorster
last collections were described as facultative and Liebenberg 1977). Pollen viability evaluated in
(Vorster and Liebenberg 1977). The only report of different plant materials of different ploidy levels in
triploidy in the E. curvula complex comes from E. curvula is high, with at least 50% of viable pollen
Leigh (1960) for a plant belonging to the robusta grains (Poverene 1988; Echenique and Polci 1994;
type. Later, Vorster and Liebenberg (1977) reported Polci 2000). This value is typical of a pseudogamous
the existence of one pentaploid plant. While the grass and guarantees a normal seed production. Esti-
heptaploid level (2n ¼ 7x ¼ 70) is more frequent, mates of pollen viability confirmed that none of the
the even number polyploidy types appear to be much accessions was highly sterile, despite the high fre-
more abundant than odd-polyploidy types in the com- quency of meiotic abnormalities observed. Several
plex (Poverene 1988). It is clear that the persistence of authors have tried to relate the number of chromo-
polyploids should be due to the apomictic reproduc- somes and the morphological types or species that
tion, but the meiotic instability that characterized them make up the complex, but the data are mixed. Each of
necessarily represents an adaptive disadvantage (Pover- the species or morphological types includes a number
ene and Curvetto 1991). Several studies have also found of different cytotypes. Intraspecific variation in chro-
aneuploids, and the spread of such cytotypes is also mosome number is very common in grasses (Quarin
possible because of apomixis (De Winter 1955; Leigh and Fernández 1982; Poverene 1988; Keeler 2004).
1960; Jones et al. 1978). E. tef, the only cereal member of the Eragrostideae,
Poverene (1988) and Cardone et al. (2006) reported is cultivated for its grain (Bekele et al. 1995). Tef is an
the presence of metacentric and submetacentric chro- allotetraploid (2n ¼ 4x ¼ 40) whose origin within the
mosomes with an average size of 2–3 mm in E. curvula. genus Eragrostis is unknown. Studies based on mor-
Meiotic chromosomes in anthers pair mainly as biva- phological, cytological, and biochemical characters
lents, but univalents and multivalents are also com- have suggested as many as 14 wild Eragrostis species
mon. Within the multivalents, tetravalents were the as potential progenitors of the crop [see Ingram and
most common configurations. Tanganyika (curvula Doyle (2003) for further details]. E. pilosa, however,
type 2n ¼ 4x ¼ 40) showed a high frequency of mul- appears to be the most likely candidate for the direct
tiple configurations (four tetravalent and one trivalent wild progenitor of E. tef. Morphologically, the two
on average). The irregularities were higher in the species are very similar, although tef plants are gener-
accessions of higher ploidy levels (Streetman 1970; ally larger with fewer tillers and later maturing than
Vorster and Liebenberg 1977). Poverene (1988), E. pilosa. Another important difference shows at
based on observations in Tanganyika (higher fre- maturity, where the lemmas, paleas, and the caryopses
quency of multiple configurations, the presence of of E. tef remain attached to the rachis, making it possi-
chromatin bridges joining these figures, and the ble for farmers to harvest the grains. On the contrary,
8 Eragrostis 139

E. pilosa shatters its grains due to the breaking apart of E. curvula tolerates several soil conditions that are
the spikelets at maturity. From the cytological point of harmful to other species including extreme pH. It
view, both species have a similar karyotype (Tavassoli grows well on acid soils and mine soils at pH 4.0 as
1986). In addition, evidence from the nuclear gene well as calcareous soils with pH near 8.0. It has been
waxy and the plastid locus rps16 (Ingram and Doyle used for the consolidation of erodible soils through the
2003) clearly indicated that E. pilosa is the closest incorporation of organic matter (Taliaferro et al. 1975;
relative of tef and also supports the hypothesis that Busso and Brevedan 1991).
E. pilosa is its progenitor. Gherbin et al. (2007) compared 23 non-native
As to the genome size of Eragrostis Wolf, an grasses along with Tall Fescue (Festuca arundinacea,
estimation by flow cytometry rendered 595.35 Mbp a native grass in the Mediterranean environment) for
for E. curvula (Echenique et al. unpub results) and their adaptability to the coastal plains of southern Italy
714–733 Mbp for the cultivated E. tef (Ayele et al. in terms of their productivity and nutritional quality. In
1996). The DNA content per haploid nucleus was esti- the second year after establishment, warm-season
mated for E. tef to be 0.68 pg (Bennett and Smith 1976). perennial grasses showed higher dry matter (DM) pro-
duction, with E. curvula producing the highest annual
DM yield (21.1 t ha1). In Argentina, E. curvula is the
most widely cultivated perennial grass with an esti-
8.2.4 Agricultural Status mated area of more than 5,000,000 ha (Covas and
Cairnie 1985). Weeping lovegrass has a remarkable
The three introduced species, weeping lovegrass, boer capacity for the production of forage in environments
lovegrass, and Lehmann lovegrass have been heavily unsuitable for most of the other species used for feed-
utilized in the United States’ agricultural system. ing cattle. This could be attributed to its drought resis-
Weeping lovegrass is unique in that it has been an tance, its capacity to produce in very loose, low
important resource for soil conservation and forage fertility soils, and its tolerance to poor management.
in semiarid regions since its introduction. It exhibits In addition, it has outstanding potential longevity that
water-saving strategies, including leaf waxing and allows pastures to be considered permanent (Covas
rolling in response to increasing water demand with- 1991). The La Pampa province native grasses are the
out necessarily exhausting its water supply (Echenique most important forage resource for cattle in Argentina.
et al. 1986a, b; Colom and Vazzana 2001), thus However, in places where agriculture is possible,
showing good performance in relation to forage pro- weeping lovegrass has increased the receptivity of
duction under water stress conditions (Ruiz et al. fields. The weeping lovegrass cycle (spring-summer)
2008). In addition, its leaf growth is relatively insensi- allows the combination of its use with the best native
tive to soil water drying (Puliga et al. 1996). However, species, which have an autumn-winter cycle, like Pip-
different water stress responses have been detected tochaetium napostaense, Poa ligularis, Stipa tenuis,
among other types within the complex including boer Bromus brevis, etc., thus avoiding the loss of these
lovegrass (cv. Consol) and robusta that are more resis- species by overgrazing. The plasticity of weeping
tant than the curvula type (cv. Ermelo) (Colom and lovegrass allows its introduction in different pro-
Vazzana 2003). These authors reported that during duction systems as it adapts to various alternatives,
drought, relative water content (RWC) decreased providing greater efficiency in all cases (Hernández
65% in cv. Ermelo and drought stress caused severe 1991).
decreases in photosynthetic rates, while cv. Consol
showed low variation in these parameters. Echenique
and Curvetto (1986) found similar results in materials
from Argentina. Balsamo et al. (2006) reported a cor- 8.3 Conservation Initiatives
relation between drought tolerance of three Eragrostis
species and leaf tensile properties (behavior during Data from the United States Department of Agricul-
mechanical stress). E. curvula (drought tolerant) had ture Germplasm Resources Information Network
higher tensile strength values than E. tef (moderately (USDA-GRIN), (USDA 2009) provide a list of 180
drought tolerant) and E. capensis (drought intolerant). Eragrostis species. Seeds for a large number of those
140 M. Zeid et al.

species from a worldwide collection are available for a number of diverse accessions from E. pilosa and
distribution, including the cultivar “A67” from which E. curvula (Bai et al. 1999b). Results indicated that
most of the weeping lovegrass varieties available variability among E. tef accessions on the DNA level
today in the United States were derived. There are was much lower (18%) than that estimated from mor-
currently 548 E. curvula, 54 E. lehmanniana, and 10 phological traits (Bai et al. 1999b). Furthermore, a
E. pilosa accessions listed in GRIN. In Europe, the poor correlation between relationships based on
System-wide Information Network for Genetic AFLP data and morphological data was reported by
Resources (SINGER, http://singer.cgiar.org, accessed Ayele and Nguyen (2000) and Ayele et al. (1999).
18 Nov 2009) shows 191 Eragrostis accessions depos- E. pilosa appeared more similar to E. tef than was
ited at the International Center for Tropical Agricul- E. curvula based on marker data, but the morphologi-
ture (CIAT) and the International Livestock Research cal data failed to differentiate the three species, thus
Institute (ILRI). Another major conservation location emphasizing the need to employ molecular markers for
is the Institute of Biodiversity Conservation (IBC) in such studies. Random amplified polymorphic DNA
Ethiopia where most of the cultivated E. tef accessions (RAPD) markers were employed by Bai et al. (2000)
and other wild Eragrostis species from Ethiopia were to estimate relationships among the same accessions
deposited. from Bai et al. (1999b), with additional new accessions
Weeping lovegrass was first introduced into from E. pilosa and E. curvula. Results were similar to
Argentina from the United States. Predominant culti- those in the study using AFLP markers, although
vars were Tanganyika, Ermelo, and Don Pablo INTA genetic similarity between species was much lower.
(Covas 1991). The introduced cultivars established in The first linkage map in the genus Eragrostis was
the north of San Luis and Córdoba, east of La Pampa, reported by (Bai et al. 1999a) using AFLP markers. A
and west of Buenos Aires provinces. Today, an active striking observation from that study, which involved a
Germplasm Bank located in the Anguil experimental set of recombinant inbred lines from a cross between
station (La Pampa), where the cultivars were first two cultivated tef varieties (Kaye Murri  Fesho),
introduced, is established. The National Institute of was the very low level of polymorphism (6.1%) for
Agriculture of Argentina (INTA), part of the Red two morphologically diverse varieties. To overcome
Nacional de Recursos Genéticos (National Network this problem of low polymorphism from intraspecific
for Genetic Resources), is the location where forage crosses, Zhang et al. (2001) used restriction fragment
species are conserved, including Eragrostis species length polymorphism (RFLP) markers and a popula-
(http://servicios.inta.gov.ar/bancos/ampliacion.html). tion of 116 F8 recombinant inbred lines (RILs) from an
interspecific cross between the cultivar Kaye Murri
(E. tef ) and its wild relative E. pilosa accession
(30-5). E. pilosa was chosen because it was possible
8.4 Role in Elucidation of Origin and
to cross with E. tef and because it was characterized by
Evolution of Allied Crop Plants a shorter stature as compared to the cultivated tef. The
source of DNA probes was a tef cDNA library, and
Attempts to quantify genetic variation in cultivated grass anchor probes from other grass species (barley,
E. tef have usually included the wild E. pilosa and rice, oat and wheat). This study realized a substan-
E. curvula as reference species. Results from Costanza tial increase in the level of polymorphism (67%) and
et al. (1979), Kefyalew et al. (2000), and Tefera et al. initiated comparative mapping of Eragrostis with
(1990) demonstrated that, although tef showed high other members of the Poaceae, since 40% of the
levels of phenotypic diversity, even more variation mapped markers were probes from other grass species.
could be observed between species including its clos- Utilizing 94 RILs of the same interspecific cross, Yu
est relative E. pilosa. Molecular marker techniques, et al. (2006b) updated the map by mapping a set of
however, were essential to overcome the problems Eragrostis-specific expressed sequence tag (EST)-
associated with the effect of environment on morpho- derived simple sequence repeats (SSR), commonly
logical traits as well as seed mixture. Amplified frag- known as EST-SSR markers (Yu et al. 2006a) and
ment length polymorphism (AFLP) analysis was heterologous markers based on EST sequences from
used to study tef accessions and elite lines as well as finger millet, rice, and wheat. Markers were grouped
8 Eragrostis 141

into 21 linkage groups with a mean distance of accession (30-5) was published by Chanyalew et al.
12.3 cm between markers (Fig. 8.2). A second linkage (2005). For this linkage map, 120 RILs were screened
map from an interspecific cross between another vari- using AFLP, EST-SSRs from wheat and Eragrostis,
ety (E. tef cv. DZ-01-2785) and the same E. pilosa SSR markers from rice, and a set of intersimple

1 2 3 4 5
RZ15 0.
TCD230 15.
TB 26. ISSR842 0. TCD3 0. SRSC2_02 0. ISSR811 0.
RZ909 33. RM170 14. ISSR842 6.
BCD1087b 32. CDO2 32. RZ460 20.
RZ27 43. CNLT151- 28.
A 53. BCD1087a 39. TCD248 40. inf30
ISSR836a 53.
TCD134 66. 65. TCD95 56. CSU7 62. TCD197 59.
RZ38 79. CNL78 92. ISSR549 68. RZ962b 72. RM14 81.
TCD182 95. CNL5 93. CSU3 94. TCD306 88. TCD24 92.
CSU60 119. ISSR54 97. PRSC1_02 113.
CDO78 109. CDO139 113.
RZ519 134. RM124 119. CNLT14 122.
BCD20 146. CNLT146- 114. RZ444 137. SRSC3_00 134. RZ44 137.
TCD327 159. BCD88 128. TCD230
PALa RZ909 162. CNLT146- 158. 155.
BCD34 177. 149.
RZ87 CNLT13 168.
BCD1087 190. 161. RZ444
RZ962 186. 179.
BCD944 200.
RZ467 209. ISSR811 217.
RZ49 218. RM10 233.
TCD99 229.
CDO116 248. TCD27 257.
TCD45* 264.
TCD99 271.
CNLT11 296.
KSUM15 298.
RM15 305.

6 7 8 9 10
CNLT49 PAL 0.
0. ISSR811 0.
CNLT49 ISSR81 22. ISSR842 0. TCD5 0.
6. ISSR840 0.
ISSR548b TCD227b 31. KSUM2 18. CNLT7 25.
32. CNL78 23.
RM170 TCD31 33. CNLT41 49.
43. ISSR841 30. RM110a* 40.
RM17 TCD11 36. CNLT41 54.
73. RM124 38. RM110 53.
TCD30 TCD227 42. ISSR842 65.
85. CNLT14 61. CNLT6 68.
ISSR549 ISSR548 60. TCD327 72.
111. RZ698 88. CDO3 64. CNLT11 91. TCD327 88.
CNLT8 114. KSUM195 101.
ISSR842 TCD32 89.
117. inf14 106.
ISSR841 ISSR81 107.
127. TCD230 130.
TCD21 135.
11 12 13 14 15 16
DupW4 0.0 CNLT61 0.0 RM124c 0.0 CDO1387 0.0 CNLT137-T13 0.0 RZ395 0.0
RZ329 15.9 RM134 11.4
ISSR842e 28.0 RZ962a 23.9 RZ467a 20.5 RZ141 27.0 CNLT154-Sa14* 23.1
RZ69* 32.6 RZ123 RM185* 29.6 ISSR836b* 34.6
37.8
RZ166 54.3 RZ413 46.6 RZ251* 50.3 RZ204 49.3 ISSR841a* 46.7
TCD415 63.2 TCD205 60.5
CNLT65 76.6
TCD397a 87.8 TCD5 85.4

17 18 19 20 21
CNLT149* 0.0 TCD503 0.0 lfm256 0.0
ISSR840b** 0.0 RZ588** 0.0
RZ214b* 9.4 RZ698b** 8.1 TCD424 6.5
DupW124 RZ519b 13.8 RM142 17.7
CNL100 16.3 22.8
RZ214a 32.6

Fig. 8.2 Genetic linkage map derived from 94 recombinant distorted segregation (P < 0.05) and single asterisk and double
inbred lines from a cross between E. tef cv. Kaye Murri and E. asterisks indicate the preferential transmission from the alleles
pilosa (30-5). Loci names with an asterisk indicate significant of E. tef, or E. pilosa, respectively (Yu et al. 2006b)
142 M. Zeid et al.

sequence repeats (ISSR) markers. The linkage map hybrids that were higher in their in vitro dry matter
covered 78% of the genome, and the average distance digestibility (IVDMD) than common weeping love-
between markers amounted to 12.7 cm. To update the grass hybrids. Boer lovegrass genotype 40-9-69FQ
existing maps, heterologous markers from pearl millet, (40-9) was inferior as a female parent because its
finger millet, and rice are being screened to insure that hybrids tended to be lower in vigor than those from
more anchor markers are included on the maps other boer lovegrass genotypes. Several hybrids from
for comparative analysis purposes (M. Zeid and the cross 40-9 x Morpa were significantly higher in
M. E. Sorrells, unpub data). Most recently, genomic IVDMD than Morpa or Ermelo, the most digestible
SSR markers have been developed to improve genome cultivars (Voigt 1984).
coverage and to identify quantitative trait loci (QTL) Breeders were successful in improving boer love-
for various traits, mainly yield and lodging resistance grass through direct selection. “A-84” boer lovegrass
in the cultivated tef species (M. Zeid and M. E. Sorrells, was first introduced to the United States in 1932 and
unpub data). The recent results from Yu et al. (2007) was officially released in 1950 (Hanson 1965). Selec-
on QTL analysis in tef have indicated that alleles from tion for seedling drought tolerance in A-84 yielded
the wild relative, E. pilosa (30-5), had an increasing the variety “Catalina” boer lovegrass. Catalina also
effect for 29% of the QTL detected. produced more and better quality forage than Leh-
mann lovegrass and was suggested as a replacement
for A-84 boer lovegrass and to partially replace A-68
Lehmann lovegrass (Voigt et al. 1970).
8.5 Crop Improvement Through In order to identify the best morphological traits to
Traditional and Advanced Tools be considered in the evaluation of E. curvula hybrids,
Di Renzo et al. (2000) estimated the degree of influ-
ence of permanent effects on the phenotypic variation.
8.5.1 Forage Crop Improvement Crown diameter, leaf length, dry matter, and panicle
number were considered in this experiment. Repeat-
8.5.1.1 Traditional Tools ability estimates for the vegetative characteristics
indicated that environmental effects were small. For
The E. curvula complex contains a vast diversity of vegetative traits, two harvests provided 98% of the
genes preserved and maintained by apomictic reproduc- accuracy of the total obtained with four cuts, and for
tion. Variation in the establishment capacity, drought panicle number, the same percentage was obtained for
resistance response, forage quality, forage yield, and three harvests. This stability of performance is a desir-
iron efficiency, among others, has been detected. This able characteristic for grass cultivars. These authors
variation can be manipulated through apomictic breed- also studied the trait association patterns and found
ing, i.e., by hybridizing rare sexual plants with apomic- that leaf length was closely associated with dry matter,
tic selections. In this way, the variation can be released with a high repeatability, allowing the use of leaf
and genotypes with new combinations of traits can be length as an indirect criterion for determining the
produced. Those hybrids, if sexual, can become the aerial biomass yield. This study was continued by Di
basis of further hybridization (Voigt 1991). Renzo et al. (2003) using 18 hybrids that were eval-
Apomictic lovegrass hybrids were developed from uated in three different environments in the semiarid
crosses of sexual boer  apomictic weeping lovegrass region of Argentina (Villa Mercedes, Rı́o Cuarto and
(Voigt 1984) with significant differences in forage Bahı́a Blanca) in order to identify environments under
vigor and quality found among the hybrids. Selection which the efficiency of indirect selection could be
for palatability gave rise to a variety of weeping love- maximized. In general, the heritability obtained was
grass, “Morpa” (more palatable), which was released higher in Villa Mercedes and Bahı́a Blanca than in Rı́o
by the USDA (Voigt et al. 1970). Morpa was selected Cuarto. The estimated decrease in efficiency of selec-
for its higher palatability relative to other weeping tion for all the traits measured in Rı́o Cuarto was
lovegrass strains, such as Common and Ermelo in higher than in the other locations, suggesting that
summer grazing. However, it is very similar to Ermelo this place was not a good choice for carrying out
in appearance. The apomictic cultivar Morpa produced indirect selection. Conversely, genetic progress
8 Eragrostis 143

would be faster if selection was carried out in Villa green) and differs from cv. Don Eduardo in leaf
Mercedes, because the evaluations provided higher color, chromosome number, and cold resistance. The
and more accurate estimates of the heritabilities than tetraploid line obtained using colchicine was grouped
in the other environments. within the weeping type and linked to Tanganyika
using molecular markers (Zappacosta 2009). The
case of the diploid somaclone is peculiar, because it
8.5.1.2 Advanced Tools presents morphological and molecular differences
with the other cultivars of the weeping group. Using
Tissue culture protocols for E. curvula have been RAPDs, AFLPs, and SSRs, it was classified within the
established in order to generate somaclonal variation curvula type, but at the morphological level, it exhibits
and to assist genetic transformation (Echenique et al. similarities with the robusta type.
1996, 2001). Plant regeneration from four genotypes Transformation of an apomictic clone is an attrac-
of E. curvula, cvs. Kromdraii (6x), Tanganyika (4x), tive strategy, as the transgene is immediately fixed in a
Morpa (4x), and Don Pablo (7x), was reported by three highly adapted genetic background capable of large-
developmental pathways: embryogenesis, organogen- scale clonal propagation. Some efforts have been
esis, and direct regeneration (Echenique et al. 1996). directed towards developing a genetic transformation
Immature inflorescences were the best explant source, protocol for the genus. Ncanana et al. (2005) reported
and fertile plants were obtained from all the genotypes the development of plant regeneration and transforma-
evaluated. The less variable apomictic cultivars, such tion protocols for E. curvula cv. Ermelo. Callus was
as Morpa, Tanganyika, and Don Pablo (Poverene generated from leaf and seed tissues and transformed
1988), showed a lower regeneration response than by particle bombardment with the yeast Saccharomy-
Kromdraii, which has the highest level of sexual ces cerevisiae Hsp12 gene under the maize ubiquitin
reproduction and is a hexaploid (2n ¼ 60). The high promoter. Although successful transformation and
ploidy level could have a “buffering” effect that would transcription of the Hsp12 gene occurred, no Hsp12
minimize genetic abnormalities; the extra chromo- protein was found present in tissue extracts of the
some copies having the capacity to compensate for transformed grass. Using embryogenic callus obtained
it. Kromdraii and Don Pablo are high polyploids, and from immature inflorescences, Dı́az (2006) evaluated
the latter is a heptaploid with a high basal level of the transient expression of uidA gene introduced by
chromosome aberrations (Poverene 1988), which may particle bombardment. Three different promoters were
have increased during in vitro culture (Echenique et al. analyzed, and the promoter of the ubiquitin gene from
1996). Somaclones were characterized at the morpho- maize produced the best response. Other physical con-
logical and molecular level using randomly amplified ditions of the bombardment device and mannitol/sor-
polymorphic DNA (RAPD) markers (Polci 2000). bitol concentration in the pretreatment medium were
After evaluation, three materials were selected from studied.
the apomictic tetraploid cv. Tanganyika (2n ¼ 40) Forage species showing summer growth, in general,
(Polci 2000; Cardone et al. 2006): (1) a sexual diploid tend to have higher fiber and lower protein contents
plant (2n ¼ 20) registered as Victoria (RC9192, when compared to those of temperate climates. This
2006–2026, Argentina), (2) a tetraploid line (2n ¼ 40) implies low digestibility and low animal productivity.
with a high level of sexual reproduction obtained E. curvula does not escape from this model, so that one
after the treatment with colchicine of the diploid of the main aspects to be considered in breeding pro-
line (seeds) and registered as Bahiense (RC9193, grams is forage quality. Implementation of molecular
2006–2026, Argentina), and (3) two polyploid highly technologies to downregulate lignin using both trans-
apomictic plants, one of which gave rise to the apo- genic (gene technology) and non-transgenic (molecu-
mictic cv. Don Luis (RC9191, 2006–2026, Argentina). lar marker technology) strategies are attractive for
This cultivar differs from other materials of E. curvula Eragrostis. Gene technology approaches for targeted
since the chromosomal number is 2n ¼ 64. It is an downregulation of key enzymes [e.g., caffeic acid O-
apomictic cultivar classified as robusta blue type. methyltransferase (OMT), cinnamyl alcohol dehydro-
It differs from cvs. Ermelo, Morpa, and Don Juan genase (CAD), cinnamoyl CoA reductase (CCR)]
because of leaf width and color (blue instead of involved in lignin biosynthesis in transgenic species
144 M. Zeid et al.

leading to altered lignification and significantly selection from landraces, since crossing techniques
enhanced dry matter digestibility have been developed were not available. Because of the small size of the
(Chen et al. 2003; He et al. 2003; Marita et al. 2003). flowers of tef and the lack of knowledge concerning
On the other hand, using cDNA libraries of E. curvula the time of flower opening and pollen shedding, early
(Cervigni et al. 2008a) to identify genes involved in the attempts to produce hybrids were not successful
lignin biosynthetic pathway, five transcripts from puta- (Ketema 1991). In 1974, the first successful crossing
tive enzymes were identified (M. Dı́az et al. unpub in tef was achieved (Berhe 1975) based on the obser-
results): phenylalanine ammonialyase (PAL), caffeic vation that flowers open earlier in the morning than
acid/5-hydroxyferulic acid O-methyltransferase previously thought. Breeders have utilized this break-
(COMT), caffeoyl-CoA O-methyltransferase through in the hybridizing techniques to increase the
(CCoAOMT), hydroxycinnamoyl-CoA reductase yield of the crop by providing farmers with improved
(CCR), and cinnamyl alcohol dehydrogenase (CAD). varieties. The urgent need for interspecific crosses
The characterization of the full-length CCoAOMT between E. tef and its closest wild relatives was
cDNA, a key enzyme of the pathway, and the later recognized in the 1990s, when yield improvement
genomic sequence isolation in cv. Tanganyika, allowed using intraspecific crosses began to plateau. Crosses
the identification of four alleles with a tissue-specific between E. cilianensis, E. minor, E. pilosa, and E. tef
expression. Three alleles, A1, A2, and A4, were found were attempted (Tavassoli 1986; Gugsa and Mengiste
to be expressed in leaves at an equal level. A1 does not 1999), but success rates in producing hybrid seeds
represent a good candidate to be downregulated, consid- were limited. E. pilosa was seen as the best candidate
ering that it is highly expressed in roots. The manipula- for such crosses because it has the same ploidy level
tion of A2 may cause an important alteration in and chromosome number as E. tef and is characterized
inflorescences essential for seed production. The third by much shorter culms, making it a good candidate for
allele, A4, constitutes a potential candidate to modified transferring the short stature trait to reduce lodging in
lignin (quality or quantity) in weeping lovegrass plants. cultivated tef. The first crosses were developed at the
Thus, molecular genetic markers may be used for the Debre Zeit Agricultural Research Center in Ethiopia,
detection and permutation of agronomically important and recombinant inbred lines from these cross are
genetic variation in sexual populations, followed by constantly being evaluated for useful agronomic traits
linked marker based transfer of the apomixis character (Tefera et al. 2003), and a candidate variety is cur-
to produce new clonal varieties. rently under evaluation (G. Belay, pers. comm.).
Those populations also provided the raw material for
molecular marker studies aimed at linkage mapping
and quantitative trait loci (QTL) analyses.
8.5.2 Improvement in Cultivated E. tef
Utilizing Its Wild Relatives 8.5.2.2 Advanced Tools

8.5.2.1 Traditional Tools The development of tissue culture techniques is an

essential tool for embryo rescue for crosses involving
Phylogenetic analysis of sequence data from the E. tef and its close relatives. Breeding new improved
nuclear gene waxy and the plastid locus rps16 (Ingram E. tef varieties is becoming very difficult due to the
and Doyle 2003) strongly support the widely held plateau reached in yield using the available breeding
hypothesis of a close relationship between tef and the material and the lack of variation in the available gene
wild allotetraploid E. pilosa. Because of this close pool for lodging resistance. While crossing with
relationship between the cultivated tef and E. pilosa, E. pilosa has been successful, other interspecific
the latter has received more attention, especially from crosses are desired for future improvement in E. tef.
plant breeders, as compared to other wild species in The crossing between E. tef and E. curvula as an
the genus. example would be a major breakthrough, for E. curvula
Efforts to improve the yield of the cultivated E. tef possesses desirable traits such as stem strength, rust
have started in the 1950s, basically through pure line resistance, and large florets to hold large seeds
8 Eragrostis 145

(Berhe 2001). Few in vitro plant regeneration techni- isms were detected in 18 contigs generated from three
ques from roots, leaf bases, and seeds of E. tef were to four libraries.
reported (Bekele et al. 1995; Mekbib et al. 1997; In addition to EST sequences, about 1,000 other
Kebebew et al. 1998). However, the most recent work nucleotide sequences are also available for 49 Era-
by Gugsa et al. (2006) reports successful results from grostis species, the majority of which are from studies
unpollinated flower explants in both E. tef and on E. tef and E. pilosa aiming to develop new molecu-
E. mexicana. Furthermore, flow-cytometry of the tissue lar markers, namely, SSRs (M. Zeid and M. E. Sorrells,
culture-derived tef plants have indicated the regenera- unpub results) and conserved intron scanning primers
tion of haploids, triploids, tetraploids, and octaploids (CISP) markers (Feltus et al. 2006) designed to utilize
with varying morphological traits as compared to the the information from sorghum and rice to develop
allotetraploid control plants. This is another promising genome analysis tools for orphan species. Fifty-seven
tool for creating useful variation in E. tef. Further protein sequences are available for 19 species mainly
studies on E. tef and its wild relatives are needed for from phylogeny and evolutionary studies (c.f. Ingram
improvement in breeding strategies and techniques in and Doyle 2003; Bell and Columbus 2008; Christin
the cultivated tef. et al. 2009).
In E. curvula, a comparative expression analysis
was performed based on EST sequencing and differ-
ential display using the libraries from the euploid
8.6 Genomics Resources Developed series (Cervigni et al. 2008b). From a total of 8,884
unigenes sequenced, 112 (1.26%) showed significant
Genomic resources may be used both for gene isola- differential expression in individuals with different
tion for transgenesis and for the development of gene- ploidy levels and/or variable reproductive mode. Inde-
associated molecular markers, such as SSRs and pendent comparisons between plants with different
single nucleotide polymorphisms (SNPs). Currently, reproductive modes (same ploidy) or different ploidy
only about 16,000 expressed sequence tags (ESTs) are levels (same reproductive mode) allowed the identifi-
available in Genbank (http://www.ncbi.nlm.nih.gov/, cation of genes modulated in response to diplosporous
accessed 18 Nov 2009) for the genus Eragrostis. development or polyploidization, respectively. EST
Seventy-seven percent of the ESTs were identified in sequencing allowed the identification of approxi-
E. curvula (Cervigni et al. 2008a) and the rest in mately 100 genes with differential expression asso-
E. pilosa and E. tef (Yu et al. 2006a, b). The E. curvula ciated with apomixis or ploidy level (Cervigni et al.
EST collection was generated from four cDNA 2008a). Interestingly, a significant number of genes
libraries, three of which were obtained from panicles were similarly expressed in the 2x sexual and the 4x
of near-isogenic lines with different ploidy levels and apomictic lines, but differentially in the 4x sexual line.
reproductive modes [described by Cardone et al. Most of those were silenced in the 2x sexual and the 4x
(2006)] and one obtained from 12 day-old plant apomictic lines. Based on these results, it was pro-
leaves. A total of 12,295 high-quality ESTs were clus- posed that apomixis in E. curvula could be the con-
tered and assembled, rendering 8,864 unigenes, sequence of failure in activation of a group of genes
including 1,490 contigs and 7,394 singletons, with a that must be expressed when ploidy level increases
genome coverage of 22%. A total of 7,029 (79.11%) (Cervigni et al. 2008b).
unigenes were functionally categorized by BLASTX
analysis against sequences deposited in public data-
bases, but only 37.80% could be classified according
to Gene Ontology. Sequence comparison against the
8.7 Scope for Domestication and
cereals genes indexes (GI) revealed 50% significant
hits. A total of 254 EST-SSRs were detected from
Commercialization
219 singletons and 35 from contigs. Di- and tri-
nucleotide motifs were similarly represented with A number of Eragrostis species is widely known for
percentages of 39 and 40%, respectively. In addition, being grazed by domestic animals in many parts of the
190 SNPs and insertion/deletion (INDEL) polymorph- world, especially in Africa. Burkill (1994) listed 27
146 M. Zeid et al.

Eragrostis species as fodder plants that are in some Dekking et al. 2005), making it a safe replacement for
cases eaten by animals for lack of better feed sources wheat flour for celiac disease patients suffering from
in the west tropical Africa. Furthermore, not all the intolerance to gluten. Tef flour also adds value to the
species are safe for grazing. In southern and eastern gluten-free products due to its high vitamin and fiber
Africa, some species (for example, E. plana) contain content (Hopman et al. 2008). To our knowledge, no
hydrocyanic acid, which is fatal for some farm animals similar studies were done on the wild species of Era-
(Watt and Breyer-Brandwijk 1962). grostis.
Weeping lovegrass could be an attractive source of
biomass for biofuels due to the amount of biomass it
generates. McMurphy et al. (1975) found that weeping
lovegrass compared favorably to switchgrass across 8.8 Some Dark Sides
all N rates. The maximum yield of each species was
9.4, 9.3, 6.4, and 8.4 t ha1 for switchgrass, weeping
8.8.1 The Invasive Nature of Some
lovegrass, Indian grass, and big bluestem, respec-
tively. Weeping lovegrass and switchgrass were the
Eragrostis Species
two most productive species at 82 kg of N and 18 kg of
P. Furthermore, Klemp (1981) reported weeping love- Seeds of E. lehmanniana originating from South
grass three years’ average yield as high as 13.3 t ha1. Africa were first introduced to Arizona, USA, in 1932
Other uses of Eragrostis species besides being a to reseed/revegetate livestock grazing areas, construc-
feed or biofuel include the use of culms for thatching tion sites, golf courses, and wildfire areas (Anable et al.
and matting (E. cilianensis, E. tremula), broom 1992). At the same time, E. curvula from Tanganyika
making (E. gangetica, E. ciliaris), and weaving into was introduced to the same region (Cox et al. 1988)
cordage (E. ciliaris, E. pilosa, E. tremula) (Abbiw and was seeded extensively for erosion control along
1990; Burkill 1994). Medicinal uses are generally banks and slopes of highways and mine spoils, on
limited to a few species. E. ciliaris is useful as a revegetated sites, range, and pasture sites (Alderson
stomachic, E. japonica and E. tremula as lactation and Sharp 1995). The newly created Eragrostis pas-
stimulants, and E. scotelliana and roots of E. tremula tures produced forage more consistently than native
are sources of aromatic substances (Burkill 1994). grass pastures, and ranchers often devoted part of
E. lehmanniana was used by the Europeans in today’s their ranches to these African grasses to ensure forage
South Africa as a remedy for colic, diarrhea, and supply in years with low precipitation (Cable 1971).
typhoid fever. Also, the Zulu used a decoction of Recent reports showed that the introduced Eragrostis
E. plana roots for treating profuse menstruation, impo- species have spread beyond initially targeted areas and
tency, and barrenness, while the Southern Sotho used invaded grassland sites that had no human or animal
the plant as a tonic and as one of the charm ingredients disturbance (Williams and Baruch 2000). Adverse
in a preparation used for treating fractures (Watt and effects of E. lehmanniana and E. curvula on the areas
Breyer-Brandwijk 1962). In Ghana, the ash of the it invaded were slowly being recognized. Anable et al.
burnt E. ciliaris plants is mixed with shea-butter and (1992) reported that E. lehmanniana has changed/
smeared over skin burns (Dokosi 1998). Plants of transformed the structure and function of large areas
E. pilosa are an effective cure for contusions, while of the semiarid grassland in Arizona since it was intro-
infused leaves of E. japonica are used for headache duced. It reduced faunal diversity (Bock et al. 1986)
(Duke and Ayensu 1985). and affected faunal pedoturbation (Hupy et al. 2004).
Grains of many Eragrostis species including The species invaded arid grasslands, forming dense
E. cilianensis, E. ciliaris, E. minor, E. pilosa, monocultures that become fire-prone in dry summer
E. tenella, E. tremula, and E. turgida are usually conditions (Cox 1999). These monocultures changed
consumed by humans in times of severe famine in fire frequency and size, competed with and replaced
many parts of Africa (Watt and Breyer-Brandwijk native species, and altered geomorphological pro-
1962; Burkill 1994; Dokosi 1998). Recent studies cesses and hydrology (Marshall et al. 2000). Similarly,
have shown that flour from grains of the cultivated many reports, ca. 25, have considered E. curvula as
E. tef lacked gluten and gluten-like proteins (Spaenij- a weed in countries like Australia, New Zealand,
8 Eragrostis 147

Mexico, Spain, and England (Global Compendium of effort will kill the remaining seeds (Invasive Alien
Weeds 2007). In Japan, E. curvula has been introduced Plants Species of Virginia 1999). Selective herbicides
to prevent erosion. This species has established large or herbicides accompanied by timed and controlled
populations, forming highly productive seed sources, burns following the reseeding of native species are
especially in mountainous regions (Washitani 2004). other options for the control of Eragrostis species
Once riparian habitats were invaded by E. curvula, and promoting native vegetation (Campbell et al.
the original conditions and vegetation of gravelly 1985; Halvorsen and Guertin 2003a, b).
floodplains, i.e., sparse vegetation cover consisting
principally of riparian endemics growing in gravelly
sandy conditions, were rapidly lost (Muranaka and 8.9 Recommendations for Future
Washitani 2003). Nakayama et al. (2007) predicted
the expansion range of the species due to seed dispersal
Actions
from source populations in order to plan effective
measures for maintaining both ecosystem integrity Eragrostis species have made many important contri-
and the biodiversity. butions to human kind, and the wide range of genetic
While E. curvula is recognized in Argentina as a variation in the genus is an indication that there is
promising forage crop, it has a mixed reputation in much potential for novel uses in the future. Clearly,
Australia as a noxious weed, pasture grass, and soil the extensive variation has played an important role in
binder (Lazarides 1997). In the United States, both the ability of these species to tolerate environmental
E. curvula and E. lehmanniana are no longer on the extremes and to adapt to forage cropping and perhaps
conservation plant release list, and the latter was lab- eventually biofuel production. Eragrostis has also
eled “potentially invasive” (http://www.plant-materials. been used for many medicinal, nutritional, and func-
nrcs.usda.gov/releases/discontinued.html, accessed 18 tional purposes. The exploitation of this variation can
Nov 2009). Eragrostis cumingii Steud (Cuming’s love- be facilitated through apomictic breeding to release
grass) and E. cilianensis (Stinking lovegrass), also new genetic variation, thus producing genotypes with
introduced to the United States from Africa, were new combinations of traits. In addition, transfo-
declared “invasive” in natural areas (http://www. rmation of an apomictic clone is a useful approach
invasive.org/weedus/grass.html, accessed 18 Nov because the transgene is immediately fixed in an elite
2009). The government in New Zealand declared E. genotype that is capable of large-scale clonal propa-
curvula among the “unwanted organisms” along with gation. Finally, it may eventually be possible to trans-
140 other plant species (http://www.biosecurity.govt. fer some of the genetic variation underlying useful
nz/pests-diseases/plants/accord/amending-list.htm, traits in these species, such as apomixis, to other
accessed 18 Nov 2009). cultivated species. However, to realize some of
Control of E. curvula plants through prescribed fire these potential benefits, much additional research is
was not successful. E. curvula benefits from fire, gen- needed to develop the necessary information base
erally increasing (Wright et al. 1978) or remaining and molecular tools.
stable in numbers after fire (Walsh 1994), deep roots
and dense crown of growth protects the plant from fire
damage (Phillips et al. 1991). E. Lehmanniana also has
high potential to reestablish after fire (Sumrall et al.
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.
Chapter 9
Festuca

Toshihiko Yamada

9.1 Basic Botany of the Species panicle; spikelets two – many-flowered, disarticulat-
ing above the glumes; glumes shorter than lemma;
upper glume usually three-nerved; lemmas membra-
The genus Festuca L. is one of the largest in Gramineae
nous to thinly coriaceous, usually glabrous, five-
and is, along with Poa L., the largest genus of the tribe
nerved, acute, awnless or awn-tipped; palea almost
Poeae. Festuca L. and its closely allied genus Lolium L.
equal to lemma, scabrous, or ciliate; lodicules two;
have long fascinated agronomists, evolutionists, and
stamens three; ovary sometimes hairy on the top
plant breeders, and these genera are among the most
(Jauhar 1993).
widely studied of the non-cereal grasses. Recent evi-
Classification of the almost cosmopolitan genus
dence from molecular phylogenetic studies shows that
Festuca has varied through the last two centuries.
the genus lacks monophyly. As a result, several species,
Hackel (1882) subdivided Festuca into six sections:
including the forage grasses, tall fescue (Fig. 9.1),
Ovinae, Bovinae, Sub-bulbosae, Variae, Scariosae,
Festuca arundinacea Schreb. [¼syn. Lolium arundina-
and Montanae based on vegetative and floral charac-
ceum (Schreb.) Darbysh.] and meadow fescue, Festuca
ters. Within sect. Ovinae and Variae, subgroups were
pratensis Huds. [¼syn Lolium pratense (Huds.) Dar-
characterized by intravaginal versus extravaginal
bysh.], formerly belonging to the genus Festuca, have
innovation. Saint-Yves (1922) largely adopted this
been recently placed into the genus Lolium (Darbyshire
system for his worldwide revisions of the genus. How-
1993). However, these species are described here as
ever, Cvelev (1971, 1976) and Alexeev (1977, 1981,
belonging to the genus Festuca.
1982, 1984, 1988) proposed a substantially new sys-
Festuca L. is a large, diverse genus whose members
tem of classification with 11 subgenera, many of
are widely adapted to a variety of ecogeographical
which were further divided into sections. The molecu-
regions. It comprises some 450 species (Clayton and
lar systematic studies using ITS/trnL-F provided new
Renvoize 1986) that range from diploid (2n ¼ 2x ¼
insights in the phylogeny of subtribe Loliinae of tribe
14) to dodecaploid (2n ¼ 12x ¼ 84) in chromosome
Poeae (Catalán et al. 2004; Torrecilla et al. 2004). The
number (Šmarda and Stančı́k 2006).
subtribe can be roughly classified into two major
A genus of Festuca L. is characterized with mor-
lineages: (1) the poorly supported “broad-leaved” fes-
phological features as follows: perennial, tufted to
cues, falling into several lineages of unclear relations
something short-rhizomatous; leaf-blades typically
and (2) the highly supported “fine-leaved” fescues
parallel-veined, mostly rolled, something flat; leaf
containing the majority of Festuca species. Two sub-
sheaths open; ligules short, membranous; auricles
groups of Festuca recognized by Hackel (1882) fall
present or absent; inflorescence a compressed or lax
into different lineages, raising the need to recognize
two new sections. M€uller and Catalán (2006) described
two sections of Lojaconoa and Dimorphae based on
T. Yamada
Field Science Center for Northern Biosphere, Hokkaido
results of recent phylogenetic studies.
University, Kita 11, Nishi 10, Kita-ku, Sapporo 060-0811, Japan Because of the diversification of the ploidy levels in
e-mail: [email protected] various taxonomically intricate groups, ploidy level is

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 153
DOI 10.1007/978-3-642-14255-0_9, # Springer-Verlag Berlin Heidelberg 2011
154 T. Yamada

Fig. 9.1 Tall fescue (Festuca arundinacea)

an important species character in recent Festuca undeveloped species, giant fescue, F. gigantea (L.)
systematic treatment. Recent studies have shown that Vill. [¼syn. Lolium giganteum (L.) Darbysh.; 2n ¼ 6
flow cytometry is useful for the determination of DNA x ¼ 42], has acquired importance because it pro-
ploidy level in fresh and herbarium materials of duces a mass of very large soft leaves of high nutri-
Festuca (Šmarda and Kočı́ 2005; Šmarda et al. 2005, tive value (Thomas and Humphreys 1991). Further,
2008b; Šmarda 2006, 2008; Šmarda and Stančı́k grass breeders have interest on some species, such as
2006). The measurement of DNA content (2C-value), diploid F. altissima All., F. donax Lowe and F.
monoploid genome size (Cx-value), average chromo- drymeja Mert. and W.D.J. Koch, tetraploid F. mairei
some size (C/n-value), and cytosine + guanine (GC) St. Yves, F. pratensis var. apennina (De Not.) Hack.
content using flow cytometry in 101 Festuca taxa and and F. arundinacea var. glaucescens Boiss., and dec-
14 of their close relatives facilitated understanding the aploid F. arundinacea var. letourneuxiana St. Yves
long-term processes of genome evolution, testing evo- (Thomas and Humphreys 1991).
lutionary hypotheses, and their usefulness for large- Tall fescue is the most important forage species
scale genomic projects, suggesting that there is an worldwide of the Festuca genus. It is indigenous
evolutionary advantage for small genomes in Festuca to Europe and also occurs naturally on the Baltic
(Šmarda et al. 2008a). coasts throughout the Caucasus in western Siberia
The genus Festuca contains two agriculturally and extending into China (Jauhar 1993). Tall fescue
important forage crops, hexaploid tall fescue (F. introductions have been made into North and South
arundinacea) (2n ¼ 6x ¼ 42) and diploid meadow America, Australia, New Zealand, Japan, and South
fescue (F. pratensis) (2n ¼ 2x ¼ 14). Other and East Asia (Barnes 1990). It was introduced into
fescues of some importance are red fescue, F. rubra USA from Europe in the early 1800s. It is widely
L. (2n ¼ 6x ¼ 42 or 8x ¼ 56), and sheep fescue, grown in southern Europe and is the predominant
F. ovina L. (2n ¼ 4x ¼ 28). The agriculturally cool-season perennial grass species in the USA,
9 Festuca 155

especially in the transition between cool-temperature diversity for future breeding objectives as well as for
and subtropical zones (Sleper and West 1996). long-term stability of ecosystems gave rise to two
Meadow fescue is one of the most widely used conservation strategies for genetic resources (CBD
forage grasses in the Scandinavia countries, due to its 2005). Ex situ conservation includes the storage in
generally high level of winter stress tolerance. It gene banks of germplasm collections intended to best
is distributed natively in Europe and western Asia represent genetic diversity. In breeding station, supe-
(Fjellheim et al. 2007). Although not widely used as rior genotypes are maintained with vegetative propa-
a forage grass species outside Scandinavia, there has gation. Ex situ conservation can be effective for
been an increasing interest in meadow fescue during protecting species or small number of threatened vari-
the last decade in Europe, North America, and Japan. eties or ecotypes. However, it is not very effective for
In North America, the acreage of meadow fescue has maintaining the vast genetic diversity characteristic of
not been significant since the early twentieth century the species and are very expensive, especially when
(Casler and van Santen 2000), because extensive test- live plants are maintained (Peeters 2004). In contrast,
ing of forage yield capacity around the turn of the in situ conservation includes the maintenance of plants
nineteenth century (Buckner et al. 1979) indicated at their specific habitats (e.g., permanent grassland),
that tall fescue was superior to meadow fescue in allowing continuing evolutionary adaptation (Frankel
vigor and resistance to crown rust (caused by Puccinia et al. 1995; Maxted et al. 1997). Ecological factors and
coronata). However, recent research has shown that agricultural practices have created a vast biodiversity
meadow fescue may be more useful than tall fescue that can only be conserved by protecting the habitats
in intensive grazing management systems in North and using management methods close to those that
America (Casler et al. 1998). In eastern Hokkaido, created that diversity (Peeters 2004).
Japan, which has severe winter climate, meadow Natural or semi-natural permanent grasslands har-
fescue is promising as a grazing species. Therefore, bor forage grass ecotype populations, which are highly
breeding program of meadow fescue was started and adapted to their individual macro- and micro-habitats,
winter-hardy and high-yielding variety “Makibasa- showing high variation for traits of adaptive signifi-
kae” has been released recently (Tase et al. unpub). cance such as ear emergence, growth habit, or resis-
Fine-leaved Festuca spp. such as F. rubra (red tance to various diseases (Wilkins 1991; Fjellheim
fescue) and F. ovina (sheep fescue) are used for et al. 2007). This wealth of variation has been used
turf species (Ruemmele et al. 2003). Fine-leaved as the main source of genetic variation for forage
Festuca spp. are generally characterized by their crop breeding since its beginning in Europe in the
fine to very narrow leaves, usually less than 1 mm early twentieth century (Humphreys 2005). Several
(Beard 1973). They have been grown on greens of studies have investigated ecotype populations of dif-
golf course in Scotland for centuries (Beard 1998). ferent forage grass species by means of molecular
Use on fairway increases with the desire for more markers (Fjellheim and Rognli 2005; Peter-Schmid
environmentally sustainable golf courses (Christians et al. 2008a), mostly showing a high variability within
2000). the populations, as it is expected for obligate outbree-
ders with a gametophytic self-incompatibility system.
Recent studies indicated that ecotype populations
may still allow to improve new varieties with favor-
9.2 Conservation Initiatives able alleles, which do not exist in present cultivars
(Fjellheim et al. 2007; Peter-Schmid et al. 2008a).
Temperate grasses including Festuca species are the Peter-Schmid et al. (2008b) demonstrated that ecotype
main component of permanent grasslands. Genetic populations and cultivars were clearly separated, and
diversity present in permanent grasslands is threatened there was a significant correlation between diversity of
by intensification of forage production. This is basi- morphological and geographic location of samplings
cally a consequence of an increasing use of external sites. Maintenance of permanent grassland in contrast-
inputs of fertilizers and large-scale resowing with ing environments appears to be a promising strategy
improved cultivars of only few species (Brown 1992; for preserving valuable genetic variation of forage
Tscharntke et al. 2005). The importance of genetic grasses (Peter-Schmid et al. 2008b).
156 T. Yamada

9.3 Role in Elucidation of Origin and 9.4 Role in Development of Cytogenetic

Evolution of Allied Crop Plants Stocks and Their Utility

Hexaploid tall fescue is endemic to much of Europe Festuca and Lolium species hybridize naturally and as
and North Africa (Borrill et al. 1971). Tall fescue is hybrids regularly exchange genes at high frequency.
an outcrossing allohexaploid with 2n ¼ 6x ¼ 42 Some Festulolium cultivars have been developed as
chromosomes. Its genome size is approximately novel temperate forage grasses in both Europe and
6  103 Mbp (Seal 1983). The genomic formula of USA (Table 9.2) (Yamada et al. 2005).
tall fescue is considered to be PPG1G1G2G2 Intergeneric hybrids between closely related Fes-
(Chandrasekharan and Thomas 1971a, b). The P tuca and Lolium species are being used to broaden the
genome is from the diploid (2n ¼ 2x ¼ 14) meadow gene pool and provide plant breeders with options to
fescue, and the G1 and G2 genomes are from the combine high quality traits with broad adaptation to a
tetraploid (2n ¼ 4x ¼ 28) F. arundinacea var. glau- range of environmental constraints (Humphreys et al.
cescens (Sleper 1985). Genome constitution of Fes- 2003). Therefore, intergeneric hybridization has been
tuca species has been considered by Bowman and carried out to develop a novel cultivar (Thomas
Thomas (1976), Thomas et al. (1997), and Harper and Humphreys 1991). Many cytological researches
et al. (2004) (Table 9.1). Further genome analyses have been published. Jauhar (1993) reviewed the stud-
are needed to prove it. ies on cytogenetics of the Festuca–Lolium complex.

Table 9.1 Chromosome number and DNA contents of some Festuca and Lolium species
Species Common name Chromosome number/genome 1C DNA content (pg)
Festuca arundinacea Tall fescue 2n ¼ 6x ¼ 42; AABBCC 6.05
F. arundinacea var. glaucescens 2n ¼ 4x ¼ 28; BBCC 4.28
Festuca gigantea Giant fescue 2n ¼ 8x ¼ 56; AAXXYYZZ 7.23
Festuca mairei Atlas fescue 2n ¼ 4x ¼ 28; BBDD 3.95
Festuca pratensis Meadow fescue 2n ¼ 2x ¼ 14; AA 2.20
Festuca scariosa 2n ¼ 2x ¼ 14; BB 2.68
Lolium perenne Perennial ryegrass 2n ¼ 2x ¼ 14 2.08
Lolium multiflorum Italian ryegrass 2n ¼ 2x ¼ 14 4.10
Modified with Kopecký et al. (2008)

Table 9.2 Festulolium cultivars developed in Europe and USA

Hybrid combination Cultivar name Type Country Year
L. multiflorum  F. pratensis Elmet Amphidiploid United Kingdom 1973
Perun Amphidiploid Czech Republic 1991
Rakopan Amphidiploid Poland 2001
F. pratensis  L. multiflorum Paulita Amphidiploid Germany 1986
Paulena Amphidiploid Germany 1995
Punia Amphidiploid Lithuania 1997
Felopa Amphidiploid Poland 1998
Sulino Amphidiploid Poland 1998
Agula Amphidiploid Poland 2002
L. perenne  F. pratensis Prior Amphidiploid United Kingdom 1973
Spring Green Amphidiploid USA 2001
L. multiflorum  F. arundinacea Kenhy Introgression USA 1977
Johnstone Introgression USA 1983
Felina Introgression Czech Republic 1988
Hykor Introgression Czech Republic 1991
Korina Introgression Czech Republic 1997
Bečva Introgression Czech Republic 1989
From Yamada et al. (2005)
9 Festuca 157

Recently, Yamada et al. (2005) and Kopecký et al. leads to restoration of fertility. Partially male and
(2008) reviewed the cytogenetics of grass species, female fertile F1 hybrids can be also obtained by
which are used in breeding of improved cultivars, crossing autotetraploid forms of Lolium sp. and
including interspecific and intergeneric hybrids, and F. pratensis. The first amphidiploid Festulolium cul-
the main attention was given to Lolium/Festuca tivars, “Elmet” (L. multiflorum  F. pratensis ) and
hybrids. “Prior” (L. perenne  F. pratensis) were bred at the
The Lolium/Festuca introgression system is based Welsh Plant Breeding Station, UK (now Aberystwyth
on a series of seven monosomic substitution lines. In University), in the early 1970s (Thomas and
each of these seven lines, one of the chromosomes of Humphreys 1991).
Lolium perenne has been replaced by its homeologous A major problem for amphidiploid breeding is the
equivalent from F. pratensis (King et al. 1998, 2002a, high level of homoeologous pairing between the dif-
b). An introgression map of L. perenne/F. pratensis ferent genomes that leads to genetic instability and
chromosome 3, homeologous to rice chromosome 1, loss of hybridity in later generations. To overcome
has been generated. The introgression map is com- these problems and reduce transfer of deleterious
posed of 16 individuals, each of which carries differ- Festuca traits, selective introgression of genes for
ent-size Festuca chromosome segments. Alignment of desirable traits from Festuca into Lolium has become
overlapping Festuca chromosome segments effec- a favored methodology. This process involves the
tively divides L. perenne/F. pratensis chromosome 3 transfer of small segments of alien Festuca chromatin
up into 18 physically demarcated bins. Screening the into the recipient Lolium genome (Thomas et al. 1988;
individuals that make up the introgression map of Humphreys 1989; Humphreys and Pašakinskiene
Lolium/Festuca chromosome 3 for the presence or 1996) and has been successfully employed to produce
the absence of Festuca polymorphisms allows genetic novel Festulolium lines (Humphreys and Thomas
markers including restriction fragment length poly- 1993; Humphreys et al. 2005). Intergeneric hybrids
morphism (RFLP), amplified fragment length poly- between L. multiflorum (2x) and F. arundinacea (6x)
morphism (AFLP), simple sequence repeat (SSR), were used in the first putatively successful introgres-
and single nucleotide polymorphism (SNP) to be sion breeding program conducted at the University of
assigned to one of the 18 introgression bins (King Kentucky, USA, in which two cultivars, “Kenhy”
et al. 2002a, b). King et al. (2007) described the (Buckner et al. 1977) and “Johnstone” (Buckner
exploitation of the published rice genome sequence et al. 1983), were developed, although in both cases,
to bin map sequences from functionally annotated the inclusion of Lolium genes was never confirmed.
gene models on approximately every fifth to tenth bac- “Kenhy” showed improved palatability and a lower
terial artificial chromosome/plasmid artificial chromo- fiber content than tall fescue “Kentucky 31” (Buckner
some (BAC/PAC) clone from rice chromosome 1 to et al. 1979). Introgression procedures for the transfer
the Lolium/Festuca chromosome 3 introgression map of genes for drought resistance from F arundinacea
to elucidate the syntenic relationship between rice and var. glucescens (2n ¼ 4x ¼ 28) into L. multiflorum
the large-genome monocots, to determine the physical (2n ¼ 2x ¼ 14) using DNA markers derived from
location of rice chromosome 1 genes in large-genome F. glucescens were described by Humphreys et al.
monocots and to determine the relationship between (2005).
gene distribution and recombination in large-genome Androgenesis was found to be an effective proce-
crop species. dure for selection of Lolium–Festuca genotypes
comprising gene combinations rarely or never recov-
ered by conventional backcross breeding programs
9.5 Role in Classical and Molecular (Leśniewska et al. 2001; Humphreys et al. 2003).
Androgenesis from Festuca–Lolium complex had
Genetic Study been studied using different parental hybrids such as
F. pratensis  L. multiflorum (Leśniewska et al. 2001;
Intergeneric hybrids between diploid L. multiflorum, Rapacz et al. 2004), L. multiflorum  F. arundinacea
L. perenne, and F. pratensis have very low fertility. (Humphreys et al. 1997, 1998b; Pašakinskienė et al.
Doubling of the chromosome number of the F1 hybrids 1997; Zwierzykowski et al. 1999; Zare et al. 1999),
158 T. Yamada

L. perenne  F. pratensis (Guo et al. 2005), and codominant multiallelic loci. Momotaz et al. (2004)
L. perenne  F. mairei St. Yves (Atlas fescue) (Cao have analyzed the genetic polymorphism of multiple
and Sleper 2001). Superior plants with high freezing genotypes derived from taxa of the Lolium/Festuca
tolerance have been found in the backcrossed progeny complex using these distinct sets of SSR markers and
(Yamada et al. 2007). Kosmala et al. (2006) used applied these data to investigate introgression and
florescence in situ hybridization (FISH) and genomic genetic relatedness in Festulolium accessions. RAPD
in situ hybridization (GISH) to identify a segment of and SSR markers were used to detect the parental
F. pratensis chromosome 2, which was introgressed genome composition of F1 hybrids and backcross,
into L. multiflorum and was located terminally on a generated from crosses between F. mairei St. Yves
non-NOR arm in two of the three most freezing- and L. perenne (Wang et al. 2009). Recently, Tamura
tolerant plants. Chromosome 4 of F. pratensis with et al. (2009) have developed the intron-flanking EST
the translocated terminal segments of L. multiflorum markers for the genetic analysis and molecular breed-
on both arms was detected in the third plant. F. mairei ing of Lolium, Festuca, and their intergeneric hybrid,
has exceptional drought tolerance but poor turf Festulolium. In this marker system, primer sets are
and forage quality. Some progeny in a high drought- designed from ESTs of the related species showing
tolerant group derived from the intergenic hybridiza- high similarity to unique rice genes. Thus, compara-
tion between F. mairei and L. perenne rated better than tive genomic analyses allow the genomic loci of mar-
the F. mairei parent (Wang and Bughrara 2008). These kers in the related species to be estimated from the
valuable breeding materials will be used in Festulo- corresponding rice loci. High species specificities of
lium introgression breeding programs to accelerate the these markers based on the intron polymorphisms are
breeding process and provide novel robust new forage advantageous for the genetic analysis and molecular
grasses for cultivation in marginal areas. breeding of interspecific or intergeneric hybrid.
The GISH technique provides the means to identify
segments of alien chromosomes introduced into the
recipient species and has proved to be a powerful tool
9.6 Role in Crop Improvement Through
for determining their chromosome location (Thomas
et al. 1994; Humphreys et al. 1995; Humphreys and Traditional and Advanced Tools
Pašakinskiene 1996). However, there are potential
difficulties in the identification of very small intro- Tall fescue is a cross-fertilizing species and is largely
gressed chromosome segments using the GISH tech- self-sterile (Sleper 1985). As a result, individuals are
nique (Humphreys et al. 1998a). highly heterozygous and populations are heteroge-
Molecular DNA markers based on Southern hybri- neous (Sleper and West 1996). Source of breeding
dization such as RFLP as well as PCR-based markers materials include plant introductions, commercial
such as random amplified polymorphic DNA (RAPD), cultivars, planned crosses among selected clones
AFLP, and SSR have been developed for grass spe- including progeny from interspecific and intergeneric
cies. Yamada and Kishida (2003) applied rice cDNA- crosses, and populations resulting recurrent selection
RFLP probes from the activity of the Rice Genome (Sleper and West 1996). Breeding objects of tall fes-
Program (RGP) of Japan to forage grasses in order cue includes forage yield, seed yield, feeding quality
to investigate genetic variation within and between for improved animal performance, and resistance to
varieties of grasses and to identify variety-specific pest and diseases (Sleper and West 1996).
RFLP markers for use in breeding programs exploiting Genetic improvement of forage grasses such as
intergeneric hybridization of Lolium and Festuca. fesues by conventional plant breeding is slow since
RFLP analysis is a highly labor-intensive methodol- many species are predominantly, if not completely,
ogy compared to the PCR-based methods. Recently, allogamous wind-pollinated grasses. Gene technology
SSR markers have been developed in tall fescue (Saha and the production of transgenic plants offer the
et al. 2004). SSR markers provide the current marker opportunity to generate unique genetic variation.
system of choice due to their abundance, ubiquitous Application of transgenesis to forage plant improve-
distribution in plant genomes, high level of reproduc- ment has been focused on the development of trans-
ibility, ease of PCR-based analysis, and detection of formation events with unique genetic variation and in
9 Festuca 159

studies on the molecular dissection of plant biosyn- sequenced tag (EST)-SSR primer pairs were designed
thetic pathways and developmental processes of high from tall fescue cDNA library sequences and tested for
relevance for forage production (Spangenberg et al. cross-species amplification in seven different grass and
2001). In recent years, biolistic transformation and cereal species (Saha et al. 2004). A subset of these
Agrobacterium-mediated transformation have become markers was used for more intensive phylogenetic anal-
the main methods for producing transgenic grasses. ysis of cool-season grasses (Mian et al. 2005). EST-
Biolistic methodology, based on particle bombard- SSR and AFLP primer pairs were used to construct
ment, employs high-velocity gold or tungsten particles a tall fescue map based on PCR-generated marker gen-
to deliver DNA into living cells for stable transforma- otyping of the F1 (HD28-56  R43-64) two-way
tion. Because biolistic methodology is a physical pro- pseudo-testcross population (Saha et al. 2005). Integra-
cess that involves only one biological system, it is a tion of the parental genetic maps produced a consensus
fairly reproducible method that can be easily adapted structure of 17 linkage groups (LGs) and a cumulative
from one laboratory to another laboratory. Transgenic map length of 1,841 cm. Homoeologous relationships
forage plants have been obtained by particle bombard- between six of the seven expected groups were
ment of embryogenic cell in tall fescue and red fescue observed. Further map enhancement would be expected
(Spangenberg et al. 1995; Cho et al. 2000; Wang et al. to identify the remaining five homologous LGs to the
2001, 2003; Chen et al. 2003, 2004) Agrobacterium- anticipated total of 22.
mediated transformation has the advantage of allow- Tall fescue RFLP markers have also been used to
ing for low copy number integration of the transgenes efficiently detect RFLPs in the diploid F. pratensis.
into the plant genome. In recent years, significant Comparative genetic mapping between the two species
progress has been made in developing transformation of fescue (Chen et al. 1998) generated a map containing
protocols using Agrobacterium tumefaciens as a vec- 66 markers on seven LGs with a total map length of
tor. Transgenics have been obtained by Agrobacter- 280 cm. A high level of conservation for the orders
ium-mediated transformation in tall fescue (Dong and of most of the genetic markers was observed, with 23 of
Qu 2005; Wang and Ge 2005) and Festololium (Guo 33 common markers located in corresponding LGs.
et al. 2009). Gao et al. (2008) described that Agrobac- A comprehensive genetic map has been constructed
terium-mediated transformation appears to be the pre- for meadow fescue using a two-way pseudo-testcross
ferred method for producing transgenic tall fescue F1 population from the pair-cross of a single geno-
plants by comparative analysis using both methods. type from a Norwegian population that has been
For downregulation of endogenous genes, the tar- selected for frost tolerance (HF2/7) and a genotype
get gene is normally isolated from a species and trans- from a Yugoslavian variety (B14/16). The combined
ferred back to the same species or closely related data for homologous and heterologous RFLP, AFLP,
species. To improve forage digestibility by downregu- SSR, and isoenzyme markers from the two parental
lation of lignin biosynthesis, genes encoding two maps defined 466 loci with a total map length of
key enzymes involved in lignin biosynthesis, cinna- 658 cm (Alm et al. 2003). Conserved synteny was
myl alcohol dehydrogenase (CAD) and caffeic acid analyzed through the use of heterologous RFLP
O-methyltransferase (COMT), were cloned from tall anchor probes derived from perennial and Italian rye-
fescue (Chen et al. 2002). Transgenic tall fescue plants grass, wheat, barley, oat, rice, maize, and sorghum,
were produced using antisense and sense CAD and demonstrating a high degree of conserved synteny
COMT gene constructs under the control of maize and colinearity with both perennial ryegrass and the
ubiquitin promoter. Severely reduced mRNA levels Triticeae consensus map. The F1 (B14/16  HF2/7)
and significantly decreased enzymatic activities were mapping population has been used for quantitative
found in some transgenic lines. These transgenic tall trait loci (QTL) analysis of vernalization requirement,
fescue plants had reduced lignin content, altered lignin heading time and number of panicles (Ergon et al.
composition, and increased in vitro dry matter digest- 2006), following phenotypic evaluation (Fang et al.
ibility (Chen et al. 2003, 2004). 2004). Molecular genetic markers have also been used
Molecular marker system is powerful tool of to monitor the introgression of a crown rust resistance
plant breeding. Some works have been done in tall determinant from F. pratensis into L. multiflorum
fescue and meadow fescue. A total of 157 expressed (Armstead et al. 2006).
160 T. Yamada

Somatic hybridization is a technique based upon meadow fescue, as well as Lolium species including
protoplast fusion, which enables hybridization between perennial ryegrass and Italian ryegrass. Introgression
sexually incompatible species or genera. This technique program could be valuable to transfer useful genes into
may also provide an alternative approach in order to cultivated species for forage and turf.
create novel genetic combination between Festuca and
Lolium species. A specific objective is the creation of
novel cytoplasmic combinations, e.g., a mixture of
9.9 Some Dark Sides and Their
both parental cytoplasms. Symmetric somatic hybrids
(Takamizo et al. 1991) and asymmetric somatic hybrids Addressing
(Spangenberg et al. 1994) have been produced between
F. arundinacea and L. multiflorum, but no further Some of species of Festuca, especially F. arundina-
research regarding their progeny with respect to agro- cea, have the potential to become weeds. For forage
nomic traits were reported. use, we should consider this invasiveness, especially
when we sow seeds of productive and vigorous culti-
vars near conserved semi-natural or natural pasture.
9.7 Genomics Resources Developed

A total of 41,516 ESTs were generated from nine 9.10 Recommendations for Future
cDNA libraries of tall fescue representing tissues Actions
from different plant organs, developmental stages,
and abiotic stress conditions. The Festuca Gene Molecular breeding is important and will be used
Index (FaGI) has been established (Mian et al. 2008). extensively in future forage and turf improvement.
FaGI provides a useful resource for genomics studies Transformation techniques using Agrobacterium or
of tall fescue and other closely related forage and turf biolistics-based method and many available molecular
grass species. Comparative genomic analyses between markers such as SSR markers and some functionally-
tall fescue and other grass species, including rye- associated genetic markers are now developed in
grasses (Lolium spp.), meadow fescue, and tetraploid Festuca species. We still have remaining challenges
fescue (F. arundinacea var. glaucescens) will benefit for the successful implementation of molecular breed-
from this database. These ESTs are an excellent ing in practical varietal development. Molecular
resource for the development of SSR and SNP-PCR- breeding needs to develop from a platform of good
based molecular markers. Information on tall fescue conventional breeding and include supporting agro-
ESTs is available in the site: http://www.plantgdb.org/ nomic research and partnering with commercial indus-
download/download.php?dir¼/Sequence/ESTcontig/ try where appropriate. With the availability of more
Festuca_arundinacea/previous_version/151a. sequencing information, such as on ESTs, gene isola-
A total of 11,346 transcript-derived fragments (TDFs) tion has become much easier than ever before. We
were detected in F. mairei (Wang and Bughrara 2007). should focus on functional characterization of genes
Analysis of the 163 differentially expressed fragments and their regulatory elements.
(DEFs) provides a first glimpse into the transcripts of F. With a widening range of traits, techniques for more
mairei during drought stress treatment. accurate, rapid and non-invasive phenotyping and geno-
typing become increasingly important. The large
amounts of data involved require good bioinfomatics
9.8 Scope for Domestication support. Data of various kinds must be integrated from
an increasingly wide range of sources such as genetic
and Commercialization
resources and mapping information for plant populations
through to the transcriptome and metabolome of individ-
The main potential economic use for wild species of ual tissues. The merging of data from diverse sources and
Festuca considered to date is as a genetic resource for multivariate data mining across datasets can reveal novel
improvement of cultivated species such as tall fescue, information concerning the biology of complexity.
9 Festuca 161

Sustainable system for production and maintenance Buckner RC, Boling JA, Burrus PB II, Bush LP, Hemken RA
of forage and turf is the most important issue. Some of (1983) Registration of Johnstone tall fescue. Crop Sci
23:399–400
the Festuca species have abiotic tolerance such as Cao M, Sleper DA (2001) Use of genome-specific repetitive
against freezing, heat, and drought. We should evalu- DNA sequences to monitor chromatin introgression from
ate many accessions of Festuca species for the materi- Festuca mairei into Lolium perenne. Theor Appl Genet
als of breeding now. 103:248–253
Casler MD, van Santen E (2000) Patterns of variation in a col-
lection of meadow fescue accessions. Crop Sci 40:248–255
Casler MD, Undersander DJ, Fredericks C, Combs DK, Reed JD
(1998) An on-farm test of perennial forage grass varieties
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Chapter 10
Lolium

Hongwei Cai, Alan Stewart, Maiko Inoue, Nana Yuyama, and Mariko Hirata

10.1 Introduction terranean climates of the world. Largely, these pastures

are allowed to regenerate naturally, but at times they
are also sown. At least four cultivars are recognized in
The genus Lolium contains two widely cultivated tem-
Australia, some are simply regional ecotypes while the
perate species, L. multiflorum (Italian or annual rye-
cultivar Guard has been bred for resistance to Anguina
grass) and L. perenne (perennial ryegrass), and it is
sp. nematode, a pivotal organism along with a bacterium,
against this background that their wild relatives will be
Corynebacterium sp., causing the annual ryegrass tox-
covered in this paper.
icity syndrome in Australia (Stynes and Bird 1993). In
The majority of L. multiflorum sown are annual or
addition, another resistant cultivar, SafeGuard, has
Westerwold ryegrass forms, but very large numbers of
been developed from hybridization between Guard
biannual or Italian ryegrass forms, having potentially
and L. multiflorum.
1,000 or more cultivars, are also used. L. perenne or
L. rigidum has also become notorious in Australia
perennial ryegrass is the most widely sown perennial
for the development of a wide range of herbicide
grass of the temperate world, both as forage and a turf
tolerances, often as few as five generations (Broster
with potentially over 2,000 cultivars. In addition, there
and Pratley 2006). However, herbicide resistance is
have been many hybrids made between L. multiflorum
also developing in naturally reseeding pastures of
and L. perenne and these are termed hybrid ryegrass of
L. multiflorum in other regions of the world. There
which there are potentially over 250 cultivars. Many
are now hundreds of cases of herbicide resistances
artificial tetraploids have also been developed in these
not only in L. rigidum (56) but also many in L. multi-
two species and they are used commercially as forage
florum (34), L. perenne (5), and L. persicum (2) (http://
along with diploids.
www.weedscience.org/In.asp), and many of these
A number of the annual Lolium species are recog-
involve multiple resistance.
nized as serious weeds in cultivated wheat and barley
The Lolium genus has clearly derived from the
fields. These include three inbreeders, L. temulentum,
closely related section Schedonorus of the genus
L. persicum, and L. remotum, as well as an outbreeder,
Festuca, the broad leaf fescues. Indeed, some taxo-
L. rigidum.
nomic revisions have included the broad leaf fescues
Apart from its weed status, L. rigidum is also cul-
in Lolium (Darbyshire 1993), while others have split
tivated as it is naturalized in annual pastures in Medi-
the broad leaf fescues into a separate genus, Schedo-
norus (Soreng and Terrell 1997). Here, we will not
enter into this debate but restrict ourselves to the
H. Cai (*)
Forage Crop Research Institute, Japan Grassland Agriculture traditional Lolium species. It is clear though that the
and Forage Seed Association, 388-5, Higashiakata, broad leaf fescues can be considered as a secondary
Nasushiobara 329-2742 Tochigi, Japan gene pool for Lolium breeding. Indeed there have been
Department of Plant Genetics and Breeding, College of
many Festulolium cultivars developed and are also
Agronomy and Biotechnology, China Agricultural University,
2, Yuanmingyuan West Road, 100193 Beijing, China being used for introgression of genes back into Lolium
e-mail: [email protected], [email protected] (Humphreys et al. 2003).

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 165
DOI 10.1007/978-3-642-14255-0_10, # Springer-Verlag Berlin Heidelberg 2011
166 H. Cai et al.

Like many grass fungal endophytes, the genus Epi- ryegrass, now distributed widely in naturally
chloe¨ or their Neotyphodium anamorphs have coevolved regenerating annual pastures in the Mediterranean
with Lolium and some are now used commercially to climatic zones, including Australia where it is a
provide protection against insects, particularly in New valuable forage
Zealand (Easton 2006). 4. L. canariense Steud.; an annual restricted to the
Canary Islands where it occurs at low altitudes in
poor conditions
10.2 Basic Botany of the Species 5. L. edwardii Scholz, Stierstorfer et Gaisberg; an
annual with a limited distribution at higher alti-
The genus Lolium consists of nine diploid species with tudes on the Canary Islands
a chromosome number of 2n ¼ 2x ¼ 14 (Terrell 6. L. temulentum L.; darnel ryegrass, widely distri-
1968; Clayton and Renvoize 1986; Jauhar 1993), buted as an annual weed of cereal crops (Fig. 10.2)
and these are commonly referred to as ryegrasses. 7. L. persicum Boiss. et Hohen.; widely distributed as
L. edwardii has only recently been named, although an annual weed of cereal crops predominantly in
Terrell (1968) recognized it as a distinct high altitude Western Asia
form of L. canariense (Scholz et al. 2000). 8. L. remotum Schrank [L. tinicolum A. Br.; L. lini-
cola Sond. Ex Koch]; an annual species known
1. L. perenne L.; the widely used perennial ryegrass
largely as a weed in flax
of commercial importance for forage and turf
9. L. subulatum Vis; mainly grows under poor and
(Fig. 10.1)
maritime conditions of Cyprus, Israel, Lebanon,
2. L. multiflorum Lam. (L. italicum A. Braun, nom.
Syria, and former Yugoslavia, and it is not consid-
inval.); The widely used Italian or annual or Wester-
ered a crop weed.
wold ryegrass of commercial importance
3. L. rigidum Gaud. [L. stricture C. Presl; L. loliaceum The genus is distributed in temperate regions of
(Bory et Chaub.) Hand.-Mazz.]; the rigid or annual Central Asia, Europe, and North Africa and has

Fig. 10.1 The plant of Lolium perenne

10 Lolium 167

molecular marker studies using random amplified

polymorphic DNA (RAPD), restriction fragment
length polymorphism (RFLP) and internal transcribed
spacer (ITS)-rDNA studies (Charmet et al. 1997).
Naylor (1960) postulated that the inbreeding group
as evolved from the outbreeding group.
Hybridization among species in Lolium is often very
easy with L. perenne, L. multiflorum, and L. rigidum
crossing very freely when and where their flowering
times overlap. The outbreeders, L. perenne and L. multi-
florum, also cross with the inbreeders, L. temulentum,
L. subulatum, and L. remotum, although some reci-
procal differences exist (Jenkins 1954; Terrell 1966;
Charmet et al. 1996). A number of the Lolium species
also cross very easily with Festuca pratensis and
F. arundinacea to form Festulolium of which there
Fig. 10.2 The panicle of Lolium temulentum have been over 20 cultivars developed (Charmet et al.
1996; Yamada et al. 2005). Some crosses have also
Table 10.1 DNA contents and breeding system in Lolium resulted in the development of a cytoplasmic male
species sterility system in L. perenne (Wit 1974).
Species Mode of Habit DNA content L. temulentum, or darnel ryegrass, has been proposed
breeding (pg/2C) as a model species for genomic studies of cool-season
L. perenne Outbreeder Perennial 4.15 (Evans et al. forage and turf grasses due to its close relationship to
1972)
the important cultivated Lolium species, its short life
L. multiflorum Outbreeder Biennial 4.31(Rees and
Jones 1967) cycle (2–3 months and lack of vernalization require-
L. rigidum Outbreeder Annual 4.33 (Hutchinson ment), and its diploid self-fertile nature (Baldwin and
et al. 1979) Dombrowski 2006).
L. canariense Outbreeder Annual 4.23 (Hutchinson L. temulentum has become valuable for the study of
et al. 1979)
self-incompatibility in Lolium. The outbreeders, peren-
L. edwardii Inbreedera Annual 6.28 (Stewart
Unpublished) nial ryegrass and Italian ryegrass, have the multiallelic
L. temulentum Inbreeder Annual 6.23 (Hutchinson two-locus (S, Z) system of self-incompatibility (Cornish
et al. 1979) et al. 1979), while the self-compatibility of L. temulen-
L. remotum Inbreeder Annual 6.04 (Hutchinson tum is controlled in a gametophytic manner, derived by
et al. 1979)
a single gene mutation of either the Z-locus or a locus
L. persicum Inbreeder Annual 6.35 (Hutchinson
et al. 1979) tightly linked to it (Thorogood and Hayward 1992).
L. subulatum Inbreeder Annual 5.49 (Hutchinson The ease of hybridization between L. temulentum
et al. 1979) and the cultivated Loliums, perennial ryegrass and
a
Probably inbreeding as no crossing was apparent when adjacent Italian ryegrass (Jenkin 1935), has also allowed the
to L. multiflorum possibility of developing inbred lines in perennial
ryegrass and Italian ryegrass in order to capture heter-
been introduced to almost all other temperate regions osis in breeding (Yamada 2001).
by man.
The genus is commonly divided into four outbree-
ders and five inbreeders (Table 10.1). Chromosome
karyotypes do not differ greatly between species 10.3 Genetic Resources
despite the approximately 40% larger chromosome
mass of the inbreeding species (Thomas 1981). There are a number of genetic resource collec-
The differentiation between the four outbreeders tions of Lolium throughout the world, 95% of which
and five inbreeders was also very apparent from are L. perenne and L. multiflorum. The three main
168 H. Cai et al.

Table 10.2 Number of Lolium accessions in major Genebanks The genetic structure of the two outbreeders,
Species Number of accessions L. perenne and L. rigidum, was studied by Balfourier
L. perenne 11,565 et al. (1998) using 12 polymorphic isozyme loci on
L. multiflorum 2,660 starch gel electrophoresis with 120 wild accessions of
L. rigidum 328 L. perenne and 50 of L. rigidum. Genetic diversity
L. temulentum 291
indices [number of alleles (A), observed (Ho) and
L. remotum 47
L. persicum 26 expected (He) heterozygosity] were significantly
L. canariense 16 higher in L. rigidum than in L. perenne. For both
L. subulatum (L. Loliaceum) 5 species, most of the diversity appeared to be within
L. edwardii 2 the populations. In L. perenne, all three genetic indices
(A, Ho, He) showed the same trend of geographical
variation, with the lowest values in the Northwest part
international collections of Lolium are in the USA with
of the distribution area (UK, Ireland) and the highest
the National Plant Germplasm System of the USDA-
ones in the Southeast (Turkey, Lebanon, Cyprus,
ARS (http://www.ars-grin.gov/npgs/index.html), in Eur-
Iraq, Iran). In the same way, as indicated by logistic
ope in the Eurisco system (http://eurisco.ecpgr.org/),
regression analyses between allelic frequencies and
and in the Margot Forde Germplasm Center in New
geographical data of L. perenne populations, the lati-
Zealand (http://www.agresearch.co.nz/seeds).
tudinal gradient of allelic frequencies appeared to be
Table 10.2 lists the total numbers of accessions of
more pronounced, although significant relationships
the nine species, although the actual numbers of dif-
also existed with longitude. In contrast, no spatial
ferent accessions will be less than this because of
organization of the diversity was detected in L. rigi-
planned duplication.
dum. They concluded that L. perenne could be derived
from a small bottleneck of L. rigidum populations in
10.4 Genetic Diversity of L. temulentum, the Middle East, and its present distribution area in
Europe could be explained either by the extension of
L. rigidum, and L. persicum
primitive agriculture from the Fertile Crescent or as a
consequence of postglacial recolonization from south-
Two studies of the diversity with L. temulentum have ern refugia. Further chloroplast studies by Balfourier
been carried out with similar results. Senda et al. (2004) et al. (2000) and McGrath et al. (2007) are also con-
studied the genetic diversity and relationships of 48 sistent with this conclusion.
accessions from eight countries using seven simple Genetic diversity of Iranian natural populations of
sequence repeat (SSR) markers and 44 amplified frag- L. persicum was studied using 12 genomic and 20
ment length polymorphism (AFLP) polymorphic loci. EST-SSR primer pairs from tall fescue (Sharifi Tehrani
Kirigwi et al. (2008) studied 41 L. temulentum, three et al. 2008). The analysis showed that the samples
F. arundinacea, two F. glaucescens, and two F. pra- could be clustered into three regional groups relating
tensis genotypes with selected 30 tall fescue employing to their geographic origin (N, NW, and SW).
expressed sequence tag (EST)-SSRs and 32 Festuca 
Lolium (F  L) genomic SSRs. The study by Senda
and coworkers resulted in four distinct geographic clus-
ters: Pakistan–Nepal complex, the Mediterranean 10.5 The Classical and Molecular
region, Ethiopia, and Japan, while Kirigwi’s study
Genetic Studies of L. temulentum
resulted in three clusters and one lone accession.
This clustering and levels of genetic diversity and Other Lolium species
within each group is consistent with the hypothesis
that the origin of L. temulentum lies in the general L. temulentum has been used as a model species
region of Southwest Asia and the Mediterranean for many studies. In seed shattering studies, results
basin and it also reflects its weedy origin. The clear of its hybridization with L. persicum indicated that
groupings of accessions from each region with high L. temulentum and L. persicum exhibited typical non-
coefficient of gene differentiation (Gst ¼ 0.688) indi- shattering (1.6% shattering) and shattering pheno-
cate that genetic exchange between them is limited. types (70.8%), respectively. The F1 hybrids of
10 Lolium 169

L. temulentum  L. persicum shatter, while the F2s bridging species L. multiflorum using a program of
segregate into 15 shattering: 1 non-shattering, indi- backcrossing and selfing.
cating that non-shattering trait is controlled by two Wang et al. (2002) developed an efficient regenera-
recessive nuclear-encoded genes (Senda et al. 2006). tion and transformation system after screening for
Paraquat resistance in L. rigidum population (AFLR1) tissue culture responses using 46 L. temulentum acces-
has been attributed to reduced paraquat translocation. sions. Embryogenic callus formation frequency ranged
Genetic inheritance of paraquat resistance was re- from 1 to 11% across all accessions tested. Embryo-
ported by Yu et al. (2009). The results suggested that genic calluses of a few responsive accessions were used
paraquat resistance in AFLR1 is inherited as a domi- to establish cell suspension cultures. The regeneration
nant or partially dominant nuclear-encoded trait. Based frequency of green plantlets from the established cell
on the results of pseudo-F2 generation seedlings treated suspension ranged from 15 to 39%. After transferring
with multiple dose rates sufficient to control the sus- the regenerants to the greenhouse, fertile plants were
ceptible parental population, and further confirmed by readily obtained without any vernalization treatment.
individual phenotyping of cloned plant genotypes. In addition, Wang et al. (2005b) also attempted
Therefore, the evolved paraquat resistance in AFLR1 is to improve tissue culture response of L. temulentum to
likely to be controlled by a single major nuclear gene. anther culture and doubled haploid production.
L. temulentum was also used for many plant physi- Ge et al. (2007) reported Agrobacterium tume-
ological studies, including salt stress (Baldwin and faciens-mediated transformation of L. temulentum. A.
Dombrowski 2006; Dombrowski et al. 2008), gibber- tumefaciens strain EHA105 harboring pCAMBIA1301
ellin structure, flowering (Evans et al. 1990; Gocal and pCAMBIA1305.2 vectors were used to infect
et al. 1999, 2001; King et al. 2001, 2008), and others embryogenic callus pieces. Hygromycin was used as a
(Gay and Thomas 1995; Gallagher and Pollock 1998, selection agent in stable transformation experiments.
2004; Baldwin et al. 2007). The transgenic nature of the regenerated plants was
Dombrowski and Martin (2009) identified and eval- checked by PCR, Southern hybridization analysis, and
uated reference genes for use in real-time quantitative b-glucuronidase (GUS) staining. Progeny analysis
PCR (RT-PCR) for abiotic stress studies in L. temulen- showed Mendelian inheritance of the transgenes.
tum. The analysis found that among nine L. temulentum Dalton et al. (1999) reported the cotransformation in
housekeeping genes, eEF-1a and UBQ5 were the most Lolium including L. temulentum produced by micro-
stable and ACT11 was the least stable of the genes projectile bombardment. In their report, 37 plants (30
tested. Analysis by geNorm software indicated that the L. multiflorum, six L. perenne, and one L. temulentum)
two most stably expressed housekeeping genes (eEF-1a were regenerated from cell suspension colonies bom-
and UBQ5) should be utilized and that are sufficient for barded with plasmid DNAs encoding a hygromycin
normalization of gene expression during stress-related resistance gene expressed under a CaMV35S promoter
studies in L. temulentum. Gocal et al. (2001) has also and a GUS-gene expressed under a truncated rice actin1
reported the expressions of two APETALA1 (AP1)-like promoter and first intron, or a maize ubiquitin promoter
genes, LtMADS1 and LtMADS2, and of L. temulentum and first intron. PCR analysis showed that the co-
LEAFY (LtLFY) gene during floral induction. transformation frequency of the GUS-gene varied from
33 to 78% of transformants, while histochemical staining
of leaf tissue from soil-grown plants showed that the co-
expression frequency varied from 37 to 50%.
10.6 Introgression Studies and Genetic
Transformation of L. temulentum
10.7 Genomics Resources: ESTs and SSR
Thomas et al. (1999) reported the transfer of the stay- Markers
green phenotype from F. pratensis into L. temulentum.
A mutant allele at the nuclear locus sid confers indefi- Unlike the two cultivated species of Lolium, there are
nite greenness on senescing leaves of the pasture grass only limited EST sequences of other Lolium species.
F. pratensis. The mutant allele (designated sidy) was In PlantGBD (http://www.plantgdb.org/), 6,336 EST
introgressed into L. temulentum strain “Ceres” via the sequences of L. temulentum can be found (May 2009).
170 H. Cai et al.

Baldwin and Dombrowski (2006) constructed a were screened on a subset of eight genotypes of
salt-stressed subtraction library of L. temulentum to L. temulentum. The selected 30 tall fescue EST-SSRs
evaluate its utility as a model grass to study salt stress. and 32 F  L genomic SSRs were used for further
A total of 528 unique sequences were identified, analysis of genotypes. The TF-EST- and the F  L
among which 167 corresponded to orthologs of previ- genomic-SSRs identified 10.3 and 9.3 alleles per
ously identified plant stress response genes. Baldwin marker, respectively, with an average PIC value of
et al. (2007) has also constructed a post harvest 0.66 (Kirigwi et al. 2008).
subtraction library of L. temulentum, and a total of Sharifi Tehrani et al. (2008) also tested the trans-
598 unique sequences were identified. Gallagher and ferability of 12 genomic and 20 EST-SSR primer pairs
Pollock (1998) also reported a cDNA library from leaf from F. arundinacea to representatives of Iranian
tissues harvested at different times. natural populations of L. persicum. The percentage
The genomes of all eukaryotes contain a class of of transferable polymorphic loci were about 75% for
sequences termed SSRs (Tautz et al. 1986) or micro- both genomic SSRs and EST-SSRs, and the PIC
satellites (Litt and Luty 1989). Microsatellites with values of utilized markers were 0.278 for EST-SSRs
tandem repeats of a basic motif of <6 bp have and 0.219 for genomic-SSRs.
emerged as an important source of ubiquitous genetic Overall, in wild species of Lolium, L. temulentum
markers for many eukaryotic genomes (Wang et al. has been the subject of most of the studies. To date,
1994). Microsatellite or SSR markers are widely used only a limited number of SSR markers have been
in plant genome analysis. Compared with other marker screened, and there has not been any genetic linkage
types, SSR markers have a number of advantages: they maps constructed. However, it is closely related to
are co-dominant PCR-based markers that occur at a the two important cultivated Lolium sp. (Italian and
high frequency in eukaryotic genomes (Li et al. 2002) perennial ryegrass) those that have rich genomic
and are generally associated with non-repetitive DNA recourses, such as bacterial artificial chromosome
regions (Morgante et al. 2002). They are also highly (BAC) libraries (Fujimori et al. 2004; Farrar et al.
reproducible across populations within the same spe- 2007), high density RFLP and SSR linkage maps
cies, and to some extent across species and genera (Jones et al. 2001, 2002; Inoue et al. 2004; Hirata
(Eujayl et al. 2004; Wang et al. 2004, 2005a; Mian et al. 2006), and more than 50,000 ESTs (Sawbridge
et al. 2005). At present, a large numbers of SSR et al. 2003; Ikeda et al. 2004), and their basic genome
markers have been developed in perennial ryegrass information should be easily established. In addition,
(Jones et al. 2002) and Italian ryegrass (Hirata et al. the genetic transformation and its advantages in plant
2006), and the related tall fescue species (Festuca physiological studies will allow L. temulentum to be
arundinacea) (Saha et al. 2004, 2005). widely used in genomic studies of cool-season forage
Senda et al. (2003) have screened a genomic library and turf grasses as a model species.
of L. temulentum with synthetic (GT)15 and (GA)15
oligonucleotides to identify microsatellite-containing
clones. After sequencing, primer design, and testing,
15 primer pairs were found to amplify single frag- 10.8 Endophytic Fungi
ments and five of them showed polymorphism in
L. temulentum and L. persicum accessions. Fifteen There are a number of fungal endophytes of the genus
primer pairs reported in L. perenne were also tested, Epichloe¨ or their Neotyphodium anamorphs, which
nine of them amplified the microsatellite alleles, coevolved with Lolium (Schardl et al. 2004). The
and two of the nine showed polymorphic alleles in annual species, L. multiflorum, L. rigidum, L. temulen-
L. temulentum and L. persicum. For the polymor- tum, and L. canariense contain N. occultans. L. edwar-
phic markers, polymorphic information content (PIC) dii has, however, been shown to have an endophyte
values ranged from 0.60 to 0.86 with an average of N. typhinum var. canariense, a derivative of E. typhina
0.76 among 10 L. temulentum accessions. (Moon et al. 2000; Stewart 2004). L. perenne contains
Tall fescue SSRs were also tested for using in N. lolii and also another type known as LP-2. These
L. temulentum. A total of 40 tall fescue (TF) EST- endophytes can have very significant effects on the
SSRs and 60 Festuca–Lolium (F  L) genomic SSRs ecology of the plants through some very potent
10 Lolium 171

chemistry. N. occultans produces lolines, which are Balfourier F, Imbert C, Charmet G (2000) Evidence for phylo-
known to be potent insecticides but with little effect on geographic structure in Lolium species related to the spread
of agriculture in Europe. A cpDNA study. Theor Appl Genet
grazing animals. The endophytes of L. perenne can 101:131–138
produce a number of alkaloids. These include lolitrems, Broster JC, Pratley JE (2006) A decade of monitoring herbicide
peramine, ergovaline, and janthitrems. Lolitrems are resistance in Lolium rigidum in Australia. Aust J Exp Agric
known to cause staggers in grazing animals; peramine 46:1151–1160
Charmet G, Balfourier F, Chatard V (1996) Taxonomic relation-
is known to be insecticidal to some insects; and ergo- ships and interspecific hybridization in the genus Lolium
valine can have both insecticidal effects and animal (grasses). Genet Resour Crop Evol 43:319–327
health effects. Ergovaline causes vascular constriction Charmet G, Ravel C, Balfourier F (1997) Phylogenetic analysis
and heat stress in grazing animals. Janthitrem has very in the Festuca–Lolium complex using molecular markers
and ITS rDNA. Theor Appl Genet 94:1038–1046
potent insecticidal effect and possible livestock tremo- Clayton WD, Renvoize SA (1986) Genera Graminum. Grasses
genic activity. Recently, New Zealand endophytes, of the world. Her Majesty’s Stationery Office, London
which produce the insecticidal compound but are Cornish MA, Hayward MD, Lawrence MJ (1979) Self incom-
safe for grazing animals, have been incorporated into patibility in ryegrass. I. Genetic control in diploid Lolium
perenne L. Heredity 43:95–106
cultivars. The endophyte in L. perenne might well Dalton SJ, Bettany AJE, Timms E, Morris P (1999) Co-
have assisted the evolutionary development of peren- transformed, diploid Lolium perenne (perennial ryegrass),
niality in this genus of annual species. Lolium multiflorum (Italian ryegrass) and Lolium temulen-
tum (darnel) plants produced by microprojectile bombard-
ment. Plant Cell Rep 18:721–726
Darbyshire SJ (1993) Realignment of Festuca subgenus
Schedonorus with the genus Lolium (Poaceae). Novon
10.9 Conclusion 3:239–243
Dombrowski JE, Martin RC (2009) Evaluation of reference
genes for quantitative RT-PCR in Lolium temulentum
The wild relatives of Lolium are of considerable under abiotic stress. Plant Sci 176:390–396
Dombrowski JE, Baldwin JC, Martin RC (2008) Cloning and
importance as a resource for the improvement of the
characterization of a salt stress-inducible small GTPase gene
widely cultivated grasses, L. multiflorum and L. per- from the model grass species Lolium temulentum. J Plant
enne. They contain endophytic diversity as well as Physiol 165:651–661
valuable genetic traits, such as non-shattering, self- Easton HS (2006) Grasses and Neotyphodium endophytes:
co-adaptation and adaptive breeding. Euphytica 154:
compatibility, larger grain sizes, and potentially
295–306
many other features as well. Until now, only 715 Eujayl I, Sledge MK, Wang L, May GD, Chekhovskiy K,
wild relatives of Lolium have been conserved in ex Zwonitzer JC, Mian MA (2004) Medicago truncatula EST-
situ genebanks, and more collections will be needed, SSRs reveal cross-species genetic markers for Medicago
spp. Theor Appl Genet 108:414–422
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.
Chapter 11
Panicum

Hem S. Bhandari, Masumi Ebina, Malay C. Saha, Joseph H. Bouton, Sairam V. Rudrabhatla,
and Stephen L. Goldman

11.1 Introduction (Clayton et al. 2006), which include 68 species of

Panicum under revised taxonomic delimitation. The
summary of the description for these 68 species is
The position of the grasses and their impact on the
found in Table 11.1.
entire globe cannot be undervalued. Indeed, the
Panicum species are either annual or perennial in
grasses are virtually ubiquitous and have been calcu-
growth habit with rhizomatous or stoloniferous roots.
lated to cover approximately 20% of the Earth’s sur-
Lateral branches are lacking or sparsely present aris-
face (Mathews 1983). Given the role the grasses play
ing from lower culms. Culms are erect or geniculately
in the nourishment for both animals and humans,
ascending or decumbent, which can be as short as
nearly 50% of the planetary digestible energy can be
2–6 cm (Panicum mohavense) to as long as 400 cm
ascribed to these plants (Evans 1993). Moreover, with
(P. altum; P. decidum) (Hitchcock 1971; Watson and
increasing demands for cheaper sources of energy,
Dallwitz 1994; Clayton et al. 2006). Culms are com-
grasses are likely to play a key role, given the oppor-
prised of internodes and nodes.
tunity to derive cellulosic-derived biofuels. Whether
These species have well developed terminal or
the Panicum species will figure prominently energy in
axillary inflorescence called a panicle. Axillary inflor-
economics remains to be seen. However, some Pani-
escences can either be compact or enclosed. In species
cum species reproduce by apomixis and it should be
such as P. campestre, P. cayennense, and P. rudgei,
possible to isolate and clone those genes responsible
axillary inflorescences are intermingled with terminal
for the stabilization of genotypes through maternal
inflorescences. Panicles are well exerted or embraced
inheritance. This in turn, would open the possibility
at the base by subtending leaf or are subtended by the
of transferring clonal propagation to sexually reprodu-
inflated leaf sheath. The peduncle can be smooth or
cing plants. The importance of transfer of those genes
hispid as well as glabrous, pubescent, hirsute, or pilose
encoding apomixis from Panicum on the economics of
above and up to 50 cm long (i.e., P. longissimum).
caloric ingestion remains to be determined.
Panicles are open or effuse in addition to being ovate,
oblong, or elliptic and can be 1 cm (P. mohavense)
to 65 cm (P. ligulare) long. Primary branches are
11.2 Botany appressed, ascending, or spreading. Panicle axis is
either smooth or scaberulous.
Description of over 460 species of grasses previously Spikelets comprise one basal sterile floret and one
classified under Panicum genera are available in fertile floret without rachilla extension. Fertile florets
Royal Botanical Garden, Kew “GrassBase” database are complete and are self-fertile (many diploids) or
self-incompatible (almost all tetraploids). Spikelets
are ovate, lanceolate, or elliptic as well as dorsally
S.L. Goldman (*)
Department of Environmental Science, The University of
compressed, obtuse, acute, or acuminate, and as short
Toledo, 2801 W. Bancroft, Toledo, OH 43606, USA as 1.2 cm (P. philadelphicum) to 9 cm (P. cervicatum)
e-mail: [email protected] long. Spikelets are usually appressed and are often

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 175
DOI 10.1007/978-3-642-14255-0_11, # Springer-Verlag Berlin Heidelberg 2011
176 H.S. Bhandari et al.

Table 11.1 Morphological variability and growth habit of different Panicum species
Species Growth Culm Culm Leaf length Leaf width Panicle Panicle type
habit typea length (cm) (cm) (mm) length (cm)
Section: Dichotomiflora
Panicum aquaticum Annual E or GA 25–200 6–30 4–8 4–27 Open/ovate
Panicum coloratum Perennial E or D 15–100 12–30 5–10 4–30 Open/oblong
Panicum Annual GA or D 100–200 12–50 3–12 12–40 Open/ovate
dichotomiflorum
Panicum Perennial na 80–100 30–40 11–20 15–30 Open/ovate
elephantipes
Panicum gouinii Perennial E 10–50 5–13 2–4 5–10 Contracted/
lanceolate
Panicum lacustre Perennial E 100–150 15–30 2 20–30 Open/ovate
Panicum pedersenii Perennial na 70–100 11–27 4–7 8–23 Open/ovate/effuse
Panicum repens Perennial E or GA 30–100 7–25 2–8 5–20 Open/oblong
Panicum sublaeve Perennial E of GA 15–20 8–10 5–10 5–8 Open/ovate
Panicum Annual D 50–70 5–10 3–8 4–7 Open/Laneolate
vaseyanum
Panicum alatum Annual E or GA 13–95 2–18 3–17 4–23 Open/pyramidal/
effuse
Section: Panicum
Panicum aquarum Annual E 90–130 11–18 2–4 35–40 Open/pyramidal
Panicum aztecanum Perennial E 50–100 14–30 1–3 15–18 Open/pyramidal/
effuse
Panicum bergii Perennial na 30–60 10–37 1–6 10–20 Open/pyramidal/
effuse
Panicum capillare Annual E of GA 20–80 7–30 5–14 15–50 Open/oblong/ovate/
effuse
Panicum Perennial E of GA 30–60 20–50 2–12 10–30 open/oblong/effuse
capillarioides
Panicum chaseae Perennial na 40 na 2–3 15–23 Open/ovate
Panicum Annual na 30–60 8–15 7–10 10–18 Open/Lanceolate
decolorans
Panicum diffusum Perennial GA or D 20–100 5–220 1–5 5–25 Open/Pyramidal
Panicum Annual E 60–70 20–30 2 30 Open/elliptic/effuse
ephemeroides
Panicum exiguum Annual D 10–30 2–6 2–4 3–8 Open/pyramidal/
effuse
Panicum fauriei Annual na 2–30 2–12 1–4 1–11 Open/linear
Panicum flexile Annual E 12–75 6–25 2–7 7–20 Open/oblong
Panicum furvum Perennial E 10–21 2–7 2–4 4–6 Open/ovate
Panicum Perennial GA or D 35–110 11–30 4–8 20–35 Open/ovate/
ghiesbreghtii pyramidal
Panicum hallii Perennial E or GA 20–80 4–30 2–10 6–20 Open/pyramidal
Panicum hirsutum Perennial na 100–130 35–70 10–30 15–50 Open/elliptic/dense
Panicum hirticaule Annual E/GA/D 25–80 7–27 6–15 13–35 Open/ovate
Panicum Annual na 50 15 8 23 Open/ovate
hispidifolium
Panicum lepidulum Perennial E 25–70 7–30 5–10 7–20 Open/elliptic
Panicum Perennial na 20–30 10–12 2 3–6 Open/ovate
magnispicula
Panicum miliaceum Annual E or GA 20–100 10–20 6–15 5–10 Contracted/obovate
Panicum Annual na 2–6 1–4 2–3 1–2 Open/oblong/dense
mohavense
Perennial na 30–90 25–60 5–15 30–40
(continued)
11 Panicum 177

Table 11.1 (continued)

Species Growth Culm Culm Leaf length Leaf width Panicle Panicle type
habit typea length (cm) (cm) (mm) length (cm)
Panicum Open/oblong/
mucronulatum obovate
Panicum Perennial na 45–240 10–40 8–25 15–60 Open/elliptic/ovate
nephelophilum
Panicum Perennial GA NA 5–12 5–10 6–15 Open/ovate
pampinosum
Panicum parcum Annual E 50–125 11–35 6–10 20–35 Open/ovate
Panicum Perennial E 20–60 7–22 2–4 7–18 Open/ovate
peladoense
Panicum Annual E 20–50 5–15 2–7 7–15 Open/ovate
philadelphicum
Panicum Perennial na 40–120 10–40 4–10 7–20 Open/ovate
quadriglume
Panicum Annual GA 20–50 10–30 4–15 5–15 Open/ovate
stramineum
Panicum Perennial E 50–150 10–50 3–8 15–40 Open/pyramidal
tamaulipense
Panicum campestre Perennial E or GA 30–74 7–25 5–7 5–15 Open/oblong/effuse
Section: Rudgeana
Panicum Annual E or GA 18–40 9–13 5–8 5–16 Open/ovate
cayennense
Panicum Perennial na 50–150 12–40 7–18 30–50 Open/oblong/ovate
cervicatum
Panicum ligulare Perennial E 130–200 30–55 9–19 47–65 Open/ovate/effuse
Panicum rudgei Perennial E or GA 75–125 20–40 8–14 30–40 Open/ovate/effuse
Panicum Perennial E 40–70 15–30 1–4 10–30 Open/ovate/effuse
chloroleucum
Section: Urvilleana
Panicum Perennial na 45–55 23–43 1–4 11–25 Contracted/linear
racemosum
Panicum Perennial E 50–100 10–20 4–7 25–30 Open/elliptic
urvilleanum
Panicum altum Perennial D 200–400 30–45 8–15 20–30 Open/ovate
Section: Virgata
Panicum amarum Perennial E 30–200 10–50 3–15 10–35 Contracted/
lanceolate
Panicum decidum Perennial na 200–400 7–9 4–8 3–7 Open/ovate/dense
Panicum glabripes Perennial E 70–130 25–45 4–5 20–26 Open/ovate
Panicum Perennial E 150–200 50–80 5–9 10–30 Contracted/linear
longissimum
Panicum Perennial na 150–200 25–35 7 30–40 Open/pyramidal
tricholaenoides
Panicum virgatum Perennial E, GA, D 30–300 10–60 3–15 15–55 Open/ovate
Source: Aliscioni et al. (2003) and Clayton et al. (2006)
a
E erect, GA geniculately ascending, D decumbent

solitary or occasionally in pairs. Fertile spikelets are vegetative leaf development, stem elongation, devel-
pedicelled. Pedicel can be clavate, scabrous, glabrous, opment of reproductive structures (panicle initiation to
or hairy at tip. Fruit is in the form of caryopsis with anthesis), and seed development and ripening are all
adherent pericarp. established. These stages are further broken down into
Plant growth stages of Panicum species follow that several substages. The duration of each stage, how-
of most perennial forage grasses (Moore et al. 1991). ever, varies according to species, ecotypes within
Five major growth stages including germination, species, genotype within each ecotype, and responses
178 H.S. Bhandari et al.

to growth conditions such as photoperiod, tempera- various stresses, such as heat and drought. Data from
ture, etc. For example, upland ecotypes of P. virgatum 14 different Panicum and other grass species indicated
L. attain maturity several days earlier than lowland that the C4 type has higher rate of photosynthesis and
ecotypes. Yet, maturity may become delayed in both immediate export compared to C3 species (Leonardos
ecotypes as one goes to northern latitudes. and Grodzinski 2000). This has made C4 species a crop
Traditionally, different species in Panicum were of choice in most of the tropical and subtropical regions
believed to possess either C3 or C4 or C3–C4 inter- of the world.
mediate photosynthetic pathway (Brown et al. 1985;
Bouton et al. 1986; Leonardos and Grodzinski
2000). The differences between C3, C4 and C3–C4
intermediate species of Panicum and other grasses in
leaf anatomy, physiology, and biochemical processes
11.3 Allied Crops
have been documented (Bouton et al. 1986; Warner
et al. 1987; Dengler et al. 1994). Existence of C3/C4 The Paniceae exhibits great diversity with respect to
pairs of closely related species as well as their molec- morphology, anatomy, photosynthetic physiology
ular phylogeny results suggests the possible two-way and karyology (Clayton and Renvoize 1986; Watson
evolutionary reversal between C3 and C4 grasses and Dallwitz 1994). Indeed, significant attempts were
(Duvall et al. 2003; Christin et al. 2009; Ibrahim made to use this information prior to the development
et al. 2009). The C3–C4 intermediate types possess of molecular markers to understand the taxonomic
Krantz like leaf anatomy that show reduced photores- relationships among related plants. On this basis, at
piration and reduced O2 inhibition of apparent photo- one time, Panicum was divided into six subgenera.
synthesis compared to their C3 counterpart (Brown These include Panicum, Dicanthelium, Steinchisma,
and Morgan 1980). It is noteworthy that closely Agrostoides, Megathyrus, and Phanopyrum (Zuloaga
related pair of C3 and C3–C4 species can hybridize et al. 2000; Giusanni et al. 2001; Aliscioni et al. 2003;
and produce viable progeny (Brown et al. 1985; Simon and Jacobs 2003; Reinheimer et al. 2005).
Bouton et al. 1986). The interspecific hybrid between The cytolological, anatomical, and morphological
C3 and C3–C4 type produced F1, which showed leaf differences and, for that matter, similarities are sum-
anatomy, morphology, O2 inhibition, and CO2 com- marized in Table 11.2. Today, each has been elevated
pensation intermediate of the two parent types. to an independent genus and more are being added
Several species of C3 or C3–C4 intermediate types of (Morrone et al. 2008).
photosynthesis pathway that were previously described
under Panicum are now assigned to other genera
(Aliscioni et al. 2003; Zuloaga et al. 2006). Apparently,
almost all of the Panicum species under revised
description follow C4 type of NAD–me photosynthetic
11.4 Distribution
pathway (Giussani et al. 2001; Aliscioni et al. 2003;
Zuloaga et al. 2006; Christin et al. 2009). These species Panicum species are adapted to tropical, subtropical,
have specialized, centrifugally placed chloroplasts in and warm temperate regions throughout the world.
the bundle sheath, without well-developed grana, and Some species are distributed more widely than others.
have lost the parenchymatous outer sheath around the For example, P. miliaceum and P. capillare of section
vascular bundles. Deviation from these structures has Panicum are found adapted to tropical, subtropical and
also been reported in Panicum species, i.e., P. prionitis temperate regions of countries in all the continents,
preserves the outer bundle sheath (Giussani et al. 2001). while P. virgatum is found primarily in the US,
The C4 photosynthesis consists of specialized anatomi- Mexico, and Brazil in addition to other countries of
cal structures and biochemical novelties that create CO2 South America, regions of Mesoamerica, and the Carib-
pump and build up CO2 concentration around Rubisco bean. Three species, P. coloratum, P. dichotomiflorum,
and reduce photorespiration (Christin et al. 2009). Due and P. repens, of section Dichotomiflora are distributed
to these features, the C4 species are well known for in tropical, subtropical, and temperate environments in
competitive adaptive advantage over C3 species under countries in North America, South America, Africa,
 11 Panicum

Table 11.2 Anatomical, morphological, and cytological similarities among Genera related to Panicum
Trait Panicum Dicanthelium Steinchisma Megathyrus Agrostoides Phanopyrum Parodioph-
yollochloa
Chromosome # x¼9 x ¼ 10 x ¼ 10 Polymorphic Section dependent Section dependent x ¼ 10
x ¼ 8 or 9 x ¼ 8 or 9 x ¼ 9 or 10
Culm Erect and cespitose North American – – – – –
types express
distinctive basal
rosettee
Vascular bundles 2 sheaths around – – – Mesotome sheath – –
bundles outer specialized
sheath contains chloroplasts
specialized
centripede
chloroplasts
Inflorescence Pyramid like – – Lax pyramid – like – Lax and open Lower flower,
many flowered lower palea
panicle absent
Spikelet Ellipsode to – Open or contracted – – –
lancelolate panicles
Glumes 7–9 nerved – – Upper glume and – Upper glume and Lower glume is
lower lemma 3–5 lower lemma 1/2 length
nerved 7–9 nerved
Anthecium Compound or – Entire upper surface Upper/transverse – – Upper anthecium
simple papillae covered with cpd. rugose þ is indurate
papillae papillae on with simple
wrinkles pillae
Photosynthesis C4 – C3/C4 intermediate C4 C4 – –
179
180 H.S. Bhandari et al.

Europe, Asia, and Australasia. Interestingly, the adap- (Nakagawa and Hanna 1990; Hamoud et al. 1993;
tation of large number of Panicum species is restricted Morrone et al. 1995, 2006; Zuloaga et al. 2006).
to countries in the western hemisphere; US, Mexico,
the Caribbean, Mesoamericana, Brazil, and other
countries in South America. Brazil and its neighboring 11.7 Origin of the Genera and Molecular
regions possess diverse species of Panicum. Of the
100 species of Panicum, 50 are of American origin,
Phylogeny
which are distributed from Canada through Argentina
(Guglieri et al. 2006); 29 were found in Brazil alone Phylogeny of the tribe Paniceae, subfamily Panicoi-
(Guglieri et al. 2004). deae, was analyzed with sequence data obtained from
the chloroplast gene ndhF (Giussani et al. 2001). Most
of the proteins involved in C4 photosynthesis are
encoded by nuclear genes. Therefore, a chloroplast
gene should provide an independent history from the
11.5 Conservation Initiatives genes selected for C4 photosynthesis. The ndhF gene
indicates a relatively high rate of molecular evolution
Two major global initiatives have been started to in the grasses (Clark et al. 1995). Phylogenetic analysis
appropriately track and catalog the world’s germplasm of 103 species of subfamily Panicoideae clearly
and derivative genetic resources. These are the System- grouped into three clades, corresponding largely to
wide Information Network for Genetic Resources the same basic chromosome number (Andropogoneae
(SINGER) and the Consultative Group on Interna- [x ¼ 10], Paniceae [x ¼ 10], and Paniceae [x ¼ 9]).
tional Agricultural Research (CGIAR). SINGER and With a few exceptions, most members of the x ¼ 9
CGIAR provide a resource to determine where partic- Paniceae clade have nine chromosomes. The x ¼ 9
ular seed lines may be found and, in addition, provide clade includes members of five subgenera of Panicum
critical information related to the collection source, (subg. Panicum [C4], Dichanthelium [C3], Agrostoides
the status of the sample, and whether the seed is [C4], Phanopyrum [C3], and Megathyrus [C4]). The
derived from domesticated or wild lines. The location subgenus Panicum was characterized as monophyletic,
of an abundance of molecular resources is also listed. and Dichanthelium appeared as polyphyletic. P. sabu-
The precise location of Panicum germplasm and lorum and P. koolauense of the subgenus Dichanthelium
accompanying molecular capital are indicated in the were unrelated to the other member, P. ovuliferum, of
body of this review as appropriate. Unlike corn, soy- this subgenus. Many of the Panicum species (P. virga-
bean, and wheat, for example, the databases and seed tum, P. elephantipes, P. nephelophilum, P. fauriel,
resources that are highly centralized, Panicum assets P. rudgel, P. repens, P. pedersenil, P. aquaticum,
are more widely dispersed in smaller collections, those P. bulbosum) fall into a group called Setaria/Uro-
in India, Japan, and along the Pacific Rim being among chloa/Panicum clade (supported by 88% bootstrap
the more notable. with 6 bp substitutions). All species in this clade
exhibit C4 photosynthesis (Giussani et al. 2001). This
study was conducted with large number of species and
further expanded the work of Gomez-Martinez and
Culham (2000) who divided the tribe Paniceae into
11.6 Cytology and Karyotype two clades corresponding to chromosome number cor-
related with biogeography (American Paniceae
As in most species of the Poaceae, the karyotype of [x ¼ 10] and Pantropical Paniceae [x ¼ 9]).
most Panicums is symmetrical and uniform with Aliscioni et al. (2003) further expanded the classi-
metacentric or submetacentric chromosomes (Zuloaga fication study of Giussani et al. (2001). A total of 123
et al. 1989). Chromosomes paired predominantly as sequences were analyzed, of which 111 represent the
bivalent. However, few monovalent and quadrivalent tribe Paniceae (66 representing Panicum s.l.). Pani-
in tetraploids and univalent, quadrivalent, or hexa- cum phylogeny was estimated from a parsimony anal-
valent in hexaploid cytotypes were also reported ysis based on all available chloroplast ndhF gene
11 Panicum 181

Table 11.3 Chromosome number and ploidy level of different Panicum and other related grass species and their configuration
during dikenensis of metaphase Ia
Taxon Basic chr. # Ploidy Configuration during dikenensis Source
of metaphase
P. Capillaire 9 2x 8 II þ 2 I Hamaud et al. (1994)
P. coloratum 9 2x, 4x 9 II “or” 18 II Hamaud et al. (1994)
P. cyanescens 18 4x 18 II or 17 II þ 2 I Morrone et al. (1995)
P. hirticaule 9 2x 9 II Morrone et al. (2006)
P. miliaceum 9 4x 18 II þ few univalent Hamaud et al. (1994)
P. virgatum 9 4x, 6x, 8x 18 II “or” 35 II þ 2 I Lu (1995)
P. cipoens 20 4x 20 II Morrone et al. (1995)
P. euprepes 20 4x 1 IV þ 18 II “or” 19 II þ 2 I Morrone et al. (1995)
P. molinioides 20 4x 19 II þ 2 I or 1 IV þ 17 II þ 2 I Morrone et al. (1995)
or 2 IV þ 16 II
P. panicillatum ca. 27 6x 27 II Morrone et al. (1995)
P. poliophyllum 20 4x 20 II or 1 IV þ 18 II Morrone et al. (1995)
or 1 IV þ 17 II þ 2 I
P. stenophyllum 10 2x 10 II Morrone et al. (1995)
P. stoloniferum 10 2x 10 II Morrone et al. (2006)
P. trichanthum 18 4x 18 II Morrone et al. (2006)
P. validum 10 2x 10 II Zuloaga et al. (1989)
P. wettsteinii 9 2x 9 II Morrone et al. (1995)
P. assurgens 9 2x 9 II Morrone et al. (1995)
P. cabrerae 9 2x 9 II Morrone et al. (1995)
P. heliophilum ca.30 6x Ca. 30 Morrone et al. (1995)
P. maximum 10 2x 10 II Nakagawa and Hanna (1990)
P. laevifolium 9 2x 8 II þ 2 I Hamaud et al. (1994)
P. antidotala 9 2x 8 II þ 2 I Hamaud et al. (1994)
P. miliare 9 4x 16 II þ 4 I Hamaud et al. (1994)
a
Previously classified under genus Panicum, #7–23 are now assigned to other genera

sequences of the tribe Paniceae. Panicum sensu stricto different ploidy levels from diploid (2n ¼ 2x ¼ 18) to
(s.s.) was restricted to the former subgenus Panicum dodecaploid (2n ¼ 12x ¼ 108) have been reported
and supported the recognition of separate genera for (Church 1940; Burton 1942; Nielsen 1944). However,
Dichanthelium, Phanopyrum, and Steinchisma. The only two major ploidy levels, the tetraploids (2n ¼ 4
subgenus Panicum was clearly monophyletic (sup- x ¼ 36) and the octoploids (2n ¼ 8x ¼ 72), are pre-
ported by 97% bootstrap) and completely included in dominant (Hopkins et al. 1996). Two cytotypes of
the x ¼ 9 Paniceae clade. switchgrass, U and L, associated with upland and
lowland ecotypes have been recognized (Hultquist
et al. 1996). The lowland cytotypes are entirely tetra-
ploids and the upland cytotypes are either tetraploids,
11.8 Cytogenetics hexaploids, or octoploids (Barnett and Carver 1967).
P. miliaceum and P. sumatrense are tetraploids (2n ¼
Panicum species vary widely in chromosome number 4x ¼ 36) species. P. capillare and P. sonorum are
as well as in ploidy levels (Nielsen 1944; McMillan reported as diploid (2n ¼ 2x ¼ 18) species (Tsvelev
and Weller 1959; Zuloaga et al. 1989; Hamoud et al. and Zhukova 1974; Hiremath et al. 1990).
1993; Morrone et al. 2006) (Table 11.3). Ploidy levels Ploidy level of several P. virgatum populations
vary from diploid to dodecaploid. The basic chromo- have been estimated using microscopy and flow cyto-
some numbers reported for the Panicum genus are metry techniques (Hopkins et al. 1996; Lu et al. 1998).
predominantly 9 or 10 with a few reported to have 7 These studies have demonstrated that plants with aver-
or 8 or 11 (Giussani et al. 2001). In switchgrass, all age nuclear DNA contents of 2.7 to 3.3 pg are
182 H.S. Bhandari et al.

tetraploid populations and 4.7–6.0 pg are octoploid DNA content of 4.76 pg), respectively. Within Cave-
populations. Nuclear DNA contents of the 31 GRIN in- rock, only one seedling each was designated as
switchgrass populations were estimated using flow tetraploid and hexaploid with a mean DNA content
cytometry (Narasimhamoorthy et al. 2008). Twenty of 2.96 and 3.68 pg, respectively. Mixed ploidy levels
accessions were classified as tetraploids, with average were only detected in Cave-in-rock with tetraploid,
DNA content ranging from 2.4 to 3.4 pg. Four culti- hexaploid, and octoploid genotypes being reported
vars, Trailblazer, Caddo, Blackwell, and Argentina, (Narasimhamoorthy et al. 2008).
were clearly classified as octoploids with average P. maximum jacq. (guineagrass), a tropical grass
DNA content ranging from 4.5 to 5.8 pg. The remain- native to Africa, has 2n ¼ 16, 32, and 48 chromo-
ing seven accessions were identified with mixed somes at diploid, tetraploid, and hexaploid levels,
ploidy levels, which include well characterized culti- respectively (Nakagawa and Hanna 1990). Diploid
vars such as Cave-in-rock, Blackwell, Shawnee, and plants of this species follow sexual mode of reproduc-
Alamo (Narasimhamoorthy et al. 2008). However, tion, while its tetraploid counterpart has shown mainly
plants obtained from commercial seed lots of Alamo apomictic mode of reproduction. Aneuploidy, such as
were all tetraploids (mean DNA content of 2.97 pg) 2n ¼ 18, 40, and 44, are also reported for this species.
while Blackwell and Shawnee were octoploids (mean Experimental evidence suggests that the hexaploid

Fig. 11.1 (continued)

11 Panicum 183

cytotypes resulted from fertilization of unreduced the simplex marker loci in AFLP primer combinations
2n ¼ 32 egg from apomictic plants by a reduced 16- was 6.4 in “Natsukaze” compared to 2.6 in the mater-
chromosome male gamete. Interestingly, the tetraploid nal “Noh PL1”. Thus, the sexual tetraploid line accu-
and hexaploid cytotypes are similar in most of the mulates duplex or multiplex loci, as compared to
morphological traits, yet 2n ¼ 40 plants show reduced “Natsukaze.” In contrast, “Natsukaze” has frequent
plant height (Nakagawa and Hanna 1990). The nuclear simplex loci. These simplex markers spread the entire
DNA content of diploid guineagrass genome is esti- linkage map, which covered 1,703.5 cm of the autotet-
mated 500 Mbp (Akiyama et al. 2008). raploid genome (2n ¼ 4x ¼ 32), with an average
In Panicum maximum Jack., linkage studies have spacing of 4.7 cm. The number of map units per
been most effective in analyzing the transmission of linkage group correlated with the number of markers
apomixis. These molecular linkage maps were con- per linkage group (R ¼ 0.68), confirming that the loci
structed by analyzing the progeny derived from a are randomly distributed across the map.
cross between the sexual, colchicine induced, autotet- In a mapping population designed to determine the
raploid “Noh PL1” and the apomict “Natsukaze” number of segregating genes regulating apomixis,
(Ebina et al. 2005). Using various combinations of the ratio of apomicts to sexual was 1:1. Among some
56 amplified fragment length polymorphism (AFLP) apomictic segregants, polyembryo was observed, and
primers and 41 random amplified polymorphic DNA conjugated cellular structure was observed in the
(RAPD) primers, a total of 360 simplex marker loci embryosac as shown in Fig. 11.2. Carman (1997)
were assigned to 39 linkage groups of the cultivar suggested that the polyspory of adventitious embryony
“Natsukaze” as shown in Fig. 11.1. The frequency of and polyembryony of gametophytic apomixis have a

Fig. 11.1 An AFLP and RAPD linkage map of aposporous loci is on the right side. The size of the linkage group is shown in
guineagrass cultivar “Natsukaze” pollen patent. Map distances the bottom of the each group. The aposporous locus (Apo) was
are in cm in left side of the linkage group. The name of marker mapped in the group 8
184 H.S. Bhandari et al.

common origin. Each is a function of asynchronous apomixis region is consistent with their mode of
expressions of sexual embryo construction genes as a reproduction. This study is now being expanded by
consequence of multiple gene copies (Carman 1997). converting these cosegregating AFLP markers into
As predicted by this model, both the sexual progeny sequence tagged markers.
of the mapping population in addition to natural Large numbers of molecular markers, including
sexual accessions never exhibit polyembryony in those completely linked to apospory, have also been
guineagrass. annotated in Paspalum notatum (Martinez et al. 2003;
In linkage group 8, shown in Fig. 11.1, the apo- Stein et al. 2004; Carneiro et al. 2006). In two species,
mixis trait cosegregated with nine AFLP markers a small genetic region is conserved due to the suppres-
mapping near the center of the linkage group. An sion of crossing over in spite of the fact that physical
expanded version of this linkage group 8 is shown in interval marking the chromosome is large. This is
Fig. 11.3. Thirteen AFLP markers flanked the apo- likely to be the first instance of recombination restric-
mixis region at 1.4 cm, and three measured 4.2 cm to tion in Pennisetum squamulatum. The physical size of
the opposite side. The marker composition of all indi- the apomixis-related gene region (ASGR) is 50 Mbp in
viduals of the mapping progeny around the aposporous P. squamulatum (Akiyama et al. 2004) and 40 Mbp in
P. notatam (Carneiro et al. 2006). The magnitude of
ASGR in guineagrass is also similar with apospory-
controlling locus in P. maximum and P. notatum with
restricted recombination across a large physical region
of the chromosome.
By analyzing putative polymorphisms focused on
the ASGR region, an attempt will be made to dissect
the phylogeny of aposporous apomixes among these
Fig. 11.2 Embryo sac analyses of the mapping population. grasses. In addition to these regional polymorphisms,
GM64 is the name of the population. An individual named
the next generation sequencers can directly read the
GM64-20 presents a typical sexual embryo sac. GM64-50 pre-
sents a typical apomixis embryo sac, however, which appears as huge physical region of the ASGR (Conner et al.
a twin embryo and cellar conjugation form 2008). In this study, one-tenth of the region has been

Fig. 11.3 An expanded

version of linkage group
8 containing the aposporous
locus (Apo) and tightly linked
markers. Markers
cosegregating and tightly
linked to Apo are shown in the
boxes at the right of the
corresponding marker
11 Panicum 185

Table 11.4 Nuclear DNA contents determined by flow cytometry and the number of ribosomal DNA (rDNA) loci in Panicum
maximum
Strains Reproduction No. of Analyzed times Mbp 5S 45S
Mode Ploidy chromosomes of DNA content C1 SD rDNA rDNA
GR-297 Sexual 2 16 12 499 3 2 4
Noh PL1 Sexual 4 32 5 995 7 4 8
Natsukaze Apospory 4 32 6 1,023 10 4 6
Natsuyutaka Apospory 4 32 5 1,045 11 4 4

sequenced by using bacterial artificial chromosome

(BAC) contigs from P. squamulatum and C. cilliaris.
In the process, many classes of transposable elements
in the ASGR were discovered. Equally notable, this
sequence has no large-scale synteny with Oryza and
Sorghum genomes. Therefore, not only further com-
parisons of the sequence between apomictic relatives
of P. squamulatum and C. cilliaris but also compar-
isons made with P. maximum and P. notatum need
definition. In principle, this technique should elucidate
common sequences important to the expression of
apospory.

11.9 Ploidy Level and Apomixis-Related

Gene Region Fig. 11.4 Physical mapping of ribosomal DNA (rDNA) on
mitotic chromosome in P. maximum (a) GR-297, (b) Noh PL1,
(c) Natsukaze and (d) Natsuyutaka. Red and Green arrow heads
Cytological analysis was performed after collecting indicate 45S and 5S rDNA, respectively. Bars, 10 mm (Akiyama
natural accessions of guineagrass (Hanna et al. 1973; et al. 2008)
Savidan 1980; Nakagawa and Hanna 1990). Guinea-
grass has a basic chromosome number of x ¼ 8; most
plants are apomictic tetraploid. Using colchicines
treatment, the chromosome number of several diploid apomicts (Table 11.4). Although this difference in
lines was doubled to obtain tetraploid plants (Savidan genome size is not necessarily congruent with the
and Pernes 1982; Nakagawa and Hanna 1992). Given size of apomixis-related region of guineagrass, a simi-
the importance of tropical forage and apomictic lines, lar number of megabases have been reported in
the basic genomic information such as genome size P. sequamulatum (Akiyama et al. 2004). Fluorescent in
and ribosomal DNA (rDNA) ideogram has only situ hybridization (FISH) analysis with rDNA probes
recently been reported (Akiyama et al. 2008). The was performed (Fig. 11.4), which shows the number of
genome of the diploid sexual line of “GR-297” was 5S rDNA loci in diploid and the tetraploids were two
determined to be approximately 500 Mbp. As such, it and four, respectively. The number of 45S rDNA loci
is equal in range to rice, and the three sexual and varied from four to eight among the three tetraploid
apomictic tetraploids accordingly corresponded to plants. Quantitative ideogram of diploid strain by
1,000 Mbp. A detailed comparison of genome size image analysis, as shown in Fig. 11.5, indicated that
between the sexual and apomictic tetraploids detected the difference in the number of 45S rDNA is dependent
a 30–50 Mbp difference in genome size specific to the on the presence or absence of locus on chromosome 2.
186 H.S. Bhandari et al.

DAPI intensity
0 50 100 150 200 250 0 50 100 150 200 250 0 50 100 150 200 250 0 50 100 150 200 250

Black region
Gray region

Chromosome 1 Chromosome 2 Chromosome 3 Chromosome 4

0 50 100 150 200 250 0 50 100 150 200 250 0 50 100 150 200 250 0 50 100 150 200 250

Chromosome 5 Chromosome 6 Chromosome 7 Chromosome 8

Fig. 11.5 Quantitative ideograms and distribution patterns 4,6- GR-297. Closed and open circle indicate loci of 5S and 45S
diamino-2-phenylindole (DAPI) intensities on chromosomes rDNA, respectively (Akiyama et al. 2008)
constructed from nine mitotic chromosome spreads of strain

11.10 Molecular Markers cytotype despite its classification as lowland cytotype

by USDA-SCS (Gunter et al. 1996; Hultquist et al.
1996). The genetic variation in the nuclear gene encod-
Molecular markers have become a reliable tool for the
ing plastid acetyl-coA carboxylase (ACCase) from six
detection of genetic variability because they directly
switchgrass cultivars was investigated using DNA
reveal changes at the DNA sequence level. In the past
sequences (Huang et al. 2003). Tetraploid and octo-
two decades, several studies have been conducted to
ploid switchgrass ecotypes formed two distinct groups
find within and among population variation in P. vir-
and suggested that the most recent polyploidization
gatum using restriction fragment length polymorphism
events that established modern switchgrass lineages
(RFLP), random amplification of polymorphic DNA
occurred less than two million years ago. RFLP mar-
(RAPD), and simple sequence repeat (SSR) molecular
kers were also used to study the genetic variation
markers. Two distinct cytoplasm types, L and U,
among 21 switchgrass genotypes randomly selected
associated with lowland and upland ecotypes, respec-
from two lowland (Alamo and Kanlow) and one upland
tively, were identified using cpDNA polymorphisms
(Summer) synthetic cultivars (Missaoui et al. 2006).
(Hultquist et al. 1996). The lowland cultivars contained
The upland cultivar showed a higher degree of genetic
a restriction site that was not present in the upland
variation than the lowlands. Analysis of chloroplast
cultivars. A broad assessment of the genetic relation-
non-coding region of the trnL DNA sequences indi-
ship among 137 genotypes representing 14 switchgrass
cated a deletion of 49 nucleotides that was specific to
populations were assayed with 91 polymorphic RAPD
lowland cytotypes (Missaoui et al. 2006).
markers (Gunter et al. 1996). The individual genotypes
A large set of microsatellite markers have been
within each population were clustered into discrete
developed from the switchgrass EST sequences
groups within the ecotypes. Both RAPD and cpDNA
(Tobias et al. 2008). Several of these markers were
analyses suggested that Cave-in-rock is an upland
11 Panicum 187

used for the genetic diversity analysis and mapping groups contained Kenyan accessions. These results are
of switchgrass populations. Microsatellite markers consistent with the hypothesis that guineagrass origi-
have also been developed from the (GA/CT)n enriched nates from East Africa, Kenya, and Tanzania. A prin-
switchgrass genomic libraries. DArT markers have ciple coordinate analysis (PCA) scatter-plot indicated
been developed and used for mapping a switchgrass that the 77 guineagrass accessions also were divided
population developed by crossing between a lowland into six major groups, as is shown in as shown in
and an upland genotype (Katrien Devos, UGA per- Fig. 11.7, hinting that the cultivars selected in different
sonal communication). A large set of SNP markers breeding sites belong to different major guineagrass
are being developed for switchgrass at the Cornell groups. Whereas Japanese cultivars selected in warm
University (Ed Buckler, personal communication). temperate zone belong to the group I, the new cultivars
The genetic structure in guineagrass was assessed of semitropical Okinawa belong to the group III. The
by the development of SSR markers (Ebina et al. importance of these SSR markers is that they should
2007). In this report, 13 SSR markers were success- be represented in new accessions and will be key in
fully developed from genomic DNA library enriched establishing new breeding lines and cultivars of gui-
for (AG)/(TC) repeats and SSR motifs survey in neagrass. Indeed, new alleles of the foundation of 13
expressed sequence tag (EST) sequences. From SSR markers are anticipated, thus facilitating mea-
Japanese accessions of guineagrass, 77 randomly sures of phylogenetic relatedness among newly iso-
selected accessions were screened and their phyloge- lated accessions.
netic relationship investigated. These 13 SSR primers
produced 1–4 amplified fragment(s) within accessions
corresponding to the autotetraploidy of guineagrass.
Although the limiting number of SSRs is caused by the 11.11 Crop Improvement
low redundancy of SSR in the genomic region and in
the ESTs, the power of discrimination (PD) and poly- Important observations using guineagrass as a model
morphic information content (PIC) were relatively plant have contributed to our understanding of cytol-
high comparing to the other self-compatible species. ogy, quantitative/population genetics, and numerical
The SSR motifs of guineagrass were 54% perfect, 31% taxonomy during the last 40 years (Combes and Pernes
interrupted, and 14% compound. The ratio of the motif 1970; Combes 1975). Indeed, not only were the dip-
variations is likely to the self-compatible allotetrapoid loid and tetraploid sexually reproducing cultivars dis-
grass species of zoysia grass (Tsuruta et al. 2005). The covered but also the apomictic accessions (Combes
limited frequency of the SSR appearance in guinea- and Pernes 1970; Hanna et al. 1973; Nakajima et al.
grass is an open question. 1979; Nakagawa and Hanna 1992). Hence, not only
The high value of PD and PIC supports the hypoth- could the inheritance patterns of important agronomic
esis of intercrossing among the sexual plants or facul- genes be determined, but these in turn could also be
tative apomicts to create genetic variability in the readily introgressed in both diploid and polyploid
natural habitat. Thirty-nine putative alleles of the lines. Similarly, effective crossing protocols have
SSR primer GNK02-02 segregates in the 77 guinea- been designed to determine the number of segregating
grass accessions where PD equals 0.968. The average genes regulating apomixes in a significant number
number of the putative alleles with 13 SSR primers of genera (Nakajima 1991; Blakey et al. 1997, 2001;
was 14.9, and the average PD value was 0.859. These Ebina and Nakagawa 2001).
values are higher than those reported in zoysia grass Gametophytic apomixis results from asexual repro-
(Tsuruta et al. 2005) and perennial ryegrass (Jones duction, and the resulting embryo is genetically iden-
et al. 2001). tical to the mother from which it was derived. It can
Using these developed SSR primers, a phenogram be classified into two forms, that is, aposporous and
was comprised of a total of 190 putative alleles in the diplosporous apomixis (Nogler 1984). In addition to
77 guineagrass accessions as shown in Fig. 11.6. The guineagrass, many other aposporous apomictic genera
phenogram contained six major groups of the guinea- have been identified among them such tropical grasses as
grass. Although two of the six major groups were not Pennisetum, Paspalum, and Brachiaria species (Asker
contained in any South African accessions, all of the and Jerling 1992; Sherwood 2001). The inheritance of
188 H.S. Bhandari et al.

Genetic similarity
0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0

GR 190 Kenya
GR 293 Tanzania I
GR 243 Kenya
GR 373 Ethiopia
214 Kenya
GR 193 Kenya II-i
73-473 Kenya
211 Kenya
73-790 Kenya
73-1006 Kenya II-ii
73-804 Kenya
GR 161 Unknown
GR 248B Kenya
453 Okinawa II
GR 374 Morocco II-iii
GR 173 Ethiopia
GR 138 Uganda
hozon 35 Okinawa
185 Kenya II-iv
K 59104 Kenya
GR 134B Uganda
GR 133B Uganda
GR 131 Uganda II-v
GR 218 Unknown
GR 233A Kenya
GR 144 Kenya
GR 434 South Africa
GR 45 Unknown
GR 377 South Africa III
72-192 Kenya
GR 238 Kenya
GR 367 South Africa
GR 371 South Africa
GR 470B South Africa
73-919B Kenya
GR 458 South Africa
GR 422-1 America
GR 464 Ghana
GR 208 Kenya
GR 376 Unknown
GR 336 South Africa IV
GR 440 Zimbabwe
K 6343 Kenya
GR 206 Tanzania
GR 37 ‘Zambezi River’
GR 124 Kenya
GR 123 Kenya
GR 329 South Africa
GR 299 Tanzania
CP 1210-o-8 Unknown
72-62 Nigeria
GR 50 Unknown V
Monbasa Unknown
GR 239A Kenya
GR 101A Nigeria
GR 334 South Africa
GR 250 Kenya
GR 120 Ethiopia
GR 214 Kenya
GR 215 Kenya
GR 441 Australia
GR 432-1 South Africa
GR 470A South Africa
Natsuyutaka Unknown
GR 452 Unknown
K 6348 Kenya
GR 186 Kenya
VI
Natsukaze Cultivar
GR 453 Unknown
GR 456 Unknown
M70-82 South Africa
GR 467 Australia
Gatton Australia
GR 88 Australia
GR 365 South Africa
M70-81 South Africa
Meidai 9 Unknown

Fig. 11.6 Phenogram of the 77 accessions of guineagrass based on the UPGMA clustering of 190 alleles obtained with 13 SSR
markers (Ebina et al. 2007)
11 Panicum 189

PC2
cv. Zambezi River
64, K
14, 15, K 8
26, T
41, S
cv. Makueni

36, K 12, N
10 2.0
40, S
33, K
75 Group IV
cv. Tanzania

Group V
48, S

52, Z
1.0 1, N
47 38, T
61, S
27, K 49
57, S 39, S
53
Group III 6, K
29, K
44, S
–2.0 –1.0 9 50, S
13, E 28, K
58, G
51, S
PC1
17,U 2, K 32, K 62 1.0 2.0 3.0
20,K 31, K 54
21 65,K
4, K
45,E 43,S
5, K 16, U 7, K
69, K 55
46, M 35, K 71 23, K 76
22, E 56 67,S
24, K
77 60, S cv. Natsukaze 66, S
18,U 37, K 30 cv. Paikaji –1.0 11,74
72 34, K cv. Natsuyutaka 73
cv. Gatton 42, S
3, K
bl. Hozon35
19, U
70, K
25, K Group I Group VI 59 bl. Meidai9
68, K
63, K
bl. Ryukyu-3
–2.0
Group II

Fig. 11.7 Principal component analysis (PCA) scatter-plot of indicate group I: open circle, group II: open triangle, group III:
the 77 accessions of guineagrass based on similarity matrix of open square, group IV: closed circle, group V: closed triangle,
Jaccard’s coefficient. PC: principle component. The doted cir- and group VI: closed square (Ebina et al. 2007)
cles indicate different groups showing as Fig. 11.1. The symbols

the apomixis in guineagrass is simple and has been (Ozias-Akins et al. 1998; Akiyama et al. 2004; Goel
ascribed to a single gene (Hanna et al. 1973; Savidan et al. 2006).
1980; Nakajima 1991; Ebina and Nakagawa 2001). In In contrast, attempts to transfer apomixis from
addition to guineagrass, apomixes in other grasses Tripsacum to maize have proven difficult. Some of
follow single gene transmission, thus raising the future these difficulties have been ascribed to the male
possibility of introducing apomixis into new species or female sterility and to epigenetic patterning that
following recombinant gene transfer. The develop- impose constraints for apomictic seed development
ment of highly efficient gene transfer protocol has (Kindiger et al. 1996; Leblanc et al. 2008). These
been reported for P. virgatum following Agrobacter- observations are further complicated by evidence sug-
ium tumefaciens-mediated transformation (Somileva gesting that the regulation of apomixis in Gama grass
2006). The effectiveness of DNA conveyance has is polygenic. Blakey et al. (1997, 2001) have shown
been proven to be a function of genotype and choice that RFLPs linked to multiple chromosomal regions
of explants. cosegregate with apomixis. Under this scenario, all of
Given the simplicity of apomictic inheritance, the regions encoding genes necessary for apomixis
attempts have been made to introduce this gene into would have to be transferred simultaneously in order
compatible sexually reproducing lines by breeding. for this trait to be expressed. To date, this transfer
In pearl millet, sexual species of Pennisetum glaucum is conspicuously absent. No apomictic maize has
have been successfully introgressed with aposporous ever been recovered from breeding programs designed
apomixis derived from P. squamulatum (Dujardin to produce maize-Tripsacum hybrids. Apomictics
and Hanna 1983; Kaushal et al. 2007). The detailed derived following mutation in sexual diploid Tripsa-
molecular mapping of ASGR, in addition to studies cum population has also never been observed.
on chromosome structure and the physical chro- The ecological significance of variability among
mosome relationships among the agamic complex apomictically reproducing species with respect to pop-
of Pennisetum and Cenchrus have been reported ulation size and the number of genes regulating
190 H.S. Bhandari et al.

apomixis has been discussed by Adolfsson and QTLs defining, for example, yield, abiotic tolerance,
Bengtsson (2007). In this connection, guineagrass etc. will directly lead to the breeding of high perfor-
has been found to express a large number of variants mance sexual lines and the production of new, value-
in nature. Seven percent of the accessions are sexual added cultivars of the guineagrass.
and it is these that create variability by generating Recurrent sexual selection pools were obtained fol-
segregating populations or alternatively as a function lowing hybridization between the doubled sexual line
of facultative apomixis (Savidan and Pernes 1982). “Noh PL1” and apomixis cultivar “Natsuyutaka” (Sato
Facultative apomixis has inherent difficulties in gui- et al. 1992; Kouki and Ebina 2007). In more than 100
neagrass due to self-incompatibility. However, F1 progeny, apomixis segregated for sex in an 1:1
between 1.3% and 4.7% off-type plants arose from ratio. Polyembryony was notably absent in the sexual
facultative apomixis (Warmke 1954). A similar pat- progeny as determined by embryo sac analysis. At
tern is observed in Brachiaria in which both sexual least three of the F2 plants of each open-pollinated F1
and apomictic plants have been found (Miles et al. plants were planted and scored for agronomic traits in
2004). Guineagrass along with Eriochloa and Uro- addition to whether they reproduced sexually or by
chloa belong to a group of closely related plants apomixis. In F2 population, 41 superior plants were
whose boundaries are poorly defined (Renvoize et al. identified with respect to agronomic superiority and,
1996). Nevertheless, it is possible that the common of these, seven plants were sexual. Three plants were
ancestor plant to each of these related genera is likely also found that are perennial. As expected, all F2
one of the original aposporous apomixis plants among sexual plants exhibit monoembryony, with polyem-
this large group of Gramineae. bryony restricted to apomicts. The selected sexual
In addition to the evaluation of a large number of plants exhibit high yield and digestibility in compari-
variable apomixis accessions, diploid and teteraploid son with several apomictic cultivars. For useful appli-
sexual lines have also been employed in breeding cation of recurrent selection, additional agronomically
programs designed to elucidate the mechanism of superior sexual plants are necessary.
sexuality (Muir and Jank 2004). Due to inherent hetero-
zyogosity associated with cross-pollination and poly-
ploidy, recurrent sexual selection will be a new
powerful instrument for any guineagrass breeding pro- 11.12 Genomics
gram (Vogel and Bruson 2004). Advanced molecular
tools, for example, those widely available apomixis Switchgrass cultivars and “prairie-remnant” popula-
markers, should make it easy to construct and select tions were analyzed with RAPD markers (Casler
highly pure recurrent sexual pools. While the region et al. 2007). The structural patterns and spatial varia-
linked to the apomixis gene is extremely large, tion suggested that there are different switchgrass gene
the AFLP markers obtained from “Natsukaze” have pools in different regions of the northern and eastern
not been tested with other cultivars and populations US. Wider within-population variation was reported
(Akiyama et al. 2008). While the common region in comparison to those among-population variation.
among the apomixis accessions remains available Thus, a relatively small number of collection sites
to discriminate asexual accessions, the cytological are considered sufficient to maintain genetic variabil-
embryo sac clearing method remains useful, for the ity within the gene pool. A collection of 31 unique
selection of recurrent sexual pool. This said, more switchgrass accessions from 168 available collections
simple diagnostic molecular methods of the pure in GRIN were analyzed with expressed sequence
sexual lines require enhancement. The prospect of tag–simple sequence repeat (EST-SSR) markers
accelerating recurrent sexual selection may be made (Narasimhamoorthy et al. 2008). Greater within-pop-
through application of genotype matrix mapping ulation (79.6%) rather than among-population (20.4%)
(GMM) selection (Isobe et al. 2007). While autotetra- variation was reported, which appears to be a common
ploidy and high heterogeneity in guineagrass have feature in many outcrossing species. The results sug-
been prevented in conventional breeding, GMM gested that the recurrent selection within a population
seeks quantitative trait loci (QTLs) and specifically would serve as breeding strategy for switchgrass. The
QTL interactions in genotype pools. The detection of genotypes were also broadly grouped into different
11 Panicum 191

adaptive zones based on the geographical locations of 11.13 Invasive Species

collections.
Recently, cDNA libraries have been constructed
Several species of Panicum such as P. capillaire,
from callus, crown, and seedling tissues of P. virga-
P. dichotomoflorum, P. repens, and P. virgatum are
tum, cultivar “Kanlow” (Tobias et al. 2008). The
notoriously invasive. For example, P. repens is a
libraries were end-sequenced. An excess of 61,500
perennial that forms large dense colonies. Concomi-
high quality ESTs were produced. The consensus
tant with this configuration long, creeping rhizomes
sequences were identified and 73% could be aligned
characterize this structure and, as a result, long floating
with Sorghum bicolor, the vast majority of these being
mats are frequently observed. This precise organiza-
identified with gene ontology. Given the scope of the
tion can result in the failure of water to flow properly,
library, it is likely that the information gleaned from
making the control and management of plant spread
scoring sequence diversity and similarity among wild
difficult. Cutting-back is ineffective; only the applica-
Panicum species could provide insight on important
tion of herbicides can ameliorate the uncontrolled
biosynthetic functionalities as cell wall biosynthesis.
proliferation and growth. This said, it is precisely the
Public EST database for guineagrass have
formation of this habit that can be used to organize
recently become available on the DDBJ at accession
effective soil erosion strategies (www.heap.org./pier/
DB999756–DB999951. These 196 ESTs are genes
species/panicum_repens.html). The lesson here is that
that were expressed in immature apomixis pistils.
the use and focused application determine whether a
These are surveyed by microarray from 4,608 apomic-
plant is an invader or a friend.
tic spikes expressed genes (Yamada-Akiyama et al.
2009). Candidates for the apomixis initiation genes
were selected from the 196 ESTs. Quantitative PCR
was performed with selected primers by using mRNA
isolated from only a few immature pistils in different
11.14 Economic Advantages
developmental stages Fig. 11.8. Due to the fact that
some guineagrass lines are facultative apomicts, the Given this, many of these species are also used as
small number of pistils screened is particularly sensi- forage for livestock. Species such as P. coloratum
tive with respect to the detection of genes involved in and P. miliaceum have been cultivated to be used as
the expression of apomixis initiation. One novel gene fodder, while P. coloratum, P. repens, and P. virgatum
homologous to a gene for immature panicle in Sor- are important constituents of native pasture (Watson
ghum bicolor exhibits higher expression in apomictic and Dallwitz 1994). P. miliaceum, P. sumatrenes, and
immature pistils of guineagrass. Another gene homol- P. sonorum are important cultivated species for food
ogous to the apomictic Cenchrus ciliaris pistil-specific for human consumption (Hitchcock 1971; Watson and
gene also demonstrated higher expression in apomictic Dallwitz 1994). In countries such as Asia, Europe, and
immature pistils of guineagrass. the Middle East, dehulled grain of P. miliaceum (proso

Fig. 11.8 The ovaries from early and middle immature panicles of P. maximum. Each five panicles were subjected to the Q-PCR
analysis. The scale bar indicates 1 mm
192 H.S. Bhandari et al.

millet) can be directly cooked like rice or ground to introduced to the Japanese semitropical islands of Oki-
make porridge. The grains are also used as birdfeed. In nawa prefecture, and new hybrid cultivars continue to
US, this crop is also used as forage. The species can be released.
thrive well under severe heat stress, poor soils, and
drought.
P. virgatum L. (switchgrass), is a native warm
11.15 Recommendations for Future
season grass of North American tallgrass prairies and
has received widespread attention due to its potential Resource Utilization
for use as feedstock for cellulose-based bioethanol
production (Sanderson et al. 1996; Vogel and Jung Several major global problems still to be faced relate
2001; McLaughlin and Kszos 2005; Bouton 2007, directly to the inequitable distribution of natural
2008). Traditionally, this crop was used primarily resources, pollution, and biodiversity [for review see,
as livestock forage, restoration of range land, or as a Mahmud et al. (2008) and Rai (2008)]. China has more
component of conservation reserve program (Bouton than 20% of the world’s population but only seven
2007). Earlier work on genetic modification of this percent of the planet’s arable land. This country, in
crop was focused mainly on the improvement of its addition to others in Asia, faces serious threats in their
forage yield and quality (Vogel and Jung 2001). After food chain as a consequence of heavy metal contami-
this crop was chosen as a main herbaceous crop for nation of the region’s major food source, specifically
production of bioethanol, the research has been shifted rice. This problem is likely to become acerbated with
to improve the biomass production and quality for time. India, with the world’s second largest area of
the conversion efficiency (Parrish and Fike 2005). tillable land, no longer exports rice but imports wheat,
Research is now underway on modification of bio- while Ganges River carries arsenic contaminants into
chemical pathways to impact the recalcitrance and Bangladesh. Not only are the land resources notori-
enhance ethanol yield (Zeng-Yu Wang, pers. comm.). ously unbalanced with respect to utilization but also is
The discovery of sexual plants of P. maximum at potable water. In fact, 20% of the Earth’s fresh water is
Tifton led to the development of winter-hardy clones to be found in the US Great Lakes System.
(Hanna et al. 1973; Hanna 1986). The progeny of the Complicating this picture further is the fact that,
same sexual lines generated new cultivars. The off- congruent with high selection pressure, plants native
spring of these lines also generated excellent succes- to these regions have evolved and reproduce success-
sions, and new cultivars were bred in several countries fully under conditions of abiotic challenges. Ferns are
of South America and Japan (Nakagawa and Hanna capable of taking up high concentrations of arsenic
1992; Muir and Jank 2004). A French collection was known as are rice lines that quite successfully package
studied in the Ivory Coast, and several accessions, such this poison in the grain, thus allowing contaminated
as “T58”, “K187,” and “C1,” were evaluated based on cereals into the food chain. Then too, there are plants
agronomic traits and subsequently distributed to some based on folklore, which are believed to have some
additional African countries (Mandret et al. 1989). This pharmaceutical properties based on empirical obser-
collection, including the sexual materials, was trans- vation. The putative treatment has been identified to a
ferred to Brazil EMBRAPA, where a robust breeding tissue or organ and anything from poultices to teas
program ensued. Three new cultivars were released, have been effectively used from one generation to
namely “Tanzania,” “Monbasa,” and “Massai” (Muir the next.
and Jank 2004). Another sexual diploid “GR 297” was While the distribution of land and water cannot be
discovered and first apomictic hybrid cultivar “Natsu- re-equilibrated, it is a certainty that the value of native
kaze” generated from crosses between it and an uniden- germplasm should at least, in portion, return to the
tified tetraploid apomictic line through open-pollination country from which it derives. To address this poten-
(Nakajima et al. 1979; Sato et al. 1993). Since “Natsu- tial source of exploitation, the United Nations Confer-
kaze” is an apomictic tetraploid, this strain is thought to ence on Environment and Development (UNED)
have originated from an unreduced egg of a diploid made available for signature The Convention on
sexual plant or a hexaploid apomictic pollen donor. Diversity. The Biodiversity Treaty, as it is commonly
Japan’s Panicum breeding program has been newly known, has been signed by an excess of 170 countries.
11 Panicum 193

One of its primary purposes calls for the “fair and Bouton JH (2007) Molecular breeding of switchgrass for use as
equitable sharing of the benefits arising out of the a biofuel crop. Curr Opin Genet Dev 17:553–558
Bouton JH (2008) Improvement of switchgrass as a bioenergy
utilization of genetic resources.” In essence, it calls crop. In: Vermerris W (ed) Genetic improvement of bioe-
for the recognition and subsequent protection of prop- nergy crops. Springer, Heidelberg, Germany, pp 295–308
erty rights devolving from biological and genetic Bouton JH, Brown RH, Evans PT, Jernstedt JA (1986) Photo-
assets. Interestingly, a number of countries home to a synthesis, leaf anatomy, and morphology of progeny from
hybrids between C-3 and C-3/C-4 Panicum species. Plant
huge biotechnology industry, including the US, have Physiol 80:487–492
not signed this treaty. Thus, issues of equity and fair- Brown RH, Morgan JA (1980) Photosynthesis of grass species
ness arise prominently where countries lacking the differing in carbon di-oxide fixation pathways 6. differential
infrastructure cannot even exploit its genetic resources. effects of temperature and light intensity on photo respira-
tion in 3 carbon 4 carbon pathway and intermediate species.
This has the potential of allowing well funded enter- Plant Physiol 66:541–544
prises from engaging in the worse kind of gene explo- Brown RH, Bouton JH, Evans PT, Malter HE, Rigsby LL (1985)
ration/abuse. For example, a gene that allows for Photosynthesis, morphology, leaf anatomy, and cytogenetics
arsenic packaging in the aerial parts of plant could be of hybrids between C-3 and C-3/C-4 Panicum species. Plant
Physiol 77:653–658
isolated, cloned by a foreign enterprise, introduced in Burton G (1942) A cytological study of some species in the tribe
rice, and sold back, for example, to Bangladesh, the Paniceae. Am J Bot 29:355–359
very source of the plant material from which it is Carman JG (1997) Asynchronous expression of duplicate genes
derived. It is to the correction of these manipulations in angiosperms may cause apomixis, bispory, tetraspory, and
polyembryony. Biol J Linn Soc 61:51–94
that principled scientists should address themselves, Carneiro VTC, Dusi DMA, Orits JPA (2006) Apomixis: occur-
speaking for those not in a position to speak for rence, applications and improvements. In: Teixeira da Silva
themselves. JA (ed) Floriculture, ornamental and plant biotechnology.
Global Science Books, Middlesex, pp 564–571
Casler MD, Stendal CA, Kapich L, Vogel KP (2007) Genetic
diversity, plant adaptation regions, and gene pools for
switchgrass. Crop Sci 47:2261–2273
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Chapter 12
Paspalum

W.M. Williams, M.L. Williamson, and D. Real

12.1 Introduction chloroplast DNA sequence (Duvall et al. 2001) and

morphology (Clayton and Renvoize 1986) and subse-
quently was absorbed into Paspalum as subgenus Har-
Paspalum L. (tribe Paniceae R. Br.) is one of the
postachys (Denham 2005).
largest C4 grass genera with approximately 400 spe-
To date, there have been very few plant breeding
cies that are native to predominantly tropical and sub-
programs. There is considerable interest in evaluating
tropical regions (Clayton and Renvoize 1986; Watson
Paspalum species in Brazil because there are about
and Dallwitz 1992), with the center of diversity in
220 species native to that country (Valls and Pozzobon
South America (Moraes Fernandes et al. 1968). The
1987). Workers are researching on a range of native
C4 decarboxylation is catalyzed by NADP malic
material with a view toward variety development and
enzyme (NADP-ME), the most common type in the
interspecific hybridization (Pagliarini et al. 2001).
Paniceae. There are a few semi-domesticated species.
Several germplasm collecting trips have been under-
P. dilatatum Poir. (dallisgrass), P. pauciciliatum
taken in the last 30 years, resulting in a collection of
(Parodi) Herter (prostrate dallisgrass), P. urvillei
over 1,500 accessions at CENARGEN/EMBRAPA by
Steud. (Vasey grass, Venuto et al. 2003), P. atratum
the early 1990s (Takayama et al. 1998; Adamowski
Swallen (Atra paspalum, Kretschmer et al. 1994),
et al. 2005). Part of this collection was duplicated at
P. glaucescens Hackel, P. guenoarum Arechav.
CPPSE/EMBRAPA and at IBONE at Corrientes,
(pasto rojo), P. nicorae Parodi (Brunswick grass),
Argentina.
and P. plicatulum Michx. (brownseed paspalum)
The predominant chromosome number is tetraploid
have been commercialized for forage production,
(2n ¼ 4x ¼ 40), but diploids are also common
while P. notatum Fl€ ugge (bahiagrass ) is used for
(Pozzobon et al. 2008). Almost all species have a
both forage and turf (Bennett 1962). P. vaginatum
basic chromosome number of x ¼ 10. However, as
Swartz (seashore paspalum) is used for dune stabiliza-
new collections are analyzed, the number of excep-
tion and turf on saline soils while P. distichum
tions has progressively increased (Penaloza et al.
L. (knotgrass) is used in wet places as a feed for
2008). Those known to date are listed in Table 12.1.
water birds. P. scrobiculatum L. (kodo millet) is used
It has been suggested that, based on secondary chro-
in Asia as a cereal grain. Many other species are used
mosome pairing observations in P. notatum, the basic
locally in native grasslands for forage and feed (Allem
number of x ¼ 10 is derived from a lower ancestral
and Valls 1987; Filgueiras 1992). The genus Thrasya
number (x ¼ 5 or 6, Forbes and Burton 1961).
Kunth (20 species) is very close to Paspalum in
Many species have multiple cytotypes (Table 12.2),
with the diploids reproducing sexually and the poly-
ploids apomictically (Quarin and Norrmann 1990).
Examples include P. dilatatum (2n ¼ 40, 50, 60;
W.M. Williams (*)
Burson 1991b) and P. notatum, P. thunbergii Kunth
AgResearch, Grasslands Research Centre, Private Bag 11008,
Palmerston North, New Zealand ex. Steud., and P. plicatulum (2n ¼ 20, 40; Darlington
e-mail: [email protected] and Wylie 1961). The mean diploid 2C DNA value is

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 197
DOI 10.1007/978-3-642-14255-0_12, # Springer-Verlag Berlin Heidelberg 2011
198 W.M. Williams et al.

Table 12.1 Paspalum species with basic chromosome numbers different from x ¼ 10
Species Group Chromosomes References
P. almum Alma 2n ¼ 12, 24 Quarin and Hanna (1980)
P. filgueirasii 2n ¼ 24 Penaloza et al. (2008)
P. convexum Plicatula 2n ¼ 32 Selva (1976), Reeder (1984)
P. lanciflorum Ceresia n¼9 Davidse and Pohl (1974)
P. stellatum Ceresia 2n ¼ 52 Honfi et al. (1990)
P. reduncum Gardneriana 2n ¼ 18 Penaloza et al. (2008)
P. ammodes Eriantha 2n ¼ 36 Penaloza et al. (2008)
P. trachycoleon Ceresia 2n ¼ 36 Penaloza et al. (2008)
P. burmanii Ceresia 2n ¼ 48 Penaloza et al. (2008)

1.6 pg and the species variation is approximately groups are not always phylogenetically informative
threefold as determined by laser flow cytometry (Souza-Chies et al. 2006).
(Watson and Dallwitz 1992). Based on morphological classification, Paspalum
Because many of the apomictic species require species have been classified into four subgenera Ana-
fertile pollen for apomixis to succeed (pseudogamy), chyris (6 species), Ceresia (approximately 25 species),
they are able to be used as male parents in hybridi- Harpostachys (39 species), and Paspalum (more than
zation. Consequently, hybridization programs have 250 species; Chase 1929; Zuloaga and Morrone 2005;
involved crosses between diploids and crosses of poly- Aliscioni and Denham 2008). Subgenus Paspalum is,
ploids with diploids. Before more targeted breeding in turn, divided into informal groups, e.g., Notata with
programs can commence, a much better understanding 21 species (Zuloaga et al. 2004; Table 12.2).
of species phylogenies is needed. Because of apo- Phylogenetic relationships were initially analyzed
mixis, it has been possible for species hybrids and using chromosome pairing in interspecific hybrids.
even intergeneric hybrids to become established This led to the identification of several distinct geno-
(Quarin 1992). mes that were annotated (Burson et al. 1991). The
occurrence of polyploidy and apomixis makes chro-
mosome analysis of phylogeny very difficult and also
hinders conventional breeding (Jarret et al. 1998).
12.2 Taxonomy Very few species have undergone meiotic analyses.
In addition to chromosome pairing abnormalities,
The systematic treatment of the genus is complex with Pagliarini et al. (2001) observed non-oriented bivalents
a large amount of interspecific hybridization and poly- at metaphase I of meiosis and early chiasma termina-
ploidy, and many taxonomic groupings are still to be lization, resulting in univalents and micronuclei at
fully resolved. Indeed, the taxonomy is complicated by anaphase I. The latter abnormalities occurred even
the presence of non-sexual polyploid complexes with when chromosome pairing appeared to be completely
sexual diploid and partially sexual tetraploid popula- normal. To date, sexual forms that have been studied
tions contributing new genetic diversity via hybridi- have normal meiosis. The meiotic abnormalities are
zation. Under these circumstances, the concept of therefore largely a characteristic of apomictic plants
“species” is difficult because continued interspecific (Burson and Bennett 1971a; Pagliarini et al. 2001).
hybridization, with the hybrids fixed by apomixis, cre-
ates continuous intermixing and intergrading of taxa,
resulting in agamic complexes (Pozzobon et al. 2008).
Various authors have used external morphological 12.3 Sexual Species of Paspalum
characteristics to subdivide the genus into subgenera
or sections and informal species groups (e.g., Chase Because of the high frequency of apomictic species
1929). However, these classifications have been and biotypes in Paspalum, the sexual species (diploids
repeatedly revised by various researchers and no sin- and facultative apomicts) represent an important
gle classification has been reached and the informal genetic resource as generators of genetic diversity for
12 Paspalum 199

Table 12.2 Approximate species numbers in subgenera, sec- plant breeding. To date, little more than one quarter
tions, and informal groups of the genus Paspalum of the genus has been characterized cytologically.
Subgenera Section or informal Number of Cultivated species
group speciesa Some of the diploid sexual species and biotypes are
Anachyris 6 summarized in Table 12.3. It is clear that sexual taxa
Ceresia Ceresia 19 are numerous and occur in almost all subgenera and
Pectinata 8
Harpostachys 39
species groups, so that potentially useful genetic
Paspalum Dilatata 4 P. dilatatum, resources are available throughout the genus.
P. urvillei, There have been a few molecular analyses of phy-
P. pauciciliatum
Livida 13 logeny of the Paspalum species. Jarret et al. (1998)
Paniculata 8 used restriction fragment length polymorphisms
Quadrifaria 10 (RFLPs) to analyze 51 accessions of 29 species mainly
Virgata 10
Notata 21 P. notatum
from South America but including one from Japan
Alma/Notata 1 (P. thunbergii Steud.) and P. scrobiculatum from India.
Ovalia 1 Species relationships generally supported those obtained
Alterniflora 4
Barbinodia 2
from morphological and cytological considerations.
Bertoniana 2
Bifida 1
Bonplandiana 6
Brevia 2 12.4 Paspalum as a Model for Apomixis
Caespitosa 17
Conjugata 1
Corcovadensia 8
Paspalum is potentially a very useful genus to research
Decumbentes 4
Dimorphostachys 1 the genetics of apomixis because of the existence of
Dissecta 7 sexual diploids, apomictic polyploids, and facultative
Disticha 4 P. distichum,
P. vaginatum
apomictic tetraploids. The genetics of apomixis in tet-
Eriantha 6 raploid P. notatum was investigated by Martinez et al.
Falcata 1 (2001) by pollinating an emasculated fully sexual
Fasciculata 1
mother plant with apomictic plants and verifying
Filiformia 5
Fimbriata 3 hybrids by RFLP analysis. F2 and backcross families
Floridana 2 were produced from both sexual and apomictic F1
Gardneriana 9
plants. The F1 produced 157 sexual plants and 56 plants
Inaequivalvia 2
Lachnea 2 that were apomictic (many of these also showed mei-
Laevia 6 otic embryo sacs), a ratio close to 3:1. Sexual F1 
Macrophylla 2 apomictic F1 and backcrosses of sexual  apomictic
Maculosa 1
Orbiculata 6 plants also produced approximate 3:1 ratios. The
Palustrine 2 results were interpreted to be consistent with deter-
Parviflora 8 mination of apospory by tetrasomic inheritance of a
Plicatula 21 P. atratum,
P. guenoarum, dominant allele at one locus, as is likely in other tropi-
P. nicorae, cal grasses. Failure to obtain a near 1:1 ratio in the F1
P. plicatulum
Racemosa 10
was attributed to segregation distortion against apos-
Recta 4 pory, possibly due to a pleiotropic lethal effect of an
Reimaria 1 allele with incomplete penetrance or to closely linked
Rupestria 3
Saccharoidea 4
genes with partial lethality. An earlier analysis (Burton
Setacea 3 and Forbes Jr 1960; Forbes and Burton 1961), conclud-
Ungrouped 34 ing that apospory was controlled by a few recessive
a
Chase (1939), Renvoize (1972), Cialdella et al. (1995), genes, was rejected on technical grounds (Martinez
Morrone et al. (1995, 1996, 2000, 2004), Denham et al. et al. 2001). Subsequently, a number of molecular
(2002), Zuloaga et al. (2004), Denham (2005), Zuloaga and
Morrone (2005), Aliscioni and Denham (2008), Hojsgaard
markers completely linked to the apospory locus
et al. (2008) have been found and analyzed (Martinez et al. 2003;
200 W.M. Williams et al.

Table 12.3 List of some species with diploid sexual biotypes. Table 12.3 (continued)
As indicated, many of these also have polyploid (apomictic) Subgenus or Species Chromosome
biotypes Group numbera
Subgenus or Species Chromosome P. paniculatum 20 (also 40)
Group numbera P. palustre 20
Subgenus P. malachophyllum 20 (most 40, P. umbrosum 20
Anachyris some 60) Recta P. rectum 20
P. procurrens 20 (also 40) P. vescum 20
P. simplex 20 (also 40, 60) Parviflora P. scalare 20
Subgenus Ceresia P. carinatum 20 (also 80) P. clavuliferum 20
P. cordatum 20 P. multicaule 20 (also 40)
P. humboldtianum 20 Disticha P. vaginatum 20 (also 40, 60)
P. lanciflorum 18 Corcovadensia P. corcovadense 20
(P. contractum) Quadrifaria P. quarinii 20 (also 40)
P. malmeanum 20
P. haumanii 20
P. pectinatum 20 (also 40, 60)
P. brunneum 20
Subgenus P. decumbens 20
P. intermedium 20 (also 40)
Harpostachys P. cinerascens 20
P. quadrifarium 20 (also 30, 40)
Subgenus
Bertoniana P. bertonii 20
Paspalum
Notata P. notatum var. saurae 20 Racemosa P. candidum 20 (also 40, 60)
P. approximatum 20 P. prostratum 20, (also 40, 60)
P. barretoi 20 Conjugata P. conjugatum 20 (also 40, 80)
P. cromyorhizon 20 (also 40) Fasciculata P. fasciculatum 20
P. nummularium 20 Orbiculata P. orbiculatum 20
P. minus 20 (also 40, 50) Livida P. plenum 20 (also 40)
P. pumilum 20 P. trichophyllum 20
Notata/ P. maculosum 20 (also 40) Dissecta P. repens 20
Maculosa Virgata P. rufum 20 (also 40)
Notata/Alma P. almum 12 (also 24) Gardneriana P. reduncum 18
a
Linearia/Notata P. crispulum 20 Main sources were Zuloaga and Morrone (2005) and Pozzobon
P. dedeccae 20 (also 40) et al. (2008)
P. ellipticum 20 (also 80)
P. filifolium 20
P. lineare 20 (also 40, 60)
P. pallens 20 Stein et al. 2004), suggesting that the locus is in a
Fasciculata/ P. equitans 20 region of suppressed recombination. The latter study
Notata obtained evidence that the apospory locus was asso-
Plicatula P. glaucescens 20 (also 40, 60)
ciated with a chromosome region that showed disomic
P. limbatum 20 (also 40)
P. guenoarum 20 (also 40)
inheritance (while the rest of the genome was largely
P. hydrophilum 20 (also 30, 40) tetrasomic). The possibility of a chromosome rear-
P. chaseanum 20 rangement (e.g., an inversion) being associated with
P. compressifolium 20 (also 40, 60) the segregation distortion as well as the suppressed
P. plicatulum 20 (also 40) recombination was discussed.
P. wrightii 20 (also 30, 40) Repression of recombination in the chromosome
Modesta/ P. modestum 20 region controlling apospory, parthenogenesis, and
Plicatula
Caespitosa/ P. arenarium 20 pseudogamy was also observed in studies of P. sim-
Setacea plex Morong (subgenus Anachyris) by Pupilli et al.
Setacea P. setaceum 20 (1997, 2001) and Caceres et al. (2001), and the region
Falcata/ P. falcatum 20 showed strong colinearity with a rice chromosome
Lachnea
Caespitosa P. indecorum 20
segment.
P. chacoense 20 P. notatum and P. simplex were used by Martinez
Paniculata P. juergensii 20 et al. (2007) to investigate the question as to why
(continued) apospory is associated with polyploidy. This work
12 Paspalum 201

showed that apomixis was not transmitted by mono- Because almost all apomictic species are polyploid
ploid gametes, explaining why hybridization does not (predominantly tetraploid), it has been postulated that
produce apomictic diploids. Gametes higher than apomixis evolved as a mechanism by which sexually
monoploid transmitted apospory but with apparently infertile wide hybrids have been rescued and restored
strong segregation distortion. Despite this, a low num- to fertility (Darlington 1939). There is evidence that 5x
ber of extra aposporic embryo sacs (always associated P. dilatatum arose from interspecific hybridization of
with normal meiotic embryo sacs) were found in sev- sexual species (Burson 1989). However, many apo-
eral diploid species by Norrmann et al. (1989). The mictic grasses appear to be autopolyploids (Quarin
species were P. intermedium Monro ex. Morong & 1992). In Paspalum, there are many species that have
Britton P. quadrifarium Lamk., P. haumanii Parodi, sexual diploid and apomictic tetraploid biotypes that
P. brunneum Mez, and P. rufum Nees ex. Steud. These are morphologically identical, suggesting that autotet-
observations, along with observations of apospory in raploidy is perhaps more likely than allotetraploidy.
P. cromyorhizon Trin. ex. Doell (Quarin 1986) and An experiment was carried out by Quarin et al. (1998)
P. equitans Mez (Quarin and Norrmann 1987) and the in which resynthesis of the tetraploid form of P. rufum
finding of facultative apospory in colchicine-doubled (Virgata group) from the diploid form strongly indi-
P. hexastachyum Parodi (Quarin and Hanna 1980), indi- cated an autopolyploid origin of the wild biotype. It is
cate that the potential for apomixis exists in some of the apparent that apomixis, while predominantly related to
sexual diploid species. Apparently, the expression of this polyploidy, is not confined to allo- or auto-polyploids,
potential can vary with environmental conditions or sea- and that some explanation other than rescue of hybrids
son (Quarin 1986; Norrmann et al. 1989). is needed for the preponderance of polyploidy in apo-
The molecular genetic basis of apomixis in mictic populations.
P. notatum was investigated by Pessino et al. (2001). Although apospory is by far the main type of apo-
A family segregating for sexual and apomictic plants mixis in Paspalum, diplospory has occasionally been
was generated by pollinating a sexual tetraploid with found. P. minus E. Fourn., a widely spread species
an apomictic tetraploid. Spikelet mRNA was extracted with 2x, 4x, and 5x races, has been shown to have
from three sexual and three apomictic progenies, and both types of apomixis in the 5x form (Bonilla and
the two lots were compared in differential display Quarin 1997).
experiments. Three transcripts that were expressed at
higher levels in the apomictic plants were cloned and
proved to show high homology. However, they
showed no homology with genes of known function, 12.5 Subgenus Anachyris Chase
although a probable homology was detected with a
KSP multiphosphorylation domain that is a phosphor- Subgenus Anachyris consists of six species (Morrone
ylation site for cdc-2-kinases that are known to be et al. 2000) distributed from Mexico to Argentina and
associated with cell division. Uruguay, characterized by boat-shaped spikelets.
A genetic linkage map of diploid P. notatum var. Three of these species (P. simplex, P. procurrens
saurae (Pensacola bahiagrass) was developed by Ortiz Quarin, and P. malachophyllum Trin.) have sexual
et al. (2001) using a mapping population from a cross diploid and facultative apomictic polyploid cytotypes
between parents from Cayasta, Argentina, and Tifton, (Table 12.3), while two (P. usterii Hack. and P. vol-
Georgia. RFLP clones from maize, rice, and oat were canensis Zuloaga, Morrone & Denham) are known
used for the framework, while random amplified poly- only as tetraploids (Hunziker et al. 1998; Hojsgaard
morphic DNA (RAPD) and amplified fragment length et al. 2008). P. simplex has been used in studies of the
polymorphism (AFLP) markers provided refinement. genetics of apomixis, as discussed above. Tetraploid
Ten linkage groups were established, with 112 mar- forms of P. malachophyllum, P. procurrens, and
kers averaging about 9 cM distance. Several RFLP P. usterii show chromosome pairing behavior that is
markers associated with apomixis in Brachiaria and consistent with autotetraploidy (Hojsgaard et al.
maize–Tripsacum hybrids were located on the map, 2008). Tetraploid interspecific hybrids between a col-
opening the way for comparative genomics of apo- chicine-doubled plant of P. simplex  P. procurrens
mixis and many other traits. and  P. usterii and in triploid and diploid hybrids
202 W.M. Williams et al.

between P. simplex and P. malachophyllum indicate ated to determine the genomic relationships between
that these species share the same homologous gen- these two species (Burson 1981a). The hybrids were
omes. Hojsgaard et al. (2008) concluded that the entire diploid with irregular chromosome behavior, typically
subgenus is probably an agamic complex, sharing a having only two bivalents and 16 univalents at meio-
common genome and speciating at the diploid level sis. This result indicated that the P. juergensii and
before polyploidization, apomixis, and geographic P. intermedium genomes were largely unrelated and
separation, led to the diversity apparent among the that each genome probably paired with different
“species” now found. P. dilatatum ssp. flavescens genomes. Hybrids were
male and female sterile.
The sexual members of the group are P. urvillei,
12.6 Subgenus Paspalum P. dasypleurum, P. dilatatum ssp. flavescens, and bio-
types P. dilatatum “Virasoro” and P. dilatatum
12.6.1 Informal Group Dilatata Chase “Vacaria,” all sharing the “I” and “J” genomes. The
pentaploid and hexaploid cytotypes are P. dilatatum
“Common” and biotypes P. dilatatum “Chiru” and
This group, which includes the agronomically important “Uruguaiana,” which contain additional genome(s) of
P. dilatatum Poir. (dallisgrass), is usually considered to uncertain origin, designated X. The genomic constitu-
be closely related to the Livida, Paniculata, Quadrifaria, tions of these closely related groups are given by
and the Virgata groups, which, along with Dilatata, Speranza (2005) and are summarized in Table 12.4.
comprise the “Dilatata complex.” Because the Dilatata The close relationship of the Livida group with Dila-
are hybrid in origin, tracing their origin has been prob- tata is unclear and needs more attention. P. denticulatum
lematic, although studies of genome relationships and Trin. (syn. P. lividum Trin. ex. Schltdl.) has been
preliminary phylogenetic studies are predominantly in
agreement (Souza-Chies et al. 2006). Four species make
Table 12.4 Ploidy level and genomic formula for Dilatata,
up the Dilatata group: P. dilatatum, P. pauciciliatum, P. Quadrifaria, Virgata, and Paniculata groups
urvillei, and P. dasypleurum Kunze ex. Desv. These Species or biotype 2n Genomic Formula
species show similarities at both the morphological Dilatata Group
and molecular levels (Chase 1929; Jarret et al. 1998; P. dilatatum ssp. dilatatum 50 IIJJX
Vaio et al. 2005, 2007; Aliscioni and Denham 2008). P. dilatatum ssp. flavescens 40 IIJJ
The Dilatata group has sexual tetraploids and apo- P. dilatatum “Virasoro” 40 IIJJ
mictic pentaploid and hexaploid cytotypes. The tetra- P. dilatatum “Vacaria” 40 IIJJ
ploid forms are hypothesized to be hybrids between P. dilatatum “Chiru” 60 IIJJXX
P. dilatatum “Uruguaiana” 60 IIJJX1X2
P. intermedium (contributing the “I” genome) and
P. urvillei 40 IIJJ
P. juergensii Hackel (contributing the “J” genome) P. dasypleurum 40 IIJJ
from groups Quadrifaria and Paniculata, respectively
Quadrifaria Group
(Burson et al. 1973; Burson 1983; Espinoza and P. intermedium 20 II
Quarin 2000; Speranza 2005). Burson et al. (1973) P. haumanii 20 II
hybridized P. dilatatum ssp. flavescens (2n ¼ 40) P. brunneum 20 II
with P. intermedium (2n ¼ 20), P. juergensii (2n ¼ P. quadrifarium 20 II
20), P. notatum (2n ¼ 20, Notata), and P. vaginatum P. densum 20 II
(2n ¼ 20, Disticha) to determine their genomic rela- P. durifolium 60 IIJ2J2XaXa
tionships. P. dilatatum ssp. flavescens  P. interme- Virgata Group
dium and P. dilatatum ssp. flavescens  P. juergensii P. conspersum 40 I2I2JJ
P. virgatum 40 IIJ2J2
hybrids were triploid (2n ¼ 30) and showed irregular
P. rufum 20 II
chromosome pairing at meiosis with ten univalents
Paniculata Group
and ten bivalents, indicating that P. dilatatum shares
P. juergensii 20 JJ
a common genome with P. intermedium and with
P. paniculatum 20 JJ
P. juergensii. Three hybrids between P. juergensii a
Unknown genome not related to other “X” genomes. Table
(2n ¼ 20) and P. intermedium (2n ¼ 20) were gener- extracted from Speranza (2005)
12 Paspalum 203

hypothesized to be the source of a genome in 4x (Bashaw and Forbes 1958; Bashaw and Holt 1958a;
P. pauciciliatum (Valls 2000). P. dilatatum and P. livi- Venuto et al. 2003; Vaio et al. 2007). Garcia et al.
dum showed close phylogenetic similarities in DNA (2007) investigated the morphological and molecular
(ITS and chloroplast) sequences (Souza-Chies et al. (RAPDs) phylogeny of 4x and 5x P. dilatatum acces-
2006). In contrast, RFLP data showed a marked separa- sions. Molecular phylogeny could, to a certain
tion between P. lividum accessions and the Dilatata extent, be used to identify germplasm from different
group (Jarret et al. 1998), although P. pubiflorum origins but could not distinguish between tetraploid
Rupr. ex. Fourn. (Livida group) showed a stronger affin- and pentaploid accessions. Morphology could distin-
ity with P. dilatatum and its relatives than with guish between most tetraploid and pentaploid acces-
P. lividum. sions but could not distinguish between germplasm
Paspalum dilatatum ssp. dilatatum (5x) seems to from different origins. It is possible that different
have originated from a natural hybridization between genomes were involved in the origins of the different
the sexual tetraploid P. dilatatum ssp. flavescens, and populations.
the apomictic hexaploid P. dilatatum “Uruguaiana” “Common” dallisgrass: The “Common” biotype
(Machado et al. 2005). As the original carrier of the (P. dilatatum ssp. dilatatum) is a purple-anthered pen-
“X” genome, the pentaploid also seems to have been taploid (2n ¼ 50, IIJJX, 2C ¼ 2.96 pg) native to the
involved in the origin of the other apomictic species in Buenos Aires province of Argentina, Uruguay and S
these groups (Speranza 2005). Chromosomes of the Brazil. It reproduces apomictically (apospory and par-
pentaploid pair as 20 bivalents and 10 univalents at thenogenesis), but the apomictic mode of reproduction
meiosis (Bashaw and Holt 1958a; Burson 1983). is a barrier to genetic improvement (Bashaw and Holt
Using RAPDs, Miz and Souza-Chies (2006) per- 1958a; Caponio and Quarin 1990; Vaio et al. 2007). It
formed a phylogenetic analysis of the species of the is highly valued as permanent pasture in southeastern
Dilatata group, including seven P. dilatatum bio- USA due to its adaptability to different soil types,
types. Also included were the possible donors of ability to grow well with other grasses and legumes,
the “I” and “J” genomes (P. intermedium, P. juer- long growing season (earlier in spring and later in
gensii, and P. paniculatum L.). Apart from P. dilata- autumn than other warm season perennial grasses),
tum “Virasoro” (2n ¼ 40), the taxa were not separable good nutrition during all phases of growth, and persis-
into single clades but formed a group consistent with tence under grazing and cutting (Venuto et al. 2003).
the existence of a large agamic complex. This high- However, its commercial success has been limited by
lighted the difficulty of using the biological species establishment problems due to poor seed quality (less
concept for a genus in which the combination of sexual than 50%), slow germination, and difficulties in seed
and apomictic reproduction can produce large, con- production due to poor seed set and ergot (Claviceps
tinuously variable, and intermingled populations in paspali) infection (Venuto et al. 2003). Burson
which enough genomic mixing occurs to blur morpho- (1991a, b) suggested that the “Common” biotype ori-
logical, cytogenetic, and molecular boundaries ginated from hybridization between tetraploid P. dila-
between populations. Microsatellite (SSR) markers tatum ssp. flavescens and hexaploid P. dilatatum
may provide a tool to enable the further resolution of “Chiru.”
this problem. To this end, Speranza and Malosetti Despite the apomictic nature of common dallis-
(2007) have developed a highly variable set of nuclear grass, genetic improvement has been possible. Intra-
and chloroplast SSR markers that may serve this specific hybridization was carried out between
purpose. “Common” (2n ¼ 50) and yellow-anthered P. dilata-
tum ssp. flavescens (2n ¼ 40; Bennett et al. 1969).
When “Common” was used as female, heptaploid
12.6.1.1 P. dilatatum Poir.: Dallisgrass (2n ¼ 70) hybrids with 10 univalents and 20 trivalents
resulted, indicating that an unreduced female gamete
There are several P. dilatatum biotypes, but desirable had been fertilized. Hybrids had reduced fertility com-
forage biotypes are obligate apomicts and have low pared to the female parent. However, when the yellow-
fertility, while sexual biotypes are remarkably uniform anthered biotype was used as a female, the resultant
with high fertility but have poor forage qualities plants had 45 chromosomes, which associated as 20
204 W.M. Williams et al.

bivalents and 5 univalents at meiosis. Subsequent chromosome behavior at meiosis, with more than half
generations (F2, F3, and F4) had only 40 chromosomes, of the pollen mother cells showing 30 bivalents (2n ¼
which associated as 20 bivalents. The F2 plants were 60, 2C ¼ 3.57). Remaining cells had up to eight
sexual and showed variety for vigor, growth habit, and univalents and rare quadrivalents (Vaio et al. 2007).
fertility. Bennett et al. (1969) were able to select for Burson et al. (1991) hypothesized an IIJJXX genomic
highly fertile (up to 90.8% seed set) and decumbent configuration for this biotype. “Chiru” (referred to as
biotypes, thereby significantly improving the agricul- “Uruguayan”) had significantly higher forage yield
tural qualities of tetraploid P. dilatatum. This was the and higher fertility than “Common,” but similar nutri-
only study identified, which resulted in combined high tional qualities (Venuto et al. 2003). It has reasonable
fertility from the tetraploid and valuable agronomic pollen stainability (42–84%) and variable seed set
traits from the pentaploid. (6–66%) (Burson et al. 1991).
P. dilatatum ssp. flavescens: P. dilatatum ssp. flaves- P. dilatatum “Torres”: The “Torres” biotype origi-
cens Roseng., B.R. Arril. & Izag. is often referred to as nated from the northeastern corner of the state of Rio
yellow-anthered dallisgrass, although two distinct yel- Grande do Sul, Brazil. It is semi-decumbent in habit
low-anthered allotetraploid dallisgrass biotypes are with a significantly lower forage yield than “Com-
mentioned in the literature (Caponio and Quarin 1990; mon” (Venuto et al. 2003). “Torres” is a hexaploid
Quarin and Caponio 1995). It is a sexually and auto- (2n ¼ 60, 2C ¼ 3.58 pg) with very irregular meiosis
gamously reproducing allotetraploid (2n ¼ 40, IIJJ, 2C with 59 univalents on average (Burson et al. 1991;
¼ 2.43 pg) with high fertility and good seed production Vaio et al. 2007). Burson et al. (1991) hypothesized
(Bashaw and Holt 1958b; Vaio et al. 2007), which has that the lack of chromosome pairing was genetically
made it a good subject for research. However, it is not controlled rather than because of a lack of homology
suitable for grazing as it is upright and has poor vigor. It between chromosomes. “Torres” has poor agronomic
is native to Argentina, Uruguay, and southern Brazil. qualities, low pollen stainability, and poor seed set
Bashaw and Patrick (1966) irradiated yellow-anthered (Burson et al. 1991; Venuto et al. 2003).
dallisgrass and obtained promising prostrate and P. dilatatum “Uruguaiana”: The “Uruguaiana” bio-
decumbent plants. However, plants had reduced fertil- type, from a small area in the western Rio Grande do
ity, and vigor declined in later generations. Following Sul, Brazil, has a semi-decumbent growth habit, simi-
crosses of P. dilatatum ssp. flavescens (2n ¼ 40) with P. lar to “Common” dallisgrass (Venuto et al. 2003). It is
notatum (2n ¼ 20, Notata) and P. vaginatum (2n ¼ 20, hexaploid (2n ¼ 60, 2C ¼ 3.69) with variable
Disticha) to determine their genomic relationships, P. chromosome configurations at meiosis, typically with
dilatatum ssp. flavescens  P. notatum and P. dilatatum 10 univalents, 25 bivalents, and rare quadrivalents
ssp. flavescens  P. vaginatum hybrids were triploid (Burson et al. 1991; Vaio et al. 2007). Burson et al.
(2n ¼ 30) with 28–29 univalents at meiosis (Burson (1991) hypothesized an IIJJX1X2 genomic configura-
et al. 1973). This confirmed that P. dilatatum ssp. tion, where there is only partial homology between the
flavescens was allotetraploid, with two unrelated gen- “X” genomes. “Uruguaiana” has low pollen stainabil-
omes, which are unrelated to either P. notatum or ity and poor seed set (Burson et al. 1991).
P. vaginatum genomes. All hybrids were male and P. dilatatum “Bajada de Pena”: P. dilatatum
female sterile. “Bajada de Pena” is a hexaploid biotype (2n ¼ 60,
P. dilatatum “Vacaria” and P. dilatatum “Vira- IIJJX1X2, 2C ¼ 3.79 pg) from Uruguay (Vaio et al.
soro”: The “Vacaria” and “Virasoro” biotypes are 2007).
tetraploid (2n ¼ 40, IIJJ) biotypes from Brazil and Synthetic pentaploid hybrids (2n ¼ 50, IIJJX) were
Argentina (Vaio et al. 2007). DNA contents (2C ¼ generated by intraspecific hybridization between tetraploid
2.33 pg and 2.29 pg, respectively) were determined by yellow-anthered P. dilatatum ssp. flavescens (2n ¼ 40,
flow cytometry (Vaio et al. 2007). IIJJ) and hexaploid P. dilatatum “Chiru” (2n ¼ 60, IIJJXX;
P. dilatatum “Chiru”: The “Chiru” biotype, also Burson 1991a, b). Hybrids had an average of 10 univalents
known as “Uruguayan,” is more upright (up to 1 m at and 20 bivalents at meiosis, suggesting that the “I” and “J”
flowering) and more robust than other hexaploid bio- genomes in each parent were very similar. Some of the
types. It is a pseudogamous, facultative apomict with hybrids resembled “Common” dallisgrass, and Burson
limited sexual reproduction. It has relatively regular (1991b) speculated that the “Common” biotype might
12 Paspalum 205

have arisen from hybridization between tetraploid (e.g., P. seven hybrids. All hybrids were triploid with ten uni-
dilatatum ssp. flavescens or P. urvillei) and hexaploid (e.g., valents and ten bivalents at meiosis (Burson 1979). This
P. dilatatum “Chiru”) cytotypes. However, it remains to be indicated that one of the P. urvillei genomes was similar
determined if the “X” genome in the “Chiru” race is similar to the P. intermedium genome. Several P. dilatatum ssp.
to the “X” genome in common dallisgrass (Burson 1991a, flavescens (2n ¼ 40)  P. urvillei (2n ¼ 40) hybrids
b). Similar results were obtained when Burson (1992) were generated by Caponio and Quarin (1990). Hybrids
hybridized P. urvillei with P. dilatatum “Chiru” (2n ¼ were tetraploid and formed 20 bivalents during meiosis,
60, IIJJXX) and with P. dilatatum “Uruguaiana” (2n ¼ indicating that they had similar genomes and confirming
60, IIJJX1X2). Hybrids were pentaploid with similar the IIJJ genome configuration for P. urvillei. Open pol-
meiotic chromosome behavior to “Common” dallis- linated, backcross and F2 seed were produced from the
grass. Thus, the “I” and “J” genomes of these two hybrids, therefore indicating that the hybrids would be
hexaploid races are the same as those in P. dilatatum useful to a breeding program.
ssp. flavescens and P. urvillei, and by extension, com-
mon dallisgrass, although the origins of the “X” gen-
omes remain uncertain.
12.6.1.4 P. dasypleurum Kunze ex E. Desv

P. dasypleurum has the southernmost distribution

12.6.1.2 P. pauciciliatum (Parodi) Herter:
of South American Paspalum species. It is native to
Prostrate Dallisgrass
central and southern Chile and southwestern Argentina.
It has desirable traits in that it is leafy and is a potential
P. pauciciliatum (syn. P. dilatatum var. paucicilia-
source of cold-tolerance (Quarin and Caponio 1995).
tum Parodi) is a decumbent perennial with forage
The species shows variation for anther color (yellow
potential. It has outyielded and was more persistent
and purple). P. dasypleurum is tetraploid (2x ¼ 40, 2C
than “Common” dallisgrass in field trials, and it has
¼ 2.39 pg) and showed 20 bivalents at meiosis (Vaio
greater resistance to foliage diseases (Venuto et al.
et al. 2007). P. dasypleurum behaves sexually, is self-
2003). However, it is more susceptible to ergot and
fertile, and is predominantly autogamous (Quarin and
less fertile than “common” dallisgrass. It is tetra-
Caponio 1995).
ploid (2n ¼ 40, 2C ¼ 2.40 pg) with up to ten
Quarin and Caponio (1995) intercrossed species of
bivalents at meiosis (Burton and Jackson 1962;
the Dilatata group to determine the genomic relation-
Vaio et al. 2007). A radiation breeding program
ship among these species. Pollination of 253 emascu-
was carried out in P. pauciciliatum and, although
lated P. dilatatum ssp. flavescens (2n ¼ 40, IIJJ)
mutants were generated, the target attributes were
spikelets with P. dasypleurum (2n ¼ 40) yielded two
not altered (Burton and Jackson 1962). One cultivar
hybrid plants. Both hybrids were indehiscent and
has been released (Burton and Wilson 1991).
self-incompatible but produced seed (8.2%) when
backcrossed to the P. dilatatum ssp. flavescens parent.
Pollination of 102 emasculated purple-anthered
12.6.1.3 P. urvillei Steud.: Vasey Grass P. dasypleurum florets with P. urvillei (2n ¼ 40, IIJJ)
generated 55 hybrids. Hybrids segregated for purple (28
P. urvillei is an allotetraploid (2n ¼ 40, IIJJ, 2C ¼ plants) and yellow (27 plants) anthers. Anthers of all
2.40 pg) that reproduces sexually and is usually self- hybrids were highly indehiscent and no self-seed was
pollinated (Vaio et al. 2007). It is a warm season recovered, although seeds were obtained when hybrids
bunch-grass indigenous to southern Brazil and north- were backcrossed to the female (4.8%) and male
ern Argentina but is now grown in warm regions (18.6%) parents. All hybrid plants tested were tetra-
around the world. It is a robust species growing up to ploid (2n ¼ 40) with 20 bivalents at meiosis, indicating
2 m in height, which makes excellent hay and is an an IIJJ genome configuration for P. dasypleurum. The
acceptable forage. success of backcrossing indicated that hybridization
Hybridization between P. urvillei (2n ¼ 40) and among these species may be a useful breeding method
P. intermedium (2n ¼ 20, Quadrifaria) generated for genetic improvement within the Dilatata group.
206 W.M. Williams et al.

12.6.2 Informal Group Livida Chase chromosomal pairing behavior during meiosis. P. juer-
gensii  P. conspersum hybrids had an average of ten
univalents and ten bivalents, suggesting that one of the
12.6.2.1 P. denticulatum Trin.: Longtom
P. conspersum genomes shared homology with the
Paspalum
P. juergensii genome. P. intermedium  P. consper-
sum hybrids typically had 20 univalents and five biva-
P. denticulatum (including P. lividum; Zuloaga and
lents suggesting that there were some similarities
Morrone 2003) is a stoloniferous perennial with forage
between the genomes. Burson (1978) proposed “J”
potential, which is often used for erosion control. It is an
for the P. juergensii genome and “I” for the P. inter-
allopolyploid (2n ¼ 40, 70) with chromosomes typi-
medium genome. He further proposed “IIJJ” for
cally forming bivalents with few univalents and quad-
P. dilatatum ssp. flavescens, and I2I2JJ for P. consper-
rivalents. P. denticulatum reproduces via apospory,
sum, indicating partial homology between P. interme-
followed by pseudospory (Burson and Bennett 1971a).
dium and the other P. conspersum genome.

12.6.3.2 P. rufum Nees ex. Steud

12.6.3 Informal Group Virgata Chase
P. rufum is a robust, erect perennial grass native to
12.6.3.1 P. conspersum Schrad.: Scattered Paraguay, southern Brazil, Uruguay, and northeastern
Paspalum Argentina, where it is normally found in marshes, wet
savannahs, or low, open grounds. Diploid (2n ¼ 20)
This is a leafy, robust, warm-season perennial species, and segmental tetraploid (2n ¼ 40) cytotypes exist
native to South America. It grows up to 2-m tall and (Quarin et al. 1998). The diploid reproduces sexually,
is distributed from Mexico to Argentina, primarily while the tetraploid has irregular meiosis and repro-
in open, moist regions along streams (Chase 1929). duces by aposporous apomixis.
A sexual segmental allotetraploid (2n ¼ 40), it resem-
bles the more common P. virgatum (Burson 1978).
Burson and Bennett (1976) investigated the feasi- 12.6.3.3 P. virgatum L.: Talquezal
bility of using P. conspersum in interspecific hybridi-
zation. Eighty P. dilatatum ssp. flavescens (2n ¼ 40, P. virgatum is a densely clumped perennial, reaching
Dilatata)  P. conspersum (2n ¼ 40) hybrids, and a 2 m at flowering. It is distributed throughout the Amer-
single P. conspersum  P. malachophyllum (2n ¼ 40, icas and Australia (Chase 1929).
Anachyris) hybrid, were developed to study the geno-
mic relationships of P. conspersum. Analysis of mei-
otic chromosome pairing in P. conspersum and
P. conspersum  P. malachophyllum led to the con-
12.6.4 Informal Group Quadrifaria
clusion that P. conspersum genomes were partially Barreto
autosyndetic, i.e., show partial homology between
genomes. P. dilatatum  P. conspersum hybrids were The Quadrifaria group was extracted from the Virgata
tetraploid (2n ¼ 40) with an average of 20 univalents, group by Barreto (1954) based on morphological char-
10 bivalents, and occasional tri- and quadrivalents. acteristics. There are approximately ten Paspalum
Burson and Bennett (1976) deduced that P. dilatatum species in the Quadrifaria Group (Adamowski et al.
and P. conspersum share one genome, and their second 2005; Vaio et al. 2005; Zuloaga and Morrone 2005).
genomes had a small amount of homology. To deter- Many species have both diploid and polyploid cytotypes.
mine which genomes were homologous, P. juergensii Diploid cytotypes are sexual and self-incompatible,
(2n ¼ 20, Paniculata) and P. intermedium (2n ¼ 20, while tetraploids reproduce apomictically. It is widely
Quadrifaria) were crossed to P. conspersum (2n ¼ 40), considered that an ancestral species from the Quad-
giving rise to two and 13 hybrids, respectively (Burson rifaria group was the donor of the Dilatata group “I”
1978). Hybrids were triploid and showed differing genome.
12 Paspalum 207

Hybrids were produced between diploid cytotypes (2n ¼ 20, II, 2C ¼ 1.51 pg; Vaio et al. 2007) and its
of P. haumanii, P. intermedium, P. brunneum (reclas- chromosomes pair as ten bivalents at meiosis (Burson
sified as P. coryphaeum Trin., Zuloaga and Morrone 1981a). It was considered a possible progenitor of one
2003), P. quadrifarium (all Quadrifaria), and P. rufum of the P. dilatatum genomes.
(Virgata) to investigate their genomic relationships
(Quarin and Norrmann 1990). All progeny were dip-
loid and all showed some bivalent pairing during mei- 12.6.4.5 P. quadrifarium Lamk.: Tussock
osis. Quarin and Norrmann (1990) concluded that all Paspalum
five species shared a common genome (I) that had
undergone some changes over time, and the Quadri- P. quadrifarium is native to Uruguay, central and
faria group of species could be included within the eastern Argentina, and Rio Grande do Sul, Brazil,
Virgata group. More recently, Vaio et al. (2005), using where it is found in low, humid, and generally salty
fluorescence in situ hybridization (FISH), rDNA, and areas. It has low forage value but was probably one of
cpDNA sequences, have confirmed differentiation the main species of the “Flooding Pampa” (Perelman
among the Quadrifaria group of species. et al. 2003). P. quadrifarium-dominated regions of the
Flooding Pampa had higher species richness but lower
alien invasion than areas from which P. quadrifarium
12.6.4.1 P. coryphaeum Trin.: Emperor had been removed (Perelman et al. 2003). Diploid
Crowngrass (2n ¼ 20, II, 2C ¼ 1.86 pg), triploid (2n ¼ 30, III0 ),
tetraploid (2n ¼ 40, III0 I0 , 2C ¼ 3.22 pg), and hexa-
P. coryphaeum (including P. brunneum; Zuloaga and ploid (2n ¼ 60) accessions have been identified
Morrone 2003) is native to tropical northeastern (Speranza et al. 2003; Vaio et al. 2007).
regions of South America and the Caribbean where it
grows on small hills or well-drained soils.
12.6.5 Informal Group Paniculata Chase

12.6.4.2 P. exaltatum J. Presl 12.6.5.1 P. juergensii Hack

P. exaltatum (including P. arechavaletae Hack. ex. P. juergensii is a sexual diploid (2n ¼ 20, JJ, 2C ¼
Arechav., Zuloaga and Morrone 2003) is an allotetra- 1.24 pg; Vaio et al. 2007) indigenous to South Amer-
ploid (2n ¼ 40, IIII, 2C ¼ 2.95 pg; Vaio et al. 2007) ica (Brazil and Uruguay). It has been used widely in
that reproduces via apospory, followed by pseudo- genomic composition studies and it is a proposed
gamy. P. arechavaletae was shown to reproduce sex- progenitor of P. dilatatum, donating the “J” genome
ually, with limited apomixis (Burson and Bennett (Burson and Bennett 1971b, 1972; Burson et al. 1973,
1971a). It is native to Brazil and Paraguay. among others). Hybridization studies between P. juer-
gensii and P. vaginatum (2n ¼ 20, Disticha) and with
12.6.4.3 P. haumanii Parodi P. setaceum var. ciliatifolium (2n ¼ 20, Setacea)
showed that the latter species did not contain “J”
P. haumanii is a diploid (2n ¼ 20, II, 2C ¼ 1.50 pg; genomes (Burson 1981a). Resulting hybrids were dip-
Vaio et al. 2007), native to Argentina, Brazil, Paraguay, loid but had very irregular chromosome behavior at
and Uruguay. meiosis, often with only a single bivalent chromosome
pair. Hybrids were male and female sterile.

12.6.4.4 P. intermedium Monro ex Morong &

Britton: Intermediate Paspalum 12.6.5.2 P. paniculatum L.: Arrocillo

P. intermedium is a coarse leaved, erect bunchgrass P. paniculatum is a semi-decumbent weedy species

native to South America (Brazil). It is a sexual diploid native to South America and now distributed in
208 W.M. Williams et al.

warm regions around the world, including Central and subspecies of P. notatum, as had previously been sug-
South America, the Caribbean, and Australia. It is a gested. This species showed chromosome pairing con-
sexual diploid (2n ¼ 20, JJ) and shares a common sistent with being a segmental allotetraploid (Burson
genome with P. dilatatum (Burson 1979). Hybridiza- and Bennett 1971a). The morphological affinities of
tion between P. dilatatum ssp. flavescens (2n ¼ 40) P. dedeccae Quarin, P. ionanthum Chase, and P. fili-
and P. paniculatum (2n ¼ 20) generated a single folium Nees ex Steud. with the Notata group were not
triploid hybrid (2n ¼ 30; Burson 1979). Chromosomes supported by DNA results. Despite its morphological
associated as ten univalents and ten bivalents at meio- similarity, P. almum Chase (x ¼ 6) was distant from
sis. This indicated that one of the P. dilatatum gen- the Notata group (Souza-Chies et al. 2006).
omes was similar to the P. paniculatum genome and
led Burson (1979) to conclude that a species or ances-
tor of the Paniculata group was one of the progenitors € gge: Bahiagrass
12.6.6.1 P. notatum Flu
of P. dilatatum.
P. notatum is one of the most important species of the
Notata group. It is a rhizomatous perennial and is
a major constituent of the native grasslands from Mex-
12.6.6 Informal Group Notata Chase ico to Argentina and Uruguay. Most biotypes of
P. notatum var. notatum are self-compatible apomictic
This group intergrades with the groups Linearia and tetraploids (2n ¼ 4x ¼ 40) with a genome formula of
Alma and with P. equitans Mez (Fasciculata). The NNNN (Burson 1983; Quarin et al. 1984). However,
boundaries of these groups remain unclear and contro- self-incompatible sexual diploid (2n ¼ 2x ¼ 20) popu-
versial (Ferreira and Souza-Chies 2005; Souza-Chies lations are native to northeastern and central-eastern
et al. 2006; Essi and Souza-Chies 2007), and reference Argentina and are collectively classified as P. notatum
to the Notata/Linearia group is frequent. For example, var. saurae Parodi (Dahmer et al. 2008). This variety
Zuloaga et al. (2004) used a single combined subgroup has become very popular as a forage grass in southern
in classifying species of the Notata group. However, USA, where it is known as Pensacola bahiagrass. It has
there have been progressive indications that the group- a small genome with a 1C DNA content of 0.6 pg
ings based on morphological traits are not monophy- (Jarret et al. 1995) and a genome formula of NN (Bur-
letic when tested by molecular data. son 1981b). Some occasional findings of triploid and
On the basis of morphological and DNA sequence pentaploid individuals have been reported (Quarin
analyses, Souza-Chies et al. (2006) identified a “core” 1992; Tischler and Burson 1995; Daurelio et al.
Notata group as P. conduplicatum Canto-Dorow, 2004). There is variability reported within the apomic-
Valls & Longhi-Wagner, P. notatum, P. minus, tic tetraploid P. notatum, but no clear separation allows
P. pumilum Nees, and P. subciliatum Chase. the division into different botanical varieties (Cidade
Essi and Souza-Chies (2007) used DNA sequence et al. 2008).
data to identify a core monophyletic group of eight Bahiagrass is also used as a turfgrass in warm,
species making up a Notata group senso stricto. These humid areas. It is characterized by slow seedling
were P. conduplicatum, P. notatum, P. minus, P. pumi- establishment and slower vegetative growth than
lum, and P. subciliatum, along with P. barretoi Canto- other subtropical grasses (Busey 1992). It performed
Dorow, P. nummularium Chase, and P. flaccidum extremely well as a forage grass in subtropical
Nees, which is morphologically distinct and previ- Australia in a trial conducted by Shaw et al. (1965).
ously classified as part of the Recta group (Zuloaga Cidade et al. (2008) used inter-simple sequence
et al. 2004). P. barretoi, P. nummularium, and repeat (ISSR) markers to show that considerable
P. pumilum are known only as sexual autogamous genetic diversity exists among tetraploid populations
diploids, whereas P. conduplicatum is hexaploid, collected near the likely center of origin in South
P. minus is pentaploid, and P. flaccidum is tetraploid Brazil, despite the apomictic breeding system. This
(Pozzobon et al. 2008). diversity was attributed to the presence of occasional
P. cromyorhizon Trin. ex Doell was found to be not sexual diploids and/or tetraploids with facultative apo-
close enough in DNA sequence to be regarded as a mixis. Thus, the species has a powerful evolutionary
12 Paspalum 209

system of generation of new genetic diversity fol- and breeding. Further development of sexual tetra-
lowed by rapid fixation of superior combinations by ploids from the above material led to the registration
apomixis. of two germplasm lines for breeding purposes (Quarin
Pillay and Mazzella (1997) compared the chloro- et al. 2003). A progeny plant from one of these lines
plast genome of tetraploid P. notatum (2n ¼ 4x) with was shown by molecular marker analysis to reproduce
that of “Common” P. dilatatum (2n ¼ 5x) using totally by sexual means, confirming the potential of
restriction site analysis. The two species had clearly this material for plant breeding (Ortiz et al. 1997).
different mutations that distinguished the chloroplast Time of pollination can be altered to achieve bipa-
genomes, a result consistent with morphological rental progeny from apomictic tetraploid P. notatum.
and nuclear cytological data, indicating that there is Martinez et al. (1994) achieved (2n þ n) progenies by
a considerable genetic distance between them. The pollinating tetraploid P. notatum 2 or 3 days before
overall Paspalum chloroplast genome was organized anthesis with either the same cytotype or with diploid
in a similar way to those of wheat and barley. Pensacola bahiagrass. Apparently, early pollination
Apomixis in P. notatum: In P. notatum, and preempted the parthenogenic development of the unre-
most apomictic Paspalum species, seeds develop fol- duced egg, allowing fertilization to occur. Thus,
lowing apospory, parthenogenesis, and pseudogamy. despite apomixis, it is possible to create new genome
In ovules, the meiotic products break down and are combinations among wild species and to breed within
replaced by embryo sacs (usually multiple) developed this complex.
from nucellar cells. These have the sporophytic chro- Acuna et al. (2007) have confirmed Burton (1955)
mosome number and are distinguished from meiotic in finding that some genotypes of diploid Pensacola
embryo sacs by a lack of the proliferated antipodal bahiagrass can show a significant amount of self-
cells characteristic of the latter. In facultative apomic- fertility, and it should not be assumed that all are
tic plants that have a high degree of sexuality, some fully outcrossing. The same applied to artificial tetra-
ovules produce both kinds of embryo sacs (Martinez ploids produced from chromosome doubling of the
et al. 2001). Autotetraploids produced by colchicine- diploids. Some artificial tetraploids also occasionally
doubling of sexual diploids are often of this facultative produced apomictic embryo sacs (Quarin et al. 2001;
apomictic type (Quarin et al. 2001). Apomixis in tet- Acuna et al. 2007), indicating that the alleles for apo-
raploid P. notatum was found to be environmentally mixis must be present in at least some diploids.
stable by Burton (1982a). Breeding diploid sexual “Pensacola bahiagrass”:
Breeding apomictic P. notatum: Although most Burton (1955) showed that this diploid material was
populations of P. notatum are tetraploid, it has been predominantly cross-fertilizing and highly variable. It
noted that the diploid type (Pensacola bahiagrass) has showed considerable (25%) loss of vegetative yield
several more desirable agronomic characteristics, with inbreeding. Two of the classical forage plant
including more rapid spread, longer leaves, greater breeding methods, recurrent restricted phenotypic
winter-hardiness, better ergot resistance, and small selection, RRPS (Burton 1974), and the improved
seeds that do not require scarification (Burton and version (Burton 1982b) were refined on Pensacola
Hanna 1986). It was, therefore, important to be able bahiagrass. Nine cycles of RRPS resulted in the deve-
to hybridize the diploid and tetraploid to combine the lopment of the cultivar Tifton 9 (Burton 1989). This
features of both. This could be done by chromosome material continues to be a source of useful sexual
doubling the diploid to obtain sexual tetraploids and genetic diversity for the improvement of the species.
crossing these with apomictic tetraploids to obtain
sexual and apomictic tetraploid families (Burton and
Forbes Jr 1960; Forbes and Burton 1961). Burton and 12.6.6.2 P. subciliatum Chase
Hanna (1986) achieved the same objective by using a
spontaneous male sterile triploid, which produced tet- This tetraploid species has been little studied cyto-
raploid progeny when pollinated with the diploid pop- genetically. One Brazilian accession analyzed by
ulation. These tetraploids were facultative apomicts Adamowski et al. (1998) showed 20 bivalents at meta-
that, when reproducing sexually, generated variable phase I of meiosis, suggesting an allotetraploid origin.
families that could form base populations for selection After this stage, one of the parental chromosome sets
210 W.M. Williams et al.

appeared to be out of phase with the other, resulting in completely sterile (Burson et al. 1973). Similarly
the elimination of chromosomes from one set. Pollen hybridization of P. juergensii and P. vaginatum was
fertility was greatly reduced, but seed viability was successful only when P. vaginatum was the male par-
maintained. Chromosome elimination during gameto- ent. Again, there was almost no chromosome pairing
genesis is unusual in plants as it usually occurs in and none of the hybrids produced seed (Burson
somatic cells during early embryogenesis. 1981a). P. vaginatum does not have either “I” or “J”
genomes found in the Dilatata complex, and the geno-
mic formula “DD” was proposed (Burson 1981a).
The existence of significant genetic variation in salt
12.6.7 Informal Group Disticha Chase tolerance among populations of P. vaginatum (Lee et al.
2005) has led to the use of this species as a tool for
The Disticha group consists of three species, including investigation of salt tolerance mechanisms (e.g., Lee
P. distichum L. and P. vaginatum Swartz. The latter is et al. 2008). It has increased osmotic potential and may
the most important among them (Hojsgaard et al. also have the ability to take up potassium and lose excess
2009) and can also be found in a diploid, tetraploid, salt through old leaves (Shonubi and Okusanya 2007).
and hexaploid forms (Quarin and Burson 1983). High frequency plant regeneration from embryo-
genic callus cultures was achieved by Cardona and
Duncan (1997). This opened the way for the potential
12.6.7.1 P. vaginatum Swartz: Seashore use of somaclonal variation to generate variants for
Paspalum genetic improvement.

Native to tropical and subtropical America, this spe-

cies is adapted to halophytic coastal environments and
12.6.7.2 P. distichum L.: Knot Grass
can stand a very wide soil fertility and pH range
(4–10). It is used for forage production and is exten- This species is native to the Americas and has been
sively used as a turfgrass for golf courses, lawns, and introduced widely around the world. It is a rhizomatous
parks and for soil conservation where salt tolerance is perennial adapted to wetlands, both saline and fresh
required (Dudeck and Peacock 1985). It is also used water. It is palatable with moderate spreading ability,
for land reclamation under saline conditions and drain- high salt tolerance, but lower shade tolerance than
age water recycling schemes. It is hard-wearing due to P. vaginatum. It is used on contaminated soils where
the shoot density, has general pest resistance, salt it takes up minerals such as lead, zinc, and cadmium
tolerance, and some drought, waterlogging, and cold from the soil (Lan et al. 1992; Yu et al. 2005).
tolerance (Dudeck and Peacock 1985; Huang et al.
1997a, b; Trenholm et al. 1999, 2000; Lee et al.
2005). It exists in dwarf and semi-dwarf forms with
short leaves, generally less than 10 cm. The species is
more tolerant of low light intensities than hybrid
12.6.8 Informal Group Plicatula Chase
Cynodon cultivars (Jiang et al. 2004). Several cultivars
have been developed in SE USA and in Australia The Plicatula group consists of up to 20 species
based on dwarf and semi-dwarf material. (Zuloaga and Morrone 2005) and shows a lot of poten-
More than 300 populations have been collected that tial for agriculture. Central and western Brazil, Boli-
show considerable diversity in leaf texture, from coarse via, and Paraguay are the center of variation for this
to very fine (Cardona and Duncan 1997). The diploid group (Quarin et al. 1997). The most important species
forms are self-incompatible and produce seeds, but known to be cultivated include the apomictic tetra-
yields are poor and seeds are generally low in viability. ploids (2n ¼ 4x ¼ 40) P. atratum Swallen, P. plicatu-
Crosses between sexual tetraploid P. dilatatum and lum Michx., P. glaucescens Hack., and P. guenoarum
P. vaginatum produced triploid hybrids only when Arechav. (Quarin et al. 1997; Takayama et al. 1998;
P. vaginatum was used as the male parent. There was Pozzobon and Valls 2000). The ancient Asian cereal
virtually no chromosome pairing and the hybrids were P. scrobiculatum L. also occurs in this group.
12 Paspalum 211

12.6.8.1 P. atratum Swallen: Atra Paspalum, by frost, but the crown survives (Shaw et al. 1965). It is
Capim-pojuca, Pasto Pojuca less tolerant of grazing than P. atratum and is not
shade-tolerant. P. plicatulum is less palatable than
This native of Brazil and Bolivia has only relatively other grasses and declines rapidly in quality with matu-
recently become used in agriculture. It is naturalized in rity. It is established from seed or cuttings. It is free
the subtropics of Australia, USA, and Southeast Asia. It of ergot attack, and dry seed can last for up to 2 years.
is in use in Thailand, Bhutan, Australia, and USA for It is also free of other diseases and has no weedy
forage or hedgerows for erosion control. It grows up to tendencies. An accession from Guatemala performed
two meters tall when flowering and is suitable for cut and extremely well in trials in Queensland, Australia
carry forage systems and shows promise for hay making. (Shaw et al. 1965). Three cultivars have been devel-
It can tolerate soils of low fertility in regions of high oped in Queensland, Australia, from different materials
rainfall (more than 1,000 mm) and tolerates waterlog- (Oram 1990).
ging. It has limited cool season growth and is damaged This is predominantly an apomictic allotetraploid
by frost but recovers quickly with onset of warm condi- species, but there are occasional self-incompatible
tions. It is shade-tolerant and useful in agroforestry. It is sexual diploids (Pagliarini et al. 2001; Adamowski
highly palatable with very good forage quality but is et al. 2005; Pozzobon et al. 2008). Chromosome dou-
better suited to cutting or lenient grazing managements bling of the diploids to produce sexual tetraploids may
(Quarin et al. 1997; Kalmbacher and Martin 1999). provide a way of breeding at the tetraploid level.
Relative to many other apomictic Paspalum species,
P. atratum is an exceptionally good seeder (Quarin et al.
1997). Nevertheless, poor seed quantity (110 kg/ha) and 12.6.8.3 P. guenoarum Arechav.: Pasto Rojo
quality limits its commercialization (Quarin et al. 1997;
Kalmbacher et al. 1999). Cultivars have been developed P. guenoarum is a leafy, upright perennial, which can
in USA, Brazil, and Thailand, but all are derived from a reach 2 m when flowering. It originated from western
single accession from Campo Grande, Brazil. South America and is currently used in South America,
P. atratum is a pseudogamous obligate apomictic Zimbabwe, and Australia. P. guenoarum is used for
tetraploid with an average of 19 bivalents and very few grazing, cut-and-carry, hay, and silage. It is moderately
uni- or quadrivalents, suggesting that it is a probable tolerant of water-logging and drought and grows well in
segmental allotetraploid (Quarin et al. 1997, Takayama areas with annual rainfalls between 1,200 and
et al. 1998). Unless a sexual diploid (or tetraploid) is 2,000 mm per year. Natural populations of P. gue-
found in future germplasm collections, population noarum are found in an unusually narrow temperature
improvement will be limited to screening of wild range (19–21 C); although “Bela Vista” (Brazil) per-
accessions (Quarin et al. 1997). forms well in areas with average annual temperatures of
27 C. Genotypes tested in Australia have shown good
frost tolerance. P. guenoarum is palatable to cattle and
horses and performs best under lenient management.
12.6.8.2 P. plicatulum Michx.: Brownseed Several cultivars are available in South America, and
Paspalum promising accessions have been identified in material
imported into Australia.
Native to southeastern USA, Central America, and the All accessions tested are tetraploids (2n ¼ 40;
Caribbean, this species is now widespread throughout Burson and Bennett 1971a; Takayama et al. 1998;
South America, China, Australia, and temperate Asia. Espinoza et al. 2001; Adamowski et al. 2005) and
Its natural adaptation is to moist habitats in high rainfall likely to be segmental allotetraploids (Burson and
regions, but it can tolerate as little as 750 mm per year. Bennett 1971a). Pollination is needed for endosperm
It is perennial and grows up to 1.3-m tall. P. plicatulum development, indicating pseudogamy in this species,
is used as permanent pasture in poorly drained low and Burson and Bennett (1971b) showed that pollen
fertility soils, tolerating high aluminum concentrations. stainability was reasonable (47.1%) and suggested
It is tolerant of waterlogging and flooding for short that P. guenoarum pollen could be used in Paspalum
periods and is moderately drought hardy. It is burned breeding programs.
212 W.M. Williams et al.

12.6.8.4 P. glaucescens Hackel tula group. In many respects, this complex typifies the
situation with regard to the potential use of wild spe-
P. glaucescens (including P. yaguaronense Henrard; cies in breeding new populations within the genus.
Pozzobon and Valls 2000) populations are from Savan-
nas and ill-drained soils in Argentina, Brazil, and Para-
guay. Material was imported into Australia in 1948 and 12.6.8.7 Other Species in the Plicatula Group
was shown to have high palatability, better annual
yields than naturalized P. dilatatum, and good frost A number of diploid species and cytotypes of the
tolerance (Shaw et al. 1965). Diploid, tetraploid, and Plicatula group might be useful for interspecific
hexaploid accessions have been identified (Pritchard breeding programs. Brazilian species P. hydrophilum
1962; Takayama et al. 1998; Pozzobon and Valls 2000; Henrard (2n ¼ 20, 30, 40), P. modestum Mez (2n ¼ 20,
Pozzobon et al. 2000; Pagliarini et al. 2001). Diploids 40), and P. palustre Mez (2n ¼ 20) were identified as
(2n ¼ 20) reproduce sexually and are typically self- species with potential for use in wetlands (Pozzobon
incompatible (Pritchard 1962; Pozzobon and Valls and Valls 2003). Espinoza et al. (2001) speculated that
2000), although Pritchard (1962) identified a single P. limbatum Henrard (2n ¼ 20, sexual) and P. lenti-
self-fertile plant. Tetraploids (2n ¼ 40) and hexaploids culare Kunth (2n ¼ 40, apomictic) were diploid and
(2n ¼ 60) are apomictic and are thought to be segmen- tetraploid accessions of the same species.
tal allotetraploids and interspecific hybrids, respec-
tively (Pozzobon and Valls 2000).
12.6.8.8 P. scrobiculatum L.: Kodo Millet

12.6.8.5 P. nicorae Parodi: Brunswick Grass One of the minor millets, this tropical species is grown
in fertility-depleted soils in Africa and Asia (Clayton
P. nicorae is native to eastern South America and 1975). Its origin is uncertain, but it appears to have
naturalized in USA and Australia. It is a variable been domesticated in India in Neolithic and post-
perennial with long, deep, and vigorous rhizomes Neolithic times (Vishnu-Mittre 1978) and is now
and is short-statured, reaching only 40 cm at heading mainly of local tribal use. It is found in the wetter
(Barreto 1956). It is used for permanent pasture in the parts of Nigeria in moist and shady places but is
tropics, being tolerant of heavy grazing, palatable, and not halophytic and so is found in non-saline regions
drought and frost-tolerant. It is an excellent seed pro- (Shonubi and Okusanya 2007). It is very invasive and
ducer, sometimes producing over 1 ton of seed/ha. It is a noxious weed in North America. Somatic embryo-
is a tetraploid and facultative apomict (Burson and genesis has been achieved (Vikrant 2002).
Bennett 1970; Pagliarini et al. 2001) that may be
interfertile with P. plicatulum. Cultivars have been
developed for pasture and turf in Georgia, USA, and
Queensland, Australia. 12.6.9 Recommendations for Further
Action

12.6.8.6 P. compressifolium Swallen The powerful evolutionary system of Paspalum needs

to be better understood, if it is to be exploited by
P. compressifolium Swallen is an important perennial plant breeders. Great genetic power is generated
forage contributor to native grasslands in South by the occurrence of occasional sexual hybridization
America, especially South Brazil, East Paraguay, and of diverse plants, followed by fixation of successful
Northeast Argentina. It has been shown to be a proba- polyploid genotypes by apomixis. If breeders could
ble agamic complex composed of occasional diploid, exploit this system, then considerable advances
predominant tetraploid, and rare hexaploid races would be made, not only in Paspalum, but also in
(Quarin et al. 1996; Adamowski et al. 2005). The other species that combine sexual and apomictic
existence of sexual diploid populations provides a modes of reproduction. In this context, the identifica-
way to breed with this complex and within the Plica- tion and conservation of the sexual diploid Paspalum
12 Paspalum 213

species requires priority action as these species will Burson BL (1981b) Genome relations among four diploid Pas-
provide the engine for generation of new and useful palum species. Bot Gaz 142:592–596
Burson BL (1983) Phylogenetic investigations of Paspalum
genetic diversity that can be used by plant breeders. dilatatum and related species. In: Proceedings of 14th inter-
national grassland congress, Lexington, Kentucky, USA,
15–24 June 1981, pp 170–173
Burson BL (1989) Phylogenetics of apomictic Paspalum dila-
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Chapter 13
Pennisetum

Thierry Robert, Nadra Khalfallah, Evelyne Martel, Françoise Lamy, Valerie Poncet, Clémentine Allinne,
Marie-Stanislas Remigereau, Samah Rekima, Magalie Leveugle, Ghayas Lakis, Sonja Siljak-Yakovlev,
and Aboubakry Sarr

13.1 Structure and Evolutionary millet with its wild relatives was proposed by Harlan
Relationships Within Pennisetum and de Wet (1971). Later Pernès (1984) extended this
gene pool classification to the concept of “complex of
Complex of Species
species”. This taxonomic representation is based not
only on the crossability of wild species with the
13.1.1 Systematic Considerations domesticated form but also on the amount of gene
of the Genus flow occurring between all the members of the gene
pool. This classification includes three gene pools. The
The genus Pennisetum belongs to the family Poaceae, primary gene pool consists of the domesticated form
group Paniceae of the subfamily Panicoideae and is (P. glaucum ssp. glaucum), the wild annual and weedy
closely related to the genera Cenchrus and Setaria. It forms (P. glaucum ssp. monodii represented by two
includes approximately 140 species (Brunken 1977) ecotypes: P. violaceum and P. mollissimum). These
distributed in tropical and subtropical regions. The wild forms cross easily with the cultivated form to
genus Pennisetum has been given various taxonomic produce viable seeds and fertile hybrids. The second-
status (see Jauhar 1981 for a review). The Stapf and ary gene pool includes a perennial fodder wild relative
Hubbard (1934) classification of Pennisetum based on to P. glaucum, P. purpureum, named as Elephant or
morphological characters divides the genus into five Niaper grass. Recently, P. squamulatum was included
sections: Penicillaria, Brevivalvula, Gymnothrix, Het- in the secondary gene pool. These two species are
erostachya, and Eu-Pennisetum. Other infrageneric easily crossable with pearl millet, but the hybrids are
classifications showing some discrepancies were pro- highly sterile. The tertiary gene pool includes true
posed by different authors (Pilger 1940; Brunken biological species compared to the primary and sec-
1977; Clayton and Renvoize 1986). The section Pen- ondary gene pool species. Indeed, strong reproductive
nicillaria includes the domesticated crop Pennisetum barriers impede natural gene flow and occurrence of
glaucum (pearl millet), which is a very important hybrids between the members of the tertiary gene pool
cereal crop in Africa and India but serves also as and the forms belonging to primary and secondary
fodder in other areas. The other species are of multiple gene pools. The tertiary gene pool includes the major-
purpose uses (forage, pasture grasses, or ornamentals, ity of the species presented in Table 13.1.
Table 13.1 and Fig. 13.1). A gene pool classification
based on the level of reproductive isolation of the pearl

A. Sarr (*) 13.1.2 Chromosome Evolution

Laboratoire Ecologie Systématique et Evolution UMR (8079)
Université Parsi-Sud, CNRS, AgroParisTech,
Université ParisSud, B 360 91 405 Orsay, France
The species of the Pennisetum genus present different
e-mail: [email protected] basic chromosome numbers x ¼ 5, 7, 8, or 9 (Jauhar
This chapter has been contributed by lead contributors, Thierry 1981), variable genome sizes, and ploidy levels rang-
Robert, Valerie Poncet, and Aboubakry Sarr. ing from diploid to octoploid.

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 217
DOI 10.1007/978-3-642-14255-0_13, # Springer-Verlag Berlin Heidelberg 2011
218 T. Robert et al.

Table 13.1 List of 82 Pennisetum species with some characteristics. Kingdom: Plantae, Subkingdom: Tracheobionta, Super-
division: Spermatophyta, Division: Magnoliophyta, Class: Liliopsida, Subclass: Commelinidae, Order: Cyperales, Family: Poaceae,
Genus Pennisetum Rich
Species Life cycle type Geographic range Chromosome number and 2C value (pg)
ploidy level (x)
n 2n
Pennisetum advena (Wipff and Veldkamp) P North America – – –
P. alopecuroides (L.) Spreng. P Asia-temperate, Asia-tropical, 18 18 (2x) 1.90
Australasia 22
P. annuum Mez A South America – – –
P. articulare Trin ex Spreng. P Pacific – – –
P. bambusiforme (Fourn) Hemsl ex B. D. Jacks. P America – 36 –
P. basedowii Summerh. and C. E. Hubb. A Australasia – 54 –
P. beckeroides Leeke P Africa – – –
P. caffrum (Bory) Leeke P Africa Indian ocean – – –
P. centrasiaticum Tsvelev P Asia-temperate – 36 –
P. chilense (E. Desv.) B. D. Jacks. ex R. E. Fr. P South America – – –
P. clandestinum Hochst. ex Chiov. P Africa, Asia-temperate, Asia- 18 36 2.30
tropical, Australasia,
Pacific, America, Antarctic.
P. complanatum (Nees) Hemsley P Pacific, North America – – –
P. crinitum (HBK) Spreng P North America, Mexico – – –
P. divisum (Forssk. ex J. F. Gmel.) Henrard P Africa, Asia-temperate, Asia- 18, 27 36 –
tropical
P. domingense (Spreng. ex Schult.) Spreng. P South America – – –
P. durum Beal P North America, Mexico. – – –
P. flaccidum Griseb P Asia-temperate, Asia-tropical 9, 18, 27 – –
P. foermeranum Leeke P Southern Africa – – –
P. frutescens Leeke P South America – – –
P. glaucifolium Hochst. ex A. Rich. P Africa, Asia-temperate – – –
P. glaucum (L.) R. Br A Wide 7 14 (2x) 4.71
P. gracilescens Hochst. P Africa – – –
P. henryanum F. Br. P Pacific – – –
P. hohenackeri Hochst. ex Steud P Africa, Asia-tropical 9 18 (2x) 1.69
P. hordeoides (Lam.) Steud A Africa, Asia-tropical – – –
P. humile Hochst. ex A. Rich. P Africa – – –
P. intectum Chase P South America – – –
P. lanatum Klotzsch P Asia-temperate, Asia-tropical 18 – –
P. latifolium Spreng P Australasia, South America – 36 –
P. laxior (Clayton) Clayton A Africa – – –
P. ledermannii Mez P Africa – – –
P. longissimum S.L. Chen & Y.X. Jin P Asia-temperate – 54 –
P. longistylum Hochst. ex A. Rich P Africa – 45 –
P. macrostachyum (Brongn.) Trin. P Asia-tropical, Pacific – 54 –
P. macrourum Trin P Africa, Asia-temperate – 36, 54 –
Australasia
P. massaicum Stapf P Africa – 32 –
P. mezianum Leeke P Africa, Asia-tropical – 16, 32 3.01 (4x)
P. mildbraedii Mez P Africa – – –
P. mollissimum Hochst. A – 14 (2x) 4.51
P. monostigma Pilg. P Africa – 18 –
P. montanum (Griseb.) Hack. P South America – 32 –
P. nervosum (Nees) Trin. P America – 36, 72 –
P. nodiflorum Franch P Africa – – –
P. nubicum (Hochst.) K. Schum. ex Engl. A Africa, Asia-temperate – – –
P. occidentale Chase P South America – – –
P. orientale Rich P Africa, Asia-temperate, 7, 18 36 (4x) 3.77
Asia-tropical, Pacific, 27, 56
South America
P. pauperum Nees ex Steud. P South America – – –
(continued)
13 Pennisetum 219

Table 13.1 (continued)

Species Life cycle type Geographic range Chromosome number and 2C value (pg)
ploidy level (x)
n 2n
P. pedicellatum Trin. A Africa, Asia-tropical, 14, 18, 27 54 (6x) 5.61
Australasia, South America
36 (4x) 4.40
P. peruvianum Trin. P South America – – –
P. petiolare (Hochst.) Chiov. A Africa – – –
P. pirottae Chiov. P Africa – – –
P. polystachion (L.) Schult. A or P Wide 27 54 (6x) 5.66
36 (4x) 4.25
34, 63
P. procerum (Stapf) Clayton P Africa – – –
P. prolificum Chase P North America – – –
P. pseudotriticoides A. Camus P Africa – – –
P. pumilum Hack. & Engl. P Africa – – –
P. purpureum Schumach. P Wide 7, 14 28 (4x) 4.59
21
P. qianningense S.L. Zhong P Asia-temperate – 36
P. ramosum (Hochst.) Schweinf. & Asch. A Africa 5 10 (2x) 4.04
20
P. rigidum (Griseb.) Hack. P South America – – –
P. riparium Hochst. ex A. Rich. P Africa – – –
P. rupestre Chase P South America – – –
P. sagittatum Henrard P South America – – –
P. schweinfurthii Pilg. A Africa – 14 (2x) 4.97
P. setaceum (Forssk.) Chiov. P Europe, Africa, Asia-temperate 18 27 (3x) 2.78
Australasia, Pacific 54 (6x) 5.28
P. shaanxiense S.L. Chen & Y.X. Jin P Asia-temperate – – –
P. sichuanense S.L. Chen & Y.X. Jin P Asia-temperate – – –
P. sieberianum (Schltdl.) Stapf & C.E. Hubb. A Africa, Asia-temperate – – –
P. sphacelatum (Nees) T. Durand & Schinz P Africa – 18 –
P. squamulatum Fresen. P Africa. 34 54 (6x) 9.56
56 (8x)
P. stramineum Peter P Africa, Asia-temperate – – –
P. tempisquense R.W. Pohl P South America – – –
P. thulinii S.M. Phillips P Africa – – –
P. thunbergii Kunth P Africa, Asia-temperate, Asia- 18 – –
tropical, Australasia
P. trachyphyllum Pilg. P Africa – – –
P. trisetum Leeke P Africa 36 – –
P. tristachyum (Kunth) Spreng. P America – – –
P. uliginosum Hack. P Africa – – –
P. unisetum (Nees) Benth. P Africa, Asia-temperate – – –
P. villosum R. Br. Ex Fresen. P Cosmopolitan 27 36 (4x) 3.47
– 45, 54
P. violaceum (Lam.) L. Rich. A Africa 7 14 (2x) 4.52
P. weberbaueri P South America – – –
P. yemense P Africa, Asia-temperate – – –
P Perennial, A annual
Sources: Clayton WD, Harman KT, Williamson H (2006 onwards). GrassBase – The Online World Grass Flora: http://www.kew.
org/data/grasses-db.html (accessed 08 March 2009)
Index to Plant Chromosome Numbers Data Base, Missouri Botanical Garden-w3TROPICOS: http://mobot.mobot.org/W3T/search/
ipcn.html (accessed 26 Feb 2009)
Botanica sistematica Italia: http://www.homolaicus.com/scienza/erbario/utility/botanica_sistematica/hypertext/1524.htm#000000
(accessed 21 April 2009)
Bennett MD, Leitch IJ (2005) Angiosperm DNA C-values database (release 4.0), Royal Botanic Gardens, Kew: http://data.kew.org/
cvalues/ (accessed 28 Feb 2009)
Fedorov (1969) Editor, Chromosome numbers of flowering plants. Academy of sciences of the USSR, V.L. Komarov Botanical
Institute, Nauka, Leningrad
220 T. Robert et al.

Fig. 13.1 Some of the studied Pennisetum species (spike, (e), P. pedicelletum (f), P. polystachyon (g), P. purpureum
rachis and spikelets): P. alopecuroides (a), P. hohenackeri (h), P. ramosum (i), P. schweinfurthii (j), P. setaceum
(b), P. mollissimum (c), P. mezianum (d), P. orientale (k), P. squamulatum (l), P. villosum (m) (Original illustrations)
13 Pennisetum 221

The diversity in basic chromosome number and present an SC on terminal position of the seventh pair,
cytological observation of different chromosome while the cultivar Ligui, originating from Chad, exhi-
sizes between species, with the larger size for the bits an intercalary SC on the sixth chromosome pair
lower numbers (Jauhar 1981), reflect chromosome (Fig. 13.2a–d). In situ hybridization analysis revealed
repatterning during evolution within the genus. The that the localization of the tandemly repeated multigene
dysploidy process, which has occurred in many plants, family coding for the 18S–5.8S–25S rRNAs (rDNA)
involves structural modification of the karyotype by and genes coding for the 5S rRNA (5S rDNA) were
change in the basic chromosome number during similar among members of the primary gene pool
phylogenetic divergence of species (Stebbins 1956; (Martel et al. 1996). Within the primary gene pool
Siljak-Yakovlev 1996). Based on chromosome pairing (P. glaucum, P. molissimum and P. violaceum),
observations in Pennisetum, an increase in chromo- rDNA sequences were detected on the telomeric part
some number from a basic chromosome number of of the short arm of pair VI and at the nucleolar orga-
x ¼ 5 has been suggested by Jauhar (1970, 1981) and nizing region (NOR) of the satellited pair (VII). The
Rao et al. (1989). However, the frequent occurrence of 5S rDNA was detected in the telomeric position of the
the basic chromosome number x ¼ 9 in the Pennise- short arm of metacentric chromosome pair IV of the
tum genus and in closely related genera Cenchrus, three species of the primary gene pool (Fig. 13.3).
Panicum, and Setaria may suggest an alternative sce- Analysis of the genome size and base composition
nario to the proposed ancestral basic chromosome of species in the primary gene pool made using flow
number of x ¼ 5. cytometric techniques has shown that there is a similar
In this section, we present the chromosome evolu- amount of DNA per basic chromosome set among
tion within Pennisetum gene pools focusing on basic these species (Martel et al. 1997). The species with
chromosome number. x ¼ 7 have monoploid genome size (1Cx-value) rang-
ing from 2.25 to 2.36 pg and are relatively GC-rich
(between 44.9% and 47.2%) compared with the other
13.1.2.1 Species with x ¼ 7 Basic Chromosome Pennisetum species analyzed (Table 13.2).
Number Within the genus Pennisetum, P. schweinfurthii is
the only other diploid species with x ¼ 7. This species
Cultivated pearl millet, P. glaucum ssp. glaucum, and belongs to the tertiary gene pool and exhibits a strong
its wild relative P. glaucum ssp. monodii belong to the reproductive barrier with the cultivated P. glaucum
Penicillaria section. These species with 2n ¼ 2x ¼ 14 (Hanna and Dujardin 1986). A small number of artificial
chromosomes (AA genome) are part of the same pri- hybrids have been obtained between P. schweinfurthii
mary gene pool (Brunken 1977). They are not repro- and cultivated P. glaucum, but these are completely
ductively isolated and can cross-hybridize when they sterile (Hanna and Dujardin 1986; Marchais and
grow in sympatry, forming fertile hybrids with normal Tostain 1997). This feature is consistent with the dif-
chromosome pairing. Despite this gene flow, several ferences in chromosome configuration observed
mechanisms such as linkage, gametophytic competi- between these species. Their karyotypes differ notably
tion, and phenology (Robert et al. 1991, 1992; Robert in the relative size of chromosome pair II, which is
and Sarr, 1992) have been shown to contribute to the larger in P. schweinfurthii (Martel et al. 1996). This
maintenance of the genetic structure of these species. difference could be attributed to the haploid genome
The cultivated forms and the wild species from which size of 2.49 pg observed for these species, which is the
they evolved (Pernès 1983) have conserved a high largest in the genus. The rDNA loci have been observed
level of similarity of their genomes. Cytogenetic on the telomeric part of the short arm of two satellited
investigations of cultivated and wild stocks of this pairs of chromosomes: in P. schweinfurthii, pair I (the
gene pool have revealed high karyotype similarity longest) and pair IV present distinctly different physical
(Fig. 13.2) with identical relative lengths of each locations with P. glaucum. Despite having the same
chromosome pair between these species (Khalfallah basic chromosome number, these species are separated
et al. 1993). The position of secondary constrictions into different gene pools presenting important differ-
(SC) is, nevertheless, different among cultivars. ences in their chromosomal organization and locations
Indeed, the majority of stocks that have been studied of their rDNA loci (Fig. 13.3). These observations may
222 T. Robert et al.

Fig. 13.2 Metaphase plate

(a) and karyotype (a0 ) of a b
Pennisetum glaucum (Ligui),
P. violaceum (b and b0 ) with
B-chromosomes (arrows),
23 DB (c and c0 ) and
Thiotande (d and d0 )

10 µm

a’

m m m m m m-sc sm
b’

m m m m m m sm-sat Bs

c d

c’

m m m m m m sm-sat
d’

m m m m m m sm-sat

show that chromosome rearrangements, such as trans- natural conditions. The A0 genome of P. purpureum
locations, may have occurred during the evolution of has been shown to be highly homeologous to the A
these species, which is consistent with the existence of genome of P. glaucum (Jauhar 1968, 1981), while the
reproductive barriers occurring in these gene pools. donor of the B genome is not known. Meiotic analysis
The forage crop “elephant grass,” P. purpureum, of these hybrids reveals formation of seven bivalents,
has the same basic chromosome number x ¼ 7 and where a maximum of five are the heteromorphic asso-
has been considered until now as the only species in ciation of chromosomes belonging to each parent.
the secondary gene pool. This allotetraploid species There are also seven univalent chromosomes that dis-
(2n ¼ 4x ¼ 28) with A0 A0 BB genomes show strong play irregular segregation causing the formation of
reproductive barriers with P. glaucum, but some par- aneuploid gametes (Jauhar 1981; Techio et al. 2006).
tially sterile male and female triploid hybrids occur in Despite their differences in ploidy level, P. purpureum
13 Pennisetum 223

Fig. 13.3 Idiograms showing

the number and position of
rDNA ribosomal genes
18S–5.8S–26S and 5S in four
Pennisetum species:
P. violaceum, P. mollissimum,
P. glaucum cv. “Massu,” and
P. schweinfurthii

P. violaceum P. mollissimum

P. glaucum cv. “Massue” P. schweinfurthii

18S-5.8S-26S 5S

and P. glaucum have almost the same genome size (2C ploid genome, which is similar to P. purpureum. This
¼ 4.59 pg and 4.71 pg, respectively) but quite different observation is consistent with the hybrid zygotes
monoploid genome size (1Cx-value), 1.15 pg and obtained by crossing tetraploid pearl millet with
2.35 pg, respectively. The chromosomes of P. purpur- P. squamulatum (Marchais and Tostain 1997) and
eum (mean chromosome size of 80 Mbp) are half the the success recombining germplasms from P. glaucum
size of the chromosome of P. glaucum (mean chromo- (pearl millet), P. purpureum, and P. squamulatum
some size of 165 Mbp) showing important chromo- (Dujardin and Hanna 1984).
somal changes linked to divergence of these species. According to the evidence detailed above, P. squa-
Another Pennisetum species may be considered to mulatum and P. schweinfurthii are the only tertiary
have x ¼ 7 as its basic chromosome number. gene pool species with x ¼ 7 and belong to the same
On cytological observations, P. squamulatum was initi- morphological section, Heterostachya.
ally reported as an allohexaploid species with x ¼ 9
and 2n ¼ 6x ¼ 54 chromosomes (Raman et al. 1959;
Patil et al. 1961). However, despite the multiple 13.1.2.2 Species with x = 5, 8, or 9 as Basic
reports of 54 chromosomes in P. squamulatum, recent Chromosome Number
studies reveal 56 chromosomes (Goel et al. 2003).
Using a fluorescent in situ hybridization method with All other species of the genus Pennisetum present
centromeric and rDNA probes, Akiyama et al. (2006) basic chromosome numbers of x ¼ 5, 8, or 9. They
observed 56 centromeric signals and detected eight are reproductively highly isolated from the pearl millet
chromosomes with rDNA loci in different accessions and belong to the tertiary gene pool.
of P. squamulatum. These results tend to confirm the P. ramosum belonging to the Gymnothrix section is
octoploid nature of this species (Akiyama et al. 2006) the only species with x ¼ 5 and has a 1Cx ¼ 2.02 pg
with 2n ¼ 8x ¼ 56 chromosomes and a basic number DNA per haploid chromosome set (Martel et al. 1997).
of x ¼ 7. According to this data, the genome size of the P. mezianum is also from Gymnothrix section with
species, with 9.56 pg/2C, has 1.19 pg DNA per mono- x ¼ 8 and presents the lowest monoploid genome
224 T. Robert et al.

Table 13.2 Chromosome number, genome size, and base composition for 15 Pennisetum species representing the taxonomic
sections (Stapf et Hub) and gene pools (Harlan and Dewet) classifications. Adapted from Martel et al. (1997)
Basic Species Section Pool Ploidy level 2C 1C DNA 1Cx- Mean GC
chromosome (chromosome DNA value (in valueb chromosome (%)
number (x) formula) value Mbp) (pg) size (in Mbp)
(pg)a
5 P. ramosum Gymnothrix III 2n ¼ 2x ¼ 10 4.04 1,976 2.02 395 43.7
(8m þ 2sm)
7 P. mollissimum Penicillaria I 2n ¼ 2x ¼ 14 4.51 2,205 2.25 315 46.5
P. violaceum Penicillaria I 2n ¼ 2x ¼ 14 4.52 2,210 2.26 316 47.2
(12m þ 2sm-sat)
P. glaucum Penicillaria I 2n ¼ 2x ¼ 14 4.71 2,303 2.35 329 44.9
(12m þ 2sm-sat)
P. purpureum Penicillaria II 2n ¼ 4x ¼ 28 4.59 2,245 1.15 160 44.3
P. schweinfurthii Heterostachya III 2n ¼ 2x ¼ 14 4.97 2,430 2.48 347 44.0
(12m þ 2sm-sat)
P. squamulatum Heterostachya III 2n ¼ 8x ¼ 56 9.56 4,675 1.19 166 44.1
8 P. mezianum Gymnothrix III 2n ¼ 4x ¼ 32 3.01 1,472 0.75 92 43.0
9 P. alopecuroides Gymnothrix III 2n ¼ 2x ¼ 18 1.90 929 0.91 103 43.5
P. hohenackeri Gymnothrix III 2n ¼ 2x ¼ 18 1.69 826 0.84 92 42.7
(18m)
P. setaceum Eu-pennisetum III 2n ¼ 3x ¼ 27 2.78 1,359 0.93 101 43.6
2n ¼ 6x ¼ 54 5.28 2,582 0.88 96 43.9
P. orientale Heterostachya III 2n ¼ 4x ¼ 36 3.77 1,844 0.94 102 44.6
P. villosum Eu-pennisetum III 2n ¼ 4x ¼ 36 3.47 1,697 0.87 94 43.6
P. pedicelletum Brevivalvula III 2n ¼ 6x ¼ 54 5.61 2,743 0.94 102 43.7
P. polystachyon Brevivalvula III 2n ¼ 6x ¼ 54 5.66 2,768 0.94 103 44.7
a
1 pg ¼ 978 Mbp (Doležel et al. 2003)
b
1 Cx-value: DNA content of monoploid genome (Greilhuber et al. 2005)

size of the genus: 1Cx ¼ 0.75 pg. Its close phyloge- P. purpureum and P. squamulatum, suggesting that
netic and morphological relationship with P. ramo- these three may have a common origin. Close relation-
sum, which has x ¼ 5, large chromosomes and a ships were also observed among species within the
large genome size, suggests that major chromosomal Brevivalvula section (Schmelzer and Renno 1999).
repatterning may have occurred during the divergence Basic chromosome number evolution within the
of P. ramosum, with an increase in the amount of genus was recently assessed by molecular phylogenies
DNA. The other Pennisetum species with the basic using the internal transcribed spacer (ITS) sequences
chromosome number x ¼ 9 (Table 13.2) have a similar of the ribosomal RNA locus (Martel et al. 2004) and
amount of DNA per monoploid genome, independent chloroplast DNA sequence data (rpl16, trnL-F region;
of their ploidy levels (from 0.85 to 0.95 pg). Donadio et al. 2009).
The phylogenetic relationships observed are con-
sistent with an ancestral x ¼ 9 small chromosome
13.1.2.3 Phylogeny and Dysploidy characters for the genus Pennisetum (Fig. 13.4a). Spe-
cies with the derived basic chromosome numbers
The genomic relationships between Pennisetum spe- x ¼ 5, 7, or 8 appear in the most recently divergent
cies have been analyzed using molecular approaches, clades, indicating that the genome structure in Penni-
through variations in chloroplast DNA (Clegg et al. setum might have evolved towards a reduced chromo-
1984; Renno et al. 2001), mitochondrial DNA some number (decreasing dysploidy) and an increased
(Chowdhury and Smith 1988), or comparisons of chromosome size (Martel et al. 2004). Therefore,
repetitive DNA sequences (Ingham et al. 1993). instead of an ascendant dysploidy (Jauhar 1981;
Significant relationships have been noticed between Jauhar et al. 2006), we propose a chromosome evolu-
the genomes of P. glaucum and the wild species tion trend in the genus Pennisetum via a decreasing
13 Pennisetum 225

dysploidy mechanism from a common ancestral pro- P. glaucum cultivars. The x ¼ 7 character observed
genitor with basic chromosome number x ¼ 9. in this clade is derived from an ancestral x ¼ 9 status.
The evolutionary reconstruction is congruent with The species of the tertiary gene pool with x ¼ 7,
the gene pool classification and in agreement with P. schweinfurthii and P. squamulatum, form a highly
some morphological grouping. Species of the primary supported clade but are not closely related to the other
gene pool are clustered together with a bootstrap sup- x ¼ 7 species (Fig. 13.4a). This indicates that genomes
port of 100%. The allotetraploid species P. purpureum with x ¼ 7 evolved twice within the Pennisetum genus.
representing the secondary gene pool of pearl millet The derived species P. mezianum with x ¼ 8 small
form a highly supported monophyletic clade with chromosomes and P. ramosum with x ¼ 5 large

a Eragrostis dielsii

Coix sp.

Sorghum nitidum
97
29 Zea mays
100
42
Tripsacum australe

Digitaria ciliaris

Setaria parviflora
a
P. polystachyon
B
P. pedicellatum*
56
8 a
P. setaceum
E
a
P. villosum
a, s
P. ramosum
92 G
23 84 a
P. mezianum
7
a
Cenchrus ciliaris
s, a
P. hohenackeri
100 G
11 s
P. alopecuroides
51
63 5 a
P. squamulatum
13 99
s H
9
P. schweinfurthii
s
P. purpureum
s
P. g. ssp. monodii,
100
mollissimum
15 s P
P. g. spp. glaucum
69 100 cv Massue
perennial 9 8 s
P. g. spp. glaucum
annual / biannual cv Souna
s
P. g. ssp. monodii,
violaceum
a H
P. orientale

Fig. 13.4 (continued)

226 T. Robert et al.

b Eragrostis dielsii

Coix sp.

Sorghum nitidum

Zea mays

Tripsacum australe

Digitaria ciliaris

Setaria parviflora

P. polystachyon
B
P.
pedicellatum*
P. setaceum
E
P. villosum
x=5
x=9 P. ramosum
G
x=8
P. mezianum

Cenchrus ciliaris

P. hohenackeri
G
P. lopecuroides

x=7 P. squamulatum
H
P. schweinfurthii

P. purpureum
P. g. ssp. monodii,
x=7 mollissimum
P. g. spp. glaucum cv P
x=9 Massue
P. g. spp. glaucum cv
increase in Souna
genome size P. g. ssp. monodii,
violaceum
P. orientale H

Fig. 13.4 Character evolution traced with ITS phylogeny. The (B), Gymnothrix (G), Heterostachya (H), and Eu-Pennisetum
strict consensus tree of the 18 most parsimonious trees from (E). Bold numbers above branches are bootstrap values (%)
parsimony analysis was generated on ITS rDNA sequences from from 1,000 replicates of the PAUP analysis. Bold numbers
16 Pennisetum species and seven outgroups. (a) Sexual expres- under branches are the number of characters state changes form-
sions optimized onto the consensus tree, with s and a: predomi- ing the corresponded consensus branches. Asterisk: Inclusion of
nant sexual and apomictic mode of reproduction, respectively. P. pedicellatum as sister species of P. polystachyon is given by
(b) Chromosome number optimized onto the consensus ITS tree. the ITS2 analysis (bootstrap value ¼ 99%)
Morphological sections are given: Penicillaria (P), Brevivalvula

chromosomes are closely related in a same clade, hav- increase in the amount of DNA during the divergence
ing evolved from an ancestral x ¼ 9 status. The evolu- of P. ramosum (Fig. 13.4b).
tive divergence of P. mezianum may be associated with These molecular phylogenies indicate that the
the loss of one chromosome pair while major chromo- genus Pennisetum is paraphyletic (Martel et al. 2004;
somal rearrangement may have occurred with an Donadio et al. 2009) with the species of Cenchrus
13 Pennisetum 227

ciliaris nested within it, as previously found by other Different types of Bs were observed by Subba Rao
authors (Gómez-Martı́nez and Culham 2000; Giussani and Pantulu (1978) and Jayalakhsmi and Pantulu
et al. 2001; Doust and Kellogg 2002; Doust et al. (1984): “Standard” Bs, which have no effects on
2007). mean A-chiasma frequency but which do affect its
Close genomic relationships have been observed variance; “Deficient” Bs, whose presence is associated
between the aposporous apomictic species Buffelgrass with a depressing effect; and “Iso-Bs,” which are
C. ciliaris (syn. Penissetum ciliare L. Link) and associated with enhanced phenotypes. They suggested
P. squamulatum. These species, used in programs to that a balance between the amount of heterochromatin
introgress apomixis from wild relatives into major and euchromatin present in the B-chromosomes is
crops species, showed almost complete macrosynteny responsible for these different effects.
at the apospory-specific genomic region (ASGR) Subba Rao (1980) studied the transmission patterns
governing apomixis in both species (Goel et al. 2006). of these three types of B-chromosome, using crosses
Species of the genus Cenchrus present basic chro- of a B-chromosome donor line and a male sterile line
mosome numbers of x ¼ 9 and 17, with the x ¼ 17 followed by recurrent backcrossing. The three types
species occurring in a most recently divergent posi- of Bs exhibit an accumulation trend in the oosphere,
tion. These x ¼ 17 species could be derived from an but this trend was not observed in the pollen grain,
ancestral tetraploid form with x ¼ 9 (Avdulov 1931) except for standard Bs. Indeed, trend of B-chromo-
through direct loss of one pair of chromosomes, or by some elimination is more frequently observed in trans-
fusion and loss of part of a chromosome containing a mission via the male gametophyte (see Jauhar 1981 for
centromere (Darlington 1956). a comprehensive review on B-chromosomes in pearl
The chromosome numbers in C. ciliaris vary from millet up to the early 1980s).
18 to 56 (Visser et al. 1998). Most genotypes are tetra- The fascinating behavior, origin, and the transmis-
ploid (2n ¼ 4x ¼ 36) with a genome size of approxi- sion patterns of Bs were subsequently studied in
mately 1,500 Mbp/C (Roche et al. 2002), which is crosses involving wild relatives of pearl millet.
roughly the same as the tetraploid species P. mezianum
(1,472 Mbp/C; Table 13.2) closely related to C. ciliaris
in the ITS phylogeny (Fig. 13.4a, b). These chromo- Behavior of B-Chromosomes in Confrontations
somal and genomic characteristics combined with Between “Wild and Domesticated” Genomes
phylogenetic relationships favor of the inclusion of
Cenchrus species within the genus Pennisetum. Accord- B-chromosomes were described by Amouroux-Pezas
ingly, we propose to reconsider the taxonomic position (1985) in a population of P. violaceum, the wild ances-
of Cenchrus species and to rename them as Pennisetum tor of pearl millet, originating from Niger. The behav-
species as they were previously known. ior of Bs was studied in crosses and recurrent
backcrosses of this P. violaceum population with
pearl millet lines not carrying Bs (Ligui originating
13.1.2.4 B-Chromosomes from Chad, Thiotande from Senegal and 23 DB from
Tifton, USA). In F1 and segregating backcross (BC)
The occurrence of B-chromosomes (Bs) in Pennise- populations, a wide range (0–13 per mitotic cell) of B-
tum typhoides (Burro.) S. & H. (now P. glaucum) was chromosomes was observed. These Bs like those
untzing (1958) and Pantulu (1960).
first reported by M€ described previously are transmissible both through
Venkateswarlu and Pantulu (1970) described the cyto- pollen grains and megaspores. A correlation between
logical behavior of these supernumerary chromo- their occurrence and the variability of some morpho-
somes. The Bs associate to form bivalents, trivalents, logical traits was also reported. In a subsequent study
quadrivalents, or multivalents. Their impact on meio- using inbred progenies (BC1 first and second genera-
sis was outlined by Pantulu and Manga (1975) who tion of selfing: BC1S1, BC1S2), Khalfallah (1990) con-
found that a reduction of mean chiasma frequency on firmed the high intra and interindividual variability in
A chromosomes is observed when the number of Bs B numbers both in mitotic (Fig. 13.5a, b) and meiotic
reaches a threshold of 4, but that the variance of mean (Fig. 13.6) cells. In meiosis of P. violaceum (wild
A-chiasma frequency increases with the number of Bs. parent bearing B-chromosomes), the Bs occurred as
228 T. Robert et al.

Fig. 13.5 Eight (a, arrows)

a b
and five (b, arrows) Bs in
mitotic metaphase cells of
hybrid individual (Ligui 
violaceum). Presence of B-
chromosome in androgenetic
doubled haploid plantlets of
pearl millet (c and d, arrows).
Adapted from Khalfallah et al.
(1993) (a and b) and Le Thi
et al. (1994) (c and d)

c d

10 µm

univalents, bivalents (Fig. 13.6a), trivalents rate has led to differing rates (high and low) within
(Fig. 13.6b), and tetravalents (Fig. 13.6c), while in accessions (Puertas et al. 1986, 1993). Furthermore, the
BCS2 progenies (Thiotande  violaceum) above a inheritance of transmission rate of B-chromosomes has
threshold of 4 Bs frequently formed multivalents com- been elucidated in both maize (Zea mays subsp. mays)
posed of 5 or 6 B-chromosomes (Fig. 13.6d–f). The and rye (Chiavarino et al. 1998, 2001; Puertas et al.
persistence of interchromosomic connections (Fig. 13.6g) 1998; Puertas 2002). In maize, the genes involved in
between A and Bs chromosomes was also observed. Bs transmission rate are located on the A chromosome
These connections could be due to shared sequences set. The accumulation of Bs resulting from non-dis-
between A- and B-chromosomes acting as end-to-end junction events during meiosis is ruled by a process
recognition sites. involving A chromosome centromeric region activity
It is also notable that the behavior of Bs is dependent (Han et al. 2007). In rye, the genes influencing
on the recipient cultivated genotype. Indeed, within B-chromosome transmission rate are located on the
Ligui derived families, the mean number of Bs is higher Bs themselves. These genes are also active in the
in BC1S1 progenies than in BC1S2 (Fig. 13.7a). In process of chiasma initiation. Accordingly, the differ-
contrast to this trend, no significant difference was ences in transmission rate of Bs in progenies derived
observed in mean B numbers between BC1S1 and from crosses between forms of the P. glaucum primary
BC1S2 (Fig. 13.7b) derived from the Thiotande  vio- gene pool (wild relative  domesticated) could reflect
laceum cross. a genetic differentiation between pearl millet cultivars.
This result is consistent with a growing number of Khalfallah (1990) hypothesized the occurrence of a
findings demonstrating that Bs now cannot be consid- differential “accumulation/loss” (A/L) mechanism in
ered only as selfish or parasitic DNA as stated by some recipient pearl millet line.
authors (Östergren 1947; Kayano 1961; Nur 1977; The Ligui and 23 DB lines exhibit an early B (A/L)
Jones 1985; Shaw and Hewitt 1985). Indeed, in rye figure, resulting in a reduction of B number from
(Secale cereale), a genotype-dependent B-transmission the BC1S1 generation, whereas in the Thiotande line
13 Pennisetum 229

a b c

d e

10 µm

f g

Fig. 13.6 B-chromosome behavior in meiosis: In P. violaceum, six (f) chromosome Bs do not follow the chromosomes A during
the Bs form two bivalents (a), one trivalent (b), or one tetrava- meiosis and were probably eliminated from cell at the stage
lent (c); in the progenies BCS2 (Thiotande  violaceum) of telophase I. Persistence of interchromosomal connections
B-chromosomes frequently form a star shape multivalent (g, arrow) between A and Bs chromosomes (Original data)
(d, diacinese, arrow). The multivalent composed by five (e) or

genetic background, this mechanism generates a high is consistent with previous genetic abnormalities (male
number of Bs in late BCS generations. Conversely, the sterility and segregation distortion) observed in the
“elimination” component of the process sets in from a offspring of Wild  Thiotande crosses (Marchais and
threshold number of Bs. Indeed, in the BCS2 progenies Pernès 1985; Sarr and Pernés 1988; Sarr et al. 1988).
(Thiotande  violaceum), B-chromosomes form a
visible star shape multivalent in telophase I of meiosis
(Fig. 13.6e, f) and are probably eliminated from the The Effect of B-Chromosomes
cell at this stage. This mechanism, which was shown
for the first time by Khalfallah et al. (1988) and In pearl millet, the occurrence of B-chromosomes
Khalfallah (1990), probably underlies the regulation could interfere with plant fertility. Indeed, the drive
of B number in the confrontation of genomes from mechanism described above as “accumulation/loss”
wild relatives and domesticated forms. The peculiar induced perturbations of meiosis (Khalfallah 1990).
expression of the A/L mechanism in the Thiotande line The same effect was observed in many animal and
230 T. Robert et al.

diversity and a higher number of Bs. Actually, the

global pattern indicate a closer genetic similarity
between wild parent and the selfed BC progenies. It
is worth mentioning that in BC1S3 progenies, the
phenotypical diversity is still high and the recovery
of domesticated like progenies is also weak. This
pattern is specific to crosses involving P. violaceum
(carrying Bs) from Niger as male parent. Indeed, in
progenies derived from crosses between P. mollisi-
mum, another wild relative (without Bs), and the
same domesticated lines, the recovery of cultivated
type in BC progenies is easier (Joly-Ichenhauser
1984; Joly-Ichenhauser and Sarr 1985; Niangado
1981). These original data laid prospects as suggested
by Jones et al. (2008) for investigations on genome
organization and the dynamics of biodiversity with
Bs as linchpin specifically around their supposed
origin from interspecific hybridization or genome
response to both biotic and abiotic environmental
stresses. For example, some specific environments
like in vitro tissue culture could induce chromosome
abnormalities and the apparition of B-chromosome.
Indeed, in doubled haploids plantlets obtained
from androgenesis of pearl millet (Fig. 13.5c, d),
B-chromosome was observed (Le Thi et al. 1994).
This phenomenon was also reported for Nicotiana
sylvestris from in vitro culture (De Paepe et al.
Fig. 13.7 Variation of B-chromosomes within inbred families 1983; Lespinasse et al. 1987).
(BC1 first and second generation of selfing: BC1S1, BC1S2);
Ligui  violaceum BC1S1 and BC1S2 (a) Thiotande  viola-
ceum BC1S1 and BC1S2 (b) (Original data)

13.1.3 Domestication Process Hallmarks

in the Genus Pennisetum
plant species; in Dactylis (Williams and Barclay 1968)
and rye (Jones and Puertas 1993), for example.
13.1.3.1 Where, When, and How Pearl Millet
Amouroux-Pezas (1985) assumed that the presence
(P. glaucum) Was Domesticated?
of Bs could stimulate plant growth at early phenologi-
cal stages (larger and higher number of leaves), but a
Origin of Pearl Millet
depressing effect is observed during heading (narrow
flag leaves and reduced productive basal tillers).
Center or Non-Center Domestication of Pearl Millet?
In the above mentioned study, Khalfallah (1990)
described the pattern of morphological trait diversity In South America, Southeast Asia, and Subsaharian
in segregating progenies derived from crosses Africa, the high and geographically structured pheno-
between a wild progenitor B donor and domesticated typic diversity that has been observed in numerous
lines empty of Bs (BC1S2 and BC1S3). This analysis crops originated from these areas has led several
(see Fig. 13.8 for BC1S2) revealed high diversity in authors to hypothesize that domestication occurred
terms of recombinant phenotypes for a set of 16 mor- several times for the same species within large geo-
phological characters. The progenies deriving from graphical regions, called non-center of domestication
the line Thiotande as female exhibit phenotypical (Harlan 1971, 1975). The question whether pearl
13 Pennisetum 231

Fig. 13.8 PCA first plan a

showing diversity of 4 BC1S2
progenies and parents (wild
and domesticated lines) (a);
Path discriminant analysis first
plan showing high diversity in
BC1S2 progenies from the
cross Thiotande (Thiotande 
violaceum) (b). (Original
data)

millet, which had been originated from Africa, was in this area of Africa. Classifications of pearl millet
domesticated or not according to a non-center model is landraces has been defined on the basis of the size and
still debated. This review proposes to enlighten this the shape of candels and the color and the shape of
question in regards to old and recent data researches in seeds (Portères 1950; Brunken 1977). Because the
archeology, paleobotanic, and genetics. distribution of the main pearl millet types shows a
clear geographic pattern, it has been supposed that
the domestication of this crop has occurred several
The Beginning of Agriculture in Africa and Pearl
times along the Sahelian region of Africa (Portères
Millet Domestication
1950; Harlan 1975; Brunken et al. 1977). This hypo-
Although pearl millet (P. glaucum ssp. glaucum) is thetical domestication non-center overlaps the actual
cultivated in every arid regions of Africa and South- distribution of the wild millets (P. glaucum ssp. mono-
west Asia, its wild closest relative P. glaucum ssp. dii). The humid climate is not propitious to the conser-
monodii has never been observed outside of the vation of carbonized remains. Conversely, the
Sahelian region where the phenotypic diversity of conditions prevailing in the Sahara desert prevented
pearl millet is the highest. This is one of the strong the formation of humus necessary to trap these
arguments favoring the hypothesis of its domestication remains to protect from oxidation, which have led to
232 T. Robert et al.

a major erosion. The reconstruction of an archeologi- crop farming (Camps 1974; Ballouche and Neumann
cal sequence in relation to the domestication is com- 1995). They are consistent with the abundant lithic
plicated by the scarcity of carbon remains in spite of tools, and footprints of pearl millet seeds that are
the intensification of excavations in recent decades. larger in the domestic form than in the wild Pennise-
However, a general pattern emerges, which may ques- tum, found in two sites dated to 6000 BP (Camps 1974;
tion the assumption of multiple domestication for Klichowska 1978; Fig. 13.9). However, these indica-
pearl millet. tions of the existence of domestic Pennisetum are still
Plant and animal domestication has been a central very tenuous and indirect. However, it seems likely
element of the Neolithic revolution during which agri- that when the period begins to be more arid in Sahara,
culture was invented. Farming in Africa probably people who migrated and settled on the banks along
started in the area, which corresponds to the actual the Nile and in the Sahel are likely to have started
Sahara desert. Indeed, around 18000 BP, the establish- agriculture. Probably, pearl millet cultivation was
ment of an intertropical rainy season has erased the already adopted all along the southern edge of the
hyper-arid desert and provides the opportunity for a Sahara, from Mauritania to Sudan, as early as 3500 BP.
wooded savannah to develop in the south part and a Fields and granary were numerous at Dhar Tichitt and
more Sahelian vegetation in the North (Adams 1997). Dhar Oualata in the actual Mauritania, where among
A dense hydrographic network of rivers and lakes was the most old domestic pearl millet remains have been
established, around which the density of human occu- found (Amblard and Pernès 1989). The most old pearl
pation has continued to increase until the middle of the millet seeds, found in northern Togo, have also been
Neolithic, through the colonization of the Sahara by dated to 3500 BP (Birimi) (Fig. 13.9; D’Andrea et al.
people from South (Camps 1974; Petit-Maire 2002). 2001; D’Andrea and Casey 2002). The Metallurgy was
However, the climate was dry enough, locally vari- already known in central Sahara as early as 3500 BP and
able, and sometimes caused droughts particularly in has spread after that to the whole of West Africa,
the eastern regions where the Atlantic monsoon was therefore, contributing to an improvement of agricul-
short (Wendorf et al. 1992). The climate became pro- tural practices and to the development of Bantu’s
gressively degraded from 6000 BP, and the formation people in the region corresponding to actual Nigeria
of the Sahara was achieved around 3500 BP. and North Camerun and to their demographic and
Until 8000 BP, Saharian people were exclusively geographic expansion (Cavalli-Sforza et al. 1994;
hunter-gatherer. Ceramics was used from 9000 BP in Bellwood 2001).
Niger, namely 2,000 years earlier than in the Middle However, up to date, no trace of undoubtedly
East according to the current state of knowledge domestic forms of pearl millet have been found in
(Fig. 13.9). Harvesting of wild grasses, especially the former sedimentary strata in these sites except on
Panicum, Digitaria, Pennisetum, and Sorghum spe- the shores of Lake Chad where pearl millet is never-
cies, was intensive as witnessed by the numerous lithic theless abundant from 3000 BP (Zach and Klee 2003;
tools found in archeological investigations of this Klee et al. 2004).
period and by the remains of seeds conserved in This gap in the sedimentary sequence indicates that
tanks used as granaries and dated as early as 9000 BP the domestication of millet could have taken place
(at Nabta Playa) (Wendorf et al. 1992; Wendorf and elsewhere, perhaps further north under the pressure
Schild 1998). Archeological reconstructions show that of the arid conditions, where wild populations were
agrarian societies using the Neolithic way of life in number (Wetterstrom 1993). Because pearl millet
existed on the African continent since at least the was not introduced as a crop in Egypt by eastern
eighth millennium BC. Saharian people who had however migrated there at
The first signs of bovine domestication (Bos Tau- this period, one could imagine that pearl millet domes-
rus) found in the west of Egypt and then in the Central tication was restricted to the west-center region of the
Sahara are dated from the same period (Wendorf and Sahara. The fact that pearl millet was already grown as
Schild 1994; Fig. 13.9). From the seventh millennium a crop in India as early as 3000 BP (Fuller et al. 2004) is
BP, the palynologic studies from this period indicate a also an argument in favor of an older domestication of
rapidly changing landscape due not only to pastoral this species. Indeed, the geographic distribution of the
practices but also clearing, which is usually linked to wild form of P. glaucum is restricted to the Sahelian
13 Pennisetum 233

Fig. 13.9 Archeological evidences of domestication in Subsaharian Africa

region of Africa so that the presence of this crop region (Tostain et al. 1987; Tostain and Marchais
species in Asia is the result of a very ancient diffusion 1989; Tostain 1992, 1993). However, genetic differ-
outside Africa. entiation between domestic and wild pearl millet could
It is obviously impossible to assess whether pearl be under-evaluated with this kind of molecular marker
millet was domesticated once or several times from since isoenzyme polymorphism is generally low.
archeological data only. So, genetic tools have also More recent data have been obtained with micro-
been used with the hope to shed light on the domesti- satellite markers, which are much more polymorphic
cation history of pearl millet. on a sample representative of the Sahelian and
Soudano-Sahelian region (Oumar et al. 2008). The
results have shown that only 74% of the genetic diver-
Contribution of the Genetic Tools in Order
sity of the wild millets is found in the domestic groups.
to Enlighten the History of Pearl Millet Domestication
This suggests that the domestication of pearl millet has
Huge collections of domestic and wild forms of pearl led to a decrease in the effective population size and
millet from the whole area of cultivation of this crop thus to a bottleneck effect. Nevertheless, this decrease
have been analyzed for the allozyme polymorphisms in pearl millet genetic diversity due to the domestica-
(Sandmeier et al. 1981; Pilate-André 1992; Tostain tion may be stronger than suggested by this data.
1993). These studies have shown that the genetic The gene flow between domestic and wild forms
diversity was similar in the forms of pearl millet and could later have contributed to the increase of the
that the genetic diversity of wild millets displays a genetic diversity of the domestic gene pool. The sim-
clear East/West geographic pattern in the Sahelian ple sequence repeat (SSR) polymorphism has also
234 T. Robert et al.

revealed a high genetic differentiation between ment length polymorphism (RFLP) analyses on the
domestic and wild populations of pearl millet nuclear adh1 gene also showed a parallel pattern in
(Oumar et al. 2008) (Fst ¼ 0.10), which was higher domestic and wild forms of pearl millet (Gepts and
than the level of genetic differentiation that has been Clegg 1989). However, the RFLP analysis on the
found between maize and teosinte, its wild ancestor. nuclear rDNA gene revealed a contrasted pattern
The analyses of the isoenzyme polymorphism on since the diversity in the domestic sample was
domestic and wild samples have yielded two main extremely low while it was high in the wild sample.
results. Firstly, the varieties from the Subsaharian According to these data, the domestication events
region, the likely area of domestication of this crop of early-flowering pearl millet could have been fol-
according to archeological data, are clearly distinct lowed by a strong secondary diversification in an area
from the East African varieties. Moreover, two main located at the west side of the actual Lake Chad
regions within the Subsaharian region can be defined on (Fig. 13.10). Late flowering, new early-flowering vari-
the basis of population differentiation (Fig. 13.10). East eties, and pearl millet varieties of East and South
Africa cannot be considered as a domestication center Africa and of India could have originated from this
on the basis of archeological data and because wild secondary center of diversification. Indeed, the low
forms of pearl millet are totally absent from this region. genetic distances between late-flowering varieties
Secondly, a more in-depth analysis considering domes- and early-flowering varieties from this region argue
tic and wild samples from the west part of the Subsa- in favor of this scenario as well as the low genetic
harian region shows that domestic and wild forms from diversity of late-flowering varieties relatively to early-
the same area clustered together (Pilate-André 1992). flowering ones (Tostain et al. 1987; Tostain and
Genetic distances between populations are dependent Marchais 1989; Tostain 1992, 1993).
on the geographic distance rather than on the taxo- More recent data obtained on the basis of SSR
nomic status. This has been considered as an argument polymorphisms (27 loci) within a sample of domestic
in favor of several events of domestication for this crop and wild pearl millet covering the entire geographic
in this region according to a non-center model (Pilate- distribution area of this species in Africa and India has
André 1992) as proposed by Portères (1950) and by shown a different pattern. Bayesian analysis of the
Harlan (1975). This is congruent with previous results SSR polymorphisms, taking into account the estimated
obtained on West African populations (from Senegal to proportion of domestic/wild admixture within the
Niger) (Tostain et al. 1987). Data from restriction frag- domestic sample suggests a single domestication center

a b c
Senegal-wild Sudan-wild
Senegal-dom. Niger-wild
Niger-dom. West-Chad-wild
Niger-dom. West-Mali-dom.
Cameroon-dom. Centre-Sahel-wild I II
Mali-dom. West-Sahel-wild
III
West-Sahel-early-dom. III
Mali-dom.
Burkina-dom. Togo-Ghana-early-dom.
Burkina-wild I: Domestication III
Niger-wild East-Sahel-dom. II: Diversification
Niger-wild West-Sahel-late-dom. III: Diffusion
India-dom
Mali-wild East and South Africa-dom.
Mali-wild
Cameroon-wild Senegal-dom (off season)

Fig. 13.10 Isoenzyme polymorphism in pearl millet. (a) anal- followed by a secondary diversification in a much restricted area
ysis based on 14 domestic (mainly early varieties) and wild (at the west side of the actual Lake Chad) and then a diffusion of
populations from the west part of the Subsaharian region (15 varieties to East and South Africa, probably mainly at Bantu’s
loci). (dom.: domestic accession) (b) Analysis on accessions migration period; (c) more likely hypothesis for pearl millet
from the all areas of pearl millet cultivation in Africa (except domestication and diffusion in Africa. (1) a primary domestica-
Maghreb). These data rather support the hypothesis of several tion (single or multiple events?) in West Africa; (2) center of
domestication primary events of early varieties in a huge non- secondary diversification (late varieties could be originating
center covering the West Sahel. This first phase would have been from this center); (3) diffusion of new early and late varieties
13 Pennisetum 235

for pearl millet located in a region corresponding to the tions located in remote areas of the original domesti-
actual West Niger and Mali. cation center have contributed significantly to the
Although no clear consensus about the history of evolution of these traits in domestic populations.
pearl millet domestication finally emerges either from In other words, different regions of the genomes of
genetic data or from archeological sequences, it domestic pearl millets could, therefore, originate from
seems, however, that the data reported from the most different ancestor wild gene pools and thus they could
recent studies using very informative molecular mar- have different histories. Therefore, the domestication
kers (SSR markers) are in favor of a pearl millet of pearl millet in particular, and perhaps of crops in
domestication localized in a restricted region of West general, can be viewed as a reticulate evolutionary
Africa. This domestication could have been followed process in which several wild gene pools have con-
by a spread of domestic populations in the all Subsa- tributed. If this did really occur for pearl millet, the
harian region. A secondary diversification of domestic reconstruction of the history of its domestication can
populations could have occurred in the eastern part of hardly be achieved using molecular markers. Yet, the
the Sahel (Tostain et al. 1987; Tostain 1992). Pearl identification of genes involved in the domestication
millet from Eastern and Southern Africa and late vari- syndrome and the analysis of their nucleotide poly-
eties are probably originated from this center of diver- morphism pattern among wild and domestic popula-
sification (Fig. 13.10). tions could be a powerful strategy in order to achieve
Inferences of domestication history from genetic this goal.
data are, however, to be made cautiously mainly
because of gene flow that have occurred between
Dynamics of Spontaneous Wild–Weed–Pearl Millet
domesticated and wild populations since the beginning
Complex
of the domestication process and is still operating now
(Pernès 1986). Indeed, since its domestication, pearl Non-intensive agrosystems often display remarkable
millet populations have been spread sometimes far occurrence of wild, weedy, and domestic forms of the
from the area where they have been domesticated. same species in sympatric/paraptric situations. Gene
Cultivated populations can, therefore, be found in flow between domesticated and spontaneous relatives
parapatric situations with other wild populations than has been considered by some authors to contribute
their wild ancestors so that introgression of new favorably to the evolution of crops in these agrosys-
genetic material can modify the original genetic com- tems. Indeed, it can lead to the production of new
position of the domestic gene pool. genotypes upon which farmers’ selection can operate
Another point to be considered is that present to produce enhanced phenotypes (Jarvis and Hodgkin
domestic populations strongly differ from a morpho- 1999) and to the spread of advantageous alleles
logical point of view, from the wild phenotypes. (Morjan and Rieseberg 2004). On the other hand,
Numerous traits contribute to the domestication syn- experimental and theoretical population genetic models
drome (cf. Sect. 13.1.3.2). It is very unlikely that all have shown that strong gene flow can be responsible for
the mutations needed to produce the domestic pheno- a decrease in local adaptation through introgression of
type observed nowadays were present in a unique wild genes from non-adapted genotypes (Antonovics and
population. For example, the increase in seed size in Bradshaw 1970; Slatkin 1987). The impact of gene
domestic pearl millet compared to the wild forms flow from the wild relatives on crop diversity has
seems to have occurred progressively and lately, far been questioned (Wood and Lenné 1997), although in
after the beginning of the domestication, according to some instances, it could produce aggressive weedy
the archeological data (Fuller 2007). A progressive forms, as clearly documented in beet (Boudry et al.
and slow evolution of the loss of shattering ability in 1993) and common bean (Papa and Gepts 2003).
domestic rice, wheat, and barley has also been sug- Areas where domestic species and their spontaneous
gested (Purugganan and Fuller 2009). Moreover, relatives coexist can, therefore, be viewed as in situ
domestic traits often display multifactorial inheritance laboratories for the study of the effect of biological
(see Sect. 13.1.3.2 for the genetic basis of the domes- factors and of anthropologic practices on the level of
tication syndrome in pearl millet). One can, therefore, gene flow and introgression between these different
imagine that several mutations present in wild popula- forms of the same species.
236 T. Robert et al.

Weedy millets (Shibra, N’doul in Haoussa and The very huge amounts of Shibra in numerous pearl
Ouofof language, respectively), with intermediate millet fields in Africa and their presence even in the
domesticated/wild phenotypes are observed in most areas where wild forms do not exist anymore or even
pearl millet fields in West Africa (Brunken et al. have probably never existed raise questions on the real
1977) including those where wild forms are totally contribution of gene flow from wild populations to
absent (Busso et al. 2000). Contrary to the well- domestic millets in order to explain their maintenance
domesticated forms, Shibra forms exhibit an intense and even, in some areas, their expansion. In fact, very
shattering habit at maturity. Their presence in drills few studies have focused on the level of genetic intro-
(holes in the ground made by farmers in which they gression of these weedy plants by genes from wild
generally sow a few dozens of seeds) decreases the or domesticated plants (Marchais 1994; Miura and
production of good quality seeds (large and easy to Terauchi 2005; Mariac et al. 2006).
thresh). Although farmers consider sometimes Shibra The studies of both Marchais (1994) and of Mariac
as invasive plants and are willing to reduce their num- et al. (2006) on the in situ morphological diversity of
bers, they do not eliminate them completely. Indeed, in weedy millets (Shibra) based on the traits defining the
harsh conditions or to bridge the gap before the main domestication syndrome have shown a large continu-
harvests, Shibra are harvested and consumed, even ous phenotypic distribution within pearl millet fields.
though their yield is very low and their seeds very Among weedy millets, plants showing different levels
hard to prepare for consumption. Weeding is supposed of combination of traits from domesticated and wild
to discard all the weeds, including Shibra, from the phenotypes are found, ranging from nearly domesti-
field. However, it is frequent that some Shibra plants cated to nearly wild phenotypes (Mariac et al. 2006).
escape this elimination so that the density of Shibra in However, Shibra that grow in drills are significantly
pearl millet fields after weeding is sometimes still very more similar to domestic plants, which suggests that
high (more than 30 Shibra/100 m2 could be observed; most of them are indeed developed from seeds sown
C. Allinne personal communication). by farmers, and thus, in most cases, from selected
Shibra plants are early maturing plants. Although candels at the harvest time (selection on the female
the flowering of their main tiller is generally very contribution to the next generation). The high diversity
early, their flowering period largely overlaps the flow- observed among Shibra plants and the multi-factorial
ering time of the domestic millet, either early or even inheritance of the traits defining the domesticated phe-
late flowering varieties, because they mostly produce a notypes (Poncet et al. 2000) suggest that Shibra are
large amount of non-synchronous axillary flowering developed from recombination events between the
tillers. They are fully interfertile with domestic and genomes of domestic and wild millets in hybrid plants.
wild pearl millet. The marked phenotypic differences between Shibra
In parapatric situations, weedy plants have been found in drills and those found outside drills (Mariac
considered as derived from recurrent pollen flow et al. 2006) could thus reflect different introgression
between domesticated and wild populations (Marchais levels by wild genes at loci involved in “domestica-
1994; Busso et al. 2000). Their presence in the fields tion” traits. Investigations carried out in several pearl
results from sowing or spontaneous seedlings devel- millet fields in Niger have shown that Shibra pheno-
oped from shattered seeds. In the former case, the types could represent up to 46% of the plants found in
seeds come from spikes selected by farmers to produce drills. However, bulked segregant analysis (BSA) of
seeds for the next growing season. Even when farmers progenies obtained from selfing Shibra plants suggests
are able to select their seeds on plants with the best that a unique region of the genome could be involved
phenotypes (well domesticated), there is no way to as a major factor explaining the morphological differ-
recognize seeds formed from pollination by weedy entiation between Shibra and plants with a truly
and wild plants. Obviously, the dynamics of weedy domestic phenotype (Miura and Terauchi 2005).
populations is dependent on farmers’ practices (espe- These apparently different conclusions are, however,
cially seed management and weeding). The relative in accordance with data from domestication quantita-
impact of these practices and gene flow from wild tive trait loci (QTL) mapping in pearl millet (see
populations on these dynamics would be a key ques- Sect. 13.1.3.2), since a region on the linkage group
tion to address from an evolutionary point of view. (LG) 6 of the pearl millet genetic map harbors a high
13 Pennisetum 237

number of major domestication QTLs, while mostly the product of crosses between Shibra or between Shibra
minor QTLs were detected on other linkage groups and domestic millets. However, selection against domi-
(Poncet et al. 2000). nant phenotypes should lead to a rapid decrease in the
The main question emerging from these data con- frequency of Shibra phenotypes in the absence of gene
cerns the roles of the different factors controlling the flow from wild populations. Quantification of the farm-
abundance and the maintenance of Shibra forms. The ers’ selection procedure is necessary to gain insight into
existence of a strong genetic differentiation between the mechanisms underlying Shibra persistence in fields.
domestic and wild pearl millets at the continental level A sizeable proportion (11.4%) of plants identified
based on neutral polymorphisms has been shown by by farmers to be domesticated was in fact morpholog-
recent studies on SSR polymorphisms (see above). ically closer to Shibra plants found in drills (Mariac
Moreover, genetic analyses using amplified fragment et al. 2006). Seeds originated from candels of Shibra-
length polymorphism (AFLP) markers on plants sam- like plants can, therefore, be unconsciously added to
pled in situations where domestic, Shibra, and wild the seed stock through the harvest of not fully domes-
forms are found in parapatric situations have also ticated phenotypes, and then be sown in the following
shown a high level of genetic differentiation between season. In addition, the harsh agricultural conditions in
wild forms on one part and the domestic on the other most African regions, where pearl millet is cultivated,
(Mariac et al. 2006), even if assignment analyses also lead to chronic deficits in seed production. In Niger,
show the existence of introgression of Shibra plants by for example, farmers are used to get their seeds not
genes from the wild. In addition, analyses of the only from their own production but also from neigh-
genetic composition of the pollen pools, which con- bors, local sellers, or regional markets, where seeds
tribute to fertilize domestic, weedy, and wild forms harvested throughout the region and even other
found in a parapatric situation, also show a clear regions of Niger are sold (Allinne et al. 2008).
genetic differentiation of the latter. A similar situation has been observed in Rajasthan,
The overall picture drawn by these data is in favor India (Vom Brocke et al. 2003), where frequent impor-
of a low occurrence of hybridization between domes- tation of seed lots by farmers explains the very low
tic and wild pearl millets despite mechanisms favoring differentiation between pearl millet populations
potential gene flow between wild and cultivated cultivated in distant villages. It has demonstrated that
(domesticated and weedy plants) populations, such as seeds sold to farmers on the markets are sometimes
outcrossing and phenological similarity, as already developed from non-selected plants, even Shibra. The
suggested by Marchais (1994). contribution of external seed sources could, therefore,
Pre-zygotic (Robert et al. 1991; Renno et al. 1997) or account for the wide presence of Shibra in drills,
post-zygotic (Amoukou and Marchais 1993) isolation which in turn could contribute, through their proge-
mechanisms already documented in pearl millet could at nies, to the seed bank and therefore to produce popula-
least partly explain this mating behavior. Then direct tions of Shibra outside drills. It is, therefore, likely that
hybridization between wild relatives (P. violaceum) and seed flow due to farmer’s strategy to get seeds is the
the domesticated forms could not be considered as the main factor explaining the spread of Shibra plants in
major process explaining the abundance and mainte- pearl millet fields in Sahel.
nance of weedy Shibra in the crop fields. Shibra them- Quantitative genetic models have shown that popu-
selves obviously contribute, through pollination, to lation differentiation can be high at the trait (pheno-
further introgression of traits characteristic of wild typic) level and low at the allelic frequency level for
plants, such as shattering or a high tillering habit, into the majority of QTLs underlying the trait, especially
the selected seeds for next sowing. The intermediate when diversifying selection is strong (Le Corre and
phenotypes of Shibra plants, despite their highest Kremer 2003). Phenotypes such as Shibra found out-
genetic similarity with domestic rather than wild millets, side the drills could be favored by natural selection.
could be explained by the dominance of alleles confer- This probably occurs since Shibra outside the drills
ring the wild phenotype, as it has been demonstrated in generally have a higher shattering ability than plants
QTL studies in pearl millet (Poncet et al. 2000 and also found in drills. They should, therefore, contribute
see Sect. 13.1.3.2). Shibra found outside drills and more than plants growing in drills to the seed bank.
developed from soil seed bank recruitment are mainly Furthermore, progenies developed from these plants
238 T. Robert et al.

will also tend to have a high shattering ability as this of accurate molecular markers (e.g., RFLP, AFLP,
phenotype is dominant in pearl millet (Poncet et al. SSR, SNP) during the last two decades (Liu et al.
1998). Consequently, if this selective advantage of 1994; Devos et al. 1995; Qi et al. 2001; Budak et al.
shattering phenotypes is effective, a hitch-hiking pro- 2003; Bertin et al. 2005). The genomic resources result-
cess may lead to an increase in other wild characters ing from sequencing technologies together with com-
such as candle morphology. This would be reinforced parative mapping studies highlighted the genomic
by linkage of domesticated genes in pearl millet (Poncet organization among species of the Poaceae family
et al. 2002 and see also Sect. 13.1.3.2). (Devos et al 2000; Qi et al. 2004; Senthilvel et al.
2008). In pearl millet like in many other species, these
maps constitute the linchpin of QTL analysis for the
Conclusion
genetic dissection of complex traits. For example, in
The data from genetic studies and investigations on pearl millet, dwarfing genes have been mapped as well
farmers’ practices suggest that the abundance of inter- as QTLs involved in tolerance to biotic or abiotic
mediate phenotypes in fields in southwestern Niger is stresses (see Yadav et al. 2002, 2004 for drought toler-
mainly because of farmers’ practices and especially ance; Jones et al. 1995, 2002; Morgan et al 1998 for
seed flow due to seed exchanges between farmers resistance to pathogens; Yadav et al 2003; Bidinger
rather than the direct pollen flow and hybridization et al 2007 for enhanced productivity under diverse
between domesticated and wild pearl millets. It is environments). Marker-assisted selection has been
well known that seed flow due to seed exchanges employed to improve efficiency of pearl millet genetic
between farmers regularly increases for several breeding schemes (Hash et al. 2003; Khairwal and
years. Moreover, farmers have no means or even will- Hash 2007). As far as wild relatives of pearl millet
ingness to specifically eliminate Shibra in drills. This are concerned, two main domains using evolutionary
provides an opportunity for reinforcing seed banks genomics and molecular genetics approaches have
through shattering plants and could lead to long-term been addressed: dissection of domestication process
persistence of Shibra plants in the fields. Thus, Shibra and interspecific hybridization (using tertiary gene
plants, rather than wild ones, provide a regular source pool species, P. squamulatum and C. ciliaris, for exam-
of traits from the wild to be further introgressed in ple) for transferring single genes of interest or unravel-
domesticated phenotypes. However, it is difficult to ing genetic basis of complex traits like apomixis
really appreciate, through the evocation of the farmers (Ozias-Akins et al. 1998, 2003; Conner et al. 2008).
whether we are facing a new and evolving process or
whether this situation corresponds to a state of
dynamic equilibrium that has lasted for decades. In The Genetic Basis of Pearl Millet Domestication
the first hypothesis, the process of homogenization of
pearl millet populations through gene flow could The QTL analysis and evolutionary genomic studies
result in a rapid degradation of the level of adaptation (Doebley and Stec 1993; Poncet et al. 2000, 2002)
of local varieties. Hence, it is crucial to analyze the have generated a strong body of knowledge on domes-
role of changes in farmers’ practices and the economi- tication process. These findings laid the foundation to
cal and ecological constraints in order to better under- conceptual inferences tracing back to the origin of
stand the evolution of the pearl millet crop/weed domesticated plants and the role of evolutionary forces
complexes. including selection (both natural and anthropogenic),
mutation, gene flow, and genetic drift (founder effect)
on the success of domestication process and the
dynamics of agrobiodiversity.
13.1.3.2 Evolutionary Genomics and the Wealth This section focuses on the current understanding of
of Wild Relatives for Deciphering the Genetic the domestication process in Pennisetum gene pool based
Basis of Quantitative Traits in Pearl Millet on genetics and genomic investigations (see Sect. 13.1
for general frame of pearl millet domestication).
The genetic maps of many cereals are becoming The perusal of literature on the analysis of genetic basis
increasingly dense, thanks to the dramatic development of domestication syndrome (i.e., traits differentiating
13 Pennisetum 239

domesticated forms from their wild progenitors as result- Table 13.3 List of morphological traits used for QTL mapping
ing from the effects of the evolutionary forces mentioned Code Units Trait description
above) clearly indicates that a few genes have been Traits measured 1 month after sowing (juvenile stage)
recruited during the domestication process. This figure NL Number of leaves on the
primary tiller (shoot)
holds for the main cereal species (maize, rice, wheat)
NT Number of basal tillers
despite differences in the architecture of their genomes LS cm Length of the sheath from base
and their mode of domestication (see modes of domesti- to the first leaf, on the
cation scheme by Small 1984; Hillman and Davies 1990; primary tiller
Doebley and Stec 1993; Dorweiler et al. 1993). Studies on DS mm Diameter of the sheath
pearl millet domestication process are consistent with this LL cm Length of the fourth appearing
leaf
general trend. WiL mm Width of the fourth appearing
leaf
TNL Total number of leaves
Genetic Dissection of Domestication Syndrome Flowering time
Traits Head Days after sowing Days to heading of the primary
tiller
Pearl millet, P. glaucum ssp. glaucum (L.) R. Br., HHead cm Plant height at first heading date
PI (BMF-BFF)/(DO) Protogyny index (with BFF:
among the species of the genus Pennisetum, is the
beginning of female
only domesticated one. This species is one of the flowering, BMF: beginning
main staple cereal crops of the semi-arid regions of of male flowering on the
Africa and India. The wild forms of pearl millet, primary tiller, DO; duration
of the female flowering)
P. mollissimum and P. violaceum (ecotypes of
Traits measured at maturity
P. glaucum ssp. monodii), are distributed only in Africa,
NNTP Number of nodes on the primary
where they have been involved in the domestication of
tiller
the crop for several thousand years (Brunken et al. 1977; NS Total number of spikes/plant
Appa Rao and de Wet 1999; see also Sect. 13.2). The NTM Number of basal tillers
main traits composing the domestication syndrome in Hmax cm Plant height
Pennisetum are listed in Table 13.3. As for many cer- HPT cm Height of the primary tiller
eals, spike “gigantism” and non-shattering of the spike- LLM cm Length of the fourth leaf at
maturity
lets at maturity are among the key traits. The wild
WiLM mm Width of the fourth leaf at
relative forms of pearl millet are still found in sympatric maturity
or parapatric distribution with the cultivated form in Traits measured on the primary mature spike
some areas of the Sahelian regions of Africa. Both WeS g Weight of the spike
P. glaucum and its two wild relatives (P. mollissimum LoS cm Length of the spike
and P. violaceum), mainly outcrossers, are fully inter- WiS cm Width of the spike
fertile and have the same genome size (see Sect. 13.1.2 PL mm Pedicel length of the floral
on Chromosome evolution). This peculiar situation in involucre measured from the
abscission layer (or rachis)
the pearl millet gene pool offers a remarkable frame- to the base of the involucre
work for addressing the evolutionary questions about the BL mm Length of the involucre bristles
number of genes involved in the domestication process Ct Scaling points Seed coating (exposed seeds as
and their genomic organization. (0 as cultivated the cultivated phenotype or
The inheritance of domestication traits distinguish- phenotype) enclosed seeds as the wild
phenotype)
ing pearl millet (P. glaucum) from one of its wild AL Presence (1) or not Functional abscission layer
relatives viz. P. mollissimum has been assessed by (0)
several authors (Joly-Ichenhauser 1984; Pernès PB Presence (1) or not Existence of a longer bristle
1986). For example, Poncet et al. (1998) tackled this (0)
GL mm Length of the upper floret
issue in F2 progenies derived from a cross between a lemma
typical early flowering landrace of pearl millet (Souna
240 T. Robert et al.

from Mali) and a wild ecotype P. mollisimum near hypothesized that two chromosome segments (segment
Gao, Mali. I and segment II) were involved in the change of the
spikelet architecture during the domestication process.
QTls mapping from the analysis of two (cultivated 
Traits of the Spikelet Structure Displayed wild) F2 populations (Poncet et al. 2000, 2002) has
a Monogenic or Digenic Inheritance confirmed the implication of major genes controlling
the differences in spikelet structure between domestic
The most critical adaptive differences between wild and wild phenotypes (Table 13.5). It has also shown
and cultivated pearl millet involve the spikelet struc- that the segments I and II previously identified corre-
ture (Table 13.3). In wild pearl millet, the presence of spond to LG 6 and 7 on the pearl millet genetic map.
an abscission layer at the basis of the involucre allows Moreover, seed coating that indicates the extent to
fertile spikelets to be shed spontaneously at maturity. which the glumes enclose the seed depends in part
Abundant bristles and the presence of a long awn, all on the length of the glumes (GL). Three unlinked
ornamented with backward-pointing bristles, enhance genetic factors were identified for this trait. A major
the dispersal abilities. Once on the ground, these bris- QTL, gl7, which explained 49.9% of the phenotypic
tles and the arrow-shaped morphology of the sessile variance, mapped to the same region as ct7 on group 7,
spikelets allow deeper penetration in the soil and thus and two other QTLs with minor effects, gl5 and gl6,
contribute to the survival of the seed until its germina- were located on LG 5 and LG 6, respectively
tion. Table 13.4 presents a comprehensive review of (Table 13.5). The wild parental alleles of QTLs, gl6
Mendelian segregation of genes involved in the inher- and gl7, increased glume length with overdominant
itance of spikelet traits. and dominant effects, respectively.
The inheritance of the length of the involucral pedi-
cel (PL) exhibit differences according to the cultivated
Some Insights in the QTL Dissection of Other
and wild parent used in the crosses. Indeed, crosses
Domestication Syndrome Traits
involving the wild ecotype, P. mollissimum, and other
cultivated forms, from different geographical origins
Spike Morphology
had always revealed a monogenic inheritance, whereas
a cross between the line 23d2B and P. violaceum Distributions for the length of the spike (LoS)
showed a digenic inheritance. The presence of an (Fig. 13.11) and the width of the spike (WiS) revealed
abscission layer (AL) and the seed coating have a transgressive segregants among the F2 progenies.
digenic inheritance in most cases. Using the order and The weight of the spike (WeS) was controlled by at
distances between genes, Joly-Ichenhauser (1984) has least three QTLs: wes2 (LG 2), wes5 (LG 5), and wes7

Table 13.4 Number of genes involved in the variation of the seed coating (Ct), the abscission layer (AL), the length of the pedicel
(PL), and their localization on the two main segments of the Pearl millet’s genetic map (segments I or II indicated within brackets) in
cultivated  wild pearl millet crosses involving plant material from different regions in Africa
Traits
Cultivated/wild combinationsa (countries) Ct AL PL
cv. Souna  P. mollissimumb (Mali  Mali) 2 (I? + II) 1 (II) 1 + minor (II)
i.l. Massue/P. mollissimumc (Mauritania  Mali) 1 (I) 1 (I) 1 (I)
cv. Tiotandé/P. mollissimumc (Senegal  Mali) 2 (I + II) 2 (I + II) 1 (I)
cv. Drôo/P. mollissimumc (Tunisia  Mali) 2 (I + II) 2 (I + II) 1 (I)
i.l. 23 d2B/P. mollissimumc (Ghana  Mali) 2 (I + II) 2 (I + II) 1 (II)
cv. Zongo/P. violaceumc (Niger  Niger) 2 (I + II) 2 (I + II) 1 (I)
cv. Drôo/P. violaceumc (Tunisia  Niger) 1 (II) 2 (I + II) 1 (II)
i.l. 23d2B/P. violaceumc (Ghana  Niger) 2 (I + II) 1 (II) 2 (I + II)
cv. cultivar, i.l. inbred line
a
The same accessions of P. mollissimum and P. violaceum were used in the different crosses
b
Poncet et al. (1998)
c
Joly-Ichenhauser (1984), backcrosses and F2 populations are analysed in each combination
13 Pennisetum 241

Table 13.5 Biometrical parameters of individual QTLs and cumulated QTLs (tot) affecting the domestication syndrome traits
Variable QTL Group Locus LOD % var a d d/a d allele
NL /
NT nt7 7 655 2.78 16.4 0.14 0.7 5.09 +
LS /
DS ds7 7 655 2.3 14.7 0.17 1.2 6.99 +
LL ll5 5 345 2.8 14.7 2.61 0.18 0.07 /
WiL /
log(Hmax) hmax7 7 655 3.13 18.9 0.04 0.054 0.02 c
hmax6 6 Est-E-al6 0.04 0.01 0.13 +
tot 5.08 27.2
HPT htp5 5 345-731 5.58 29.9 21.41 27.46 1.28 c
htp2 2 176-592 10.44 16.5 1.58 +
tot 7.66 38.6
HPT-HHead (htp-hhead)1 1 515-756 2.68 15.7 4.61 11.57 2.5 c
NNPT nnpt2 2 856-176 3.5 17.6 0.29 0.86 2.97 c
nnpt6 6 Est-E-al6 2.7 13.9 0.29 0.82 2.82 c
tot 6.0 28.5
sr(NS) ns1 1 461-858 2.38 20.4 0.24 1.37 5.73 +
ns2 2 856-176 2.78 14.3 0.82 0.06 0.08
ns6 6 Est-E-al6 3.37 25.0 1.12 0.1 0.09
ns7 7 655 2.8 15.4 0.63 0.74 1.17 +
tot 11.4 61.3
NTM nbt2 2 332 2.68 13.7 3.31 1.24 0.37 /
nbt5-1 5 (1) 345 4.31 21.4 4.54 0.85 0.19 /
nbt5-2 5 (2) 735 2.79 17.2 3.78 2.26 0.60 c
tot 11.7 57.6
LLM /
WiLM wilm2-1 2 (1) 332 5.72 27.6 0.42 0.041 0.1 /
wilm2-2 2 (2) 592 4.8 24.3 0.41 0.15 0.37 +
wilm4 4 464-515B 3.36 30.9 0.36 0.35 0.99 +
wilm6 6 al6-713 2.34 20.8 0.27 0.35 1.28 c
tot 10.53 54.9
HHead hhead5 5 651-735 6.65 34.7 24.75 26.13 1.06 c
hhead7 7 812 2.47 14.2 16.16 8.04 0.50 c
tot 9.15 42.9
Head head5 5 651-735 10.75 59.0 10.93 10.23 0.94 c
head7 7 655 3.68 24.2 5.52 7.05 1.28 +
tot 15.16 67
BFF bff2 2 176-592 3.2 3.04 0.95 +
bff5 5 651-735 12.78 64.0 10.24 11.1 1.08 c
bff6 6 202 2.4 14.6 2.28 8.07 3.53 c
bff7 7 655 2.62 0.57 0.22 /
tot 19.21 73.5
BFF-Head (bff-head)1 1 858-515 5.04 26.2 1.88 0.12 0.06 /
(bff-head)7 7 655-812 9.4 43.4 1.96 1.38 0.7 +
tot 13.67 58.1
BMF-BFF /
FFF-BFF /
PI /
log(WeS) wes2 2 176-592 15.43 59.7 0.28 0.00 0.00
wes5 5 651-735 2.63 15.8 0.14 0.1 0.72 +
wes6 6 Est-E-al6 2.41 18.8 0.13 0.12 0.9 c
tot 24.5 82
log(LoS) los1 1 858-515 2.34 12.7 0.04 0.00 0.07
(continued)
242 T. Robert et al.

Table 13.5 (continued)

Variable QTL Group Locus LOD % var a d d/a d allele
los2 2 592-738 7.11 34.7 0.07 0.02 0.26
los7 7 655 2.41 13.7 0.03 0.04 1.61 +
tot 12.3 54
WiS wis5 5 735 2.62 15.4 0.34 0.10 0.31 /
wis7 7 655 15.58 62.0 0.54 0.21 0.39 /
tot 22.2 75.8
sr(PL) pl6 6 Est-E- al6 11.86 59.5 0.62 0.15 0.24 /
pl7 7 655 0.28 0.13 0.48 +
tot 17.22 73.4
log(BL) bl6 6 Est-E- al6 3.75 54.7 0.09 0.3 3.01 c
bl7 7 655 8.90 44.6 0.15 0.12 0.77 +
tot 10.02 71.4
GL gl5 5 651-735 3.24 17.9 0.44 0.02 0.05 /
gl6 6 Est-E-al6 2.87 16.4 0.23 0.39 1.70 +
gl7 7 655 10.72 49.9 0.52 0.37 0.71 +
tot 18.48 69.5
% var percentage of phenotypic variance explained, a additive effect of the wild allele (when the positive effect is attributed to the
wild allele, figures are in italic), d dominance effect, d allele the dominant allele is c cultivated or þ wild allele

(LG 7), which accounted for 82% of the phenotypic for sheath (LS) and leaf (LL) length, and to a less
variation (Table 13.5). The major QTL, wes2, alone extent for the number of leaves (NL, TNL), the
explained more than 50%. For all the QTLs, the width of the leaf (WiL), and the number of basal tillers
cultivated alleles increased weight, as predicted by (NT).
the parental means. The length of the spike was mainly A few QTLs were detected (Table 13.5) for the
determined by three QTLs. The major QTL, los2 juvenile traits, nt7, dg7, and lof5. The QTLs nt7 and
(34.7% of the phenotypic variance) was located on dg7 were mapped in the same interval on LG 7. In both
LG 2. The additive effect of the cultivated allele cases, the wild allele exhibited a positive additive
increased the length. In contrast, the positive effect effect and overdominance in their favor.
of the two minor QTLs located on LG 1 (los1) and The maximum height of the plant (Hmax) was
LG 7 (los7) was attributed to the wild alleles influenced by a QTL on LG 7, which was responsible
(Table 13.5). Thus, when positive alleles of the three for 18.9% of the phenotypic variation. The additive
QTLs are accumulated in the same genotype, trans- effect of the cultivated allele of hmax7 increased the
gressive phenotypes with longer spikes than the height.
cultivated parent could be obtained. This can occur Four unlinked regions contributing to variation in
quite easily as these QTLs are located on different the total number of fertile tillers (NS) were identi-
chromosomes. One of the QTLs for both weight and fied: ns1, ns2, ns6, and ns7. All the QTLs were of
LoS, wes2 and los2, coincided on LG 2. For these two similar effect, each explained between 14.3% and
QTLs, the magnitude and direction of allele effects 25% of the phenotypic variation, totaling 61.3%. As
were similar. Two QTLs were detected for the WiS. expected, all wild alleles increased the number of
The effect of wis7 (LG 7) was the largest and explained fertile tillers.
62% of the phenotypic variance. The additive effect of The QTLs detected for the number of basal tillers
the cultivated allele of wis7 reduced the width. A minor at maturity (NTM) were located on LG 2 and LG 5.
QTL on LG 5 was also detected where the wild allele They were distinct from the QTLs for the total
exerted a positive effect (Table 13.5). number of spikes (NS). However, ns2 and ns6 were
closely linked to two QTLs controlling the number
of nodes on the primary tiller (NNPT). The QTLs
Plant Architecture
ns7 and nt7 may correspond to the same QTL,
Differences between the wild and cultivated pheno- whereas ns1, ns2, and ns6 could be involved in
types were observed at the juvenile stage, especially axillary tillering.
13 Pennisetum 243

Fig. 13.11 Frequency 15

distribution for spike length in
14 F2 _ F1 Cultivated
the F2 population. The means _ _
13 X = 12.9 (±2.6) X = 14.4 (±1.9) X = 14.6 (±1.8)
and standard deviations (in
brackets) are given for 12
parents, F1 and F2 populations 11

Number of observations
10
9
8
7 Wild
6 X = 7.1 (±1.5)
5
4
3
2
1
0
7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23
Length of the spike (LoS) in cm

Flowering of the major quantitative traits of the domestication

syndrome (Fig. 13.12). This finding is consistent with
The two (cultivated  wild) F2 progenies analyzed
the current data on the domestication of the principal
shows a variation in flowering time. The dates of
cereal species. Indeed, comparative mapping studies
heading and female and male flowering were all asso-
suggest that domestication was achieved by selection
ciated with a major QTL on LG 5 that was responsible
(along with the other evolutionary forces) on only
for a minimum of 50% of the phenotypic variation
few genomic regions (Peng et al. 2003; Li et al.
observed for each trait (Table 13.5). The cultivated
2006; Purugganan and Fuller 2009). Nevertheless,
alleles at this QTL delayed heading and flowering in a
this general statement could be relative by consider-
dominant manner. LG 6 and 7 also carried QTLs with
ing the methodological aspects of QTL mapping. The
weaker effects on these characters. Variation in the
usual sizes of the mapping population might be too
delay between heading (Head) and beginning of
small for the detection of scattered loci with low
stigma emergence (BFF) was observed. Two QTLs
effect on domestication quantitative traits. The very
on LG 7 and LG 1 jointly accounted for 58% of the
importance of these types of QTL as well as the
phenotypic variation for this trait.
pleiotropic and/or epistatic effects of linked genes
on QTL expression are still to be investigated in
Pennisetum by means of QTL fine-mapping.
Concluding Remarks on the Wealth of Wild Relatives
for Unraveling the Evolutionary Mechanisms
Underlying the Domestication Process
Transgressive Phenotypes Resulting from Wild
QTL Effects: An Evolutionary Fancy and Basis for
The genetic control of the domestication syndrome
New Genetic Resources Enhancement Strategies
in Pennisetum sp. at the spikelet level involved fac-
tors with large effects that were located on LG 6
Transgressions and Genetic Resource
(Fig. 13.12) and/or on LG 7. The evolution from
wild to cultivated spikelet morphology occurred For most of the major QTLs, the directions of the
mostly through recessive mutations. effects of the parental alleles were as predicted by
QTL analysis revealed that a few loci mapped on the parental phenotypic means. However, for some
few genomic regions are involved in the inheritance traits, Hmax, LoS, WiS, BL, and GL, positive alleles
244 T. Robert et al.

GL1 GL2 GL4

GL6
Xpsm461 Xpsm464
Xpsm322
GL3
Xpsm202 R2 > 20%
Xpsm856 Xpsm592b
R2 < 20%

Xpsm176
Xpsm476
Generative structures Xpsm588
Xpsm592 Xpsm515b
Panicle length Xpsm568
Panicle weight Xpsm84
Xpsm858
Glume length Xpsm416
Xpsm696
Xpsm515
Panicle width Xpsm738
Xpsm756 EST-E
Bristle length
Peduncle length Xpsm336
GL5 Xpsm416b
Xpsm386

Vegetative structures Xpsm345

GL7
4th leaf width
Number of tillers Xpsm655
Number of fertile tillers Xpsm749
Xpsm731
Main culm height
Plant height Xpsm651
Nb of nodes on main culm Xpsm812
Xpsm713

Xpsm735

Fig. 13.12 Consensus map of domestication QTLs identified in two cultivated  wild F2 progenies. Most of the traits analyzed
involved major QTLs. The linkage group 6 and 7 harbors a high number of QTLs, especially those involved in the variation of
spikelet structure

were contributed by both the parents. When an indi- genes not only to enhance stress tolerance but also to
vidual received positive alleles from both the parents substantially increase yield. Such favorable genes have
for a particular trait, its phenotypic value exceeds the already been detected for fruit in tomato (Fulton et al.
parental ones. Transgressive phenotypes occurred 1997) and for the number of grains per panicle in rice
even easier when a trait is controlled by unlinked (Xiao et al. 1996). Introgression of Oryza rufipogon
QTLs (e.g., LoS). wild alleles at two specific loci into a top-performing
The existence of wild alleles that reinforce the rice variety was associated with yield increases of 18%
cultivated phenotype in the F2 background (WiS, and 17%. Based on this finding, new strategies were
LoS, Hmax, etc.) raises questions on the mechanism designed for massive introgression of QTL alleles
allowing their maintenance in the wild populations from wild relatives (Tanksley and McCouch 1997;
while they are expected to decrease the adaptive McCouch et al. 2006). Improvement of yield compo-
value of the wild plant in natural conditions. This nents in pearl millet, such as LoS, could thus be
paradox could be explained by epistasis of the wild obtained by introgression of wild alleles. But this strat-
genetic background on the effect of the QTL leading to egy should be carefully designed because of the strong
its silencing. This hypothesis is still to be tested. gene flow supported by the allogamous breeding sys-
According to the evolutionary history of a species tem and the sympatric or parapatric distribution of
gene pool, the bottleneck affecting the genetic diver- pearl millet with its wild relatives (P. mollissimum or
sity and the differentiation are more or less important. P. violaceum). In addition, this transgression phenom-
However, such results open promising possibilities enon could be the source of vigorous introgressed
to enhance pearl millet landraces by using genetic intermediate forms, which are often considered as
resources from the wild. Indeed, despite their overall aggressive weeds by farmers (see Sect. 13.1.3.2).
lower value in terms of agronomic performance, the This point was raised by Rieseberg et al. (2007) in
closest wild relatives of crops can be sources of novel sunflower.
13 Pennisetum 245

13.2 Enhancement of Genetic Resources van Dijk 2007). Molecular markers and genomic tools
Using Genes from Wild Pennisetum have greatly enhanced the understanding of his trait.
Apomictic meiosis has for long been regarded as by-
Relatives
passed or non-functional. Recently, using genomic
approaches, it has been demonstrated in Arabidopsis
As shown in previous sections, the Pennisetum com- mutants that the gene sw1 (DYAD) causes synapsis to
plex of species provides a favorable biological context fail in female meiosis and yields two unreduced
for addressing fundamental questions on the evolution gametes (Noyes 2008).
of grasses. As for other cereal species belonging to the The genus Pennisetum contains many apomictic
Poaceae family, wild relatives have been targeted as species (see Tables 13.1 and 13.2), for example:
either sources of genes of interest for tolerance to P. setaceum and P. squamulatum. C. ciliaris (previ-
biotic and abiotic stresses or raw material for direct ously P. ciliare), a close relative to Pennisetum species
use. Many wild Pennisetum species were used in grass (see Fig. 13.4), also deserves a mention.
lands of tropical areas or as fodder crop. For example, A large (>50 Mb) ASGR was observed for
P. purpureum, P. squamulatum, P. orientale, P. poly- P. squamulatum (Goel et al. 2003; Akiyama et al.
stachion, P. flaccidum, and P. pedicellatum contribute 2004, 2005). Gametophytic apomixis in C. ciliaris
significantly to non-cultivated forage production in the is also controlled by ASGR (Akiyama et al. 2005).
tropics. Some species (P. alopecuroides, P. orientale, This trait is highly conserved and macrosyntenic
P. setaceum) are grown as ornamental plants. with ASGR of P. squamulatum. Recently, Conner
A sustainable use of wild genetic resources depends et al. (2008) have sequenced and analyzed a portion
by far on the accuracy and the thoughtfulness of fun- of ASGR in Pennisetum and Cenchrus. ASGR
damental knowledge about the structure of genetic sequences reveal the presence of transposable ele-
diversity and genome organization as revealed by ments and contain multiple regions of microsynteny
evolutionary genomics studies on the gene pools. with Oryza sativa. Forty potentially transcribed
In this section, we put emphasis on some meaning- genes were identified, some of them may play a
ful strategies developed for optimizing the use of wild role in apomictic development.
Pennisetum genetic resources. Given the large non-recombining property of
ASGR and the presence of repetitive DNA, the identi-
fication of apomixis gene by genetic mapping seems
13.2.1 Wild Pennisetum Species as particularly difficult.
Apomictic trait has already been transferred from
Sources of Major Genes of
wild progenitors to cultivated forms for the purpose of
Agronomic Interest (GAI) propagating heterozygous genotypes and hopefully
heterosis.
13.2.1.1 Apomixis In breeding for fodder, the aposporous apomictic
trait from P. squamulatum is being transferred to pearl
Apomixis is an asexual reproduction system that pro- millet by means of backcrosses. Using a molecular
duces progenies via seeds without sperm–egg fusion. cytogenetic method genomic in situ hybridization
Embryos arise from cells of the maternal genotype. (GISH), it was demonstrated that in BC8, one to
This trait occurs in wild relatives of crop plants and three chromosomes from P. squamulatum are present
can be introduced by crossing. The inheritance of this in the apomictic plants. This finding evidences for
breeding system has been studied in many plant many hopes for the release of a apomictic pearl millet
families both in monocots and dicots (for example: variety for fodder purpose. Nevertheless, many diffi-
Poaceae, Liliaceae, Brassicaceae, Compositeae, etc.). culties are still to be resolved before producing desired
Many reviews and books have recently been devoted apomictic pearl millet varieties for fodder purpose
to understanding apomixis (Savidan 2001; Ozias- (Ozias-Akins et al. 2003; Akiyama et al. 2004; Jauhar
Akins 2006; Hoerandl et al. 2007; Ozias-Akins and et al. 2006).
246 T. Robert et al.

Male Sterility When Natural Reproductive Barriers crosses. Wild P. glaucum ssp. violaceum from primary
Turn to be Source of GAI gene pool and species from tertiary gene pool were
evaluated for disease resistance, mainly for Pyricu-
The gene pool classification is based on natural cross- laria grisea, Puccinia substriata var. indica (Wilson
ability between the wild relatives and the domesticated and Hanna 1992). Many resources of germplasms
form. The hybrid breeding strategy widely uses cyto- present at least resistance to one of the five pathogens
plasmic male sterility either naturally occurring in the tested and offer global valuable diversity for disease
cultivated form or induced using interspecific hybridiza- resistance.
tion, for example. In pearl millet breeding for F1 vari- Resistance to P. grisea (Cke) Sarc. leaf spot of
eties, A1 cytoplasmic male sterility (CMS) has been pearl millet was discovered in P. glaucum subsp.
widely used. Nevertheless, in order to avoid cytoplasmic monodii originating from Senegal, and was transferred
uniformity and the risks of vulnerability to disease and effectively to pearl millet (Hanna and Wells 1989). So,
insect pests, other CMS sources were identified as alter- both a resistant inbred line and a resistant commercial
native to A1 CMS. Two new CMS systems have been forage hybrid were produced.
identified in wild species P. glaucum ssp. violaceum Another important pathogen is a parasitic weed
(¼ monodii) from Senegal: Av CMS (Marchais and Striga hermonthica that infects grain fields resulting
Pernès 1985) and A4 CMS (Hanna 1989). Evaluation in an annual grain yield losses. Resistance to S. her-
of three CMS systems (Av, A4, and A5; Rai 1995) monthica was evaluated in more than 250 wild
indicates that the A4 CMS system provides a better P. glaucum subsp. monodii and stenostachyum acces-
alternative to the A1 CMS system than the Av system sions (Wilson et al. 2000, 2004) in diverse regions in
(Rai et al. 2001). Chandra-Shekara et al. (2007) confirms West Africa. Four accessions were classified as resis-
the potentiality of A4 and A5 CMS sources as potential tant and are useful sources of striga resistance for pearl
alternatives to diversify the male sterile cytoplasm in millet breeding in West Africa.
pearl millet hybrid breeding. Rai et al. (2009), comparing Tolerance to abiotic stresses like drought or salinity
the three CMS systems (A1, A4 and A5) suggested that have been tested in wild relatives. For example, acces-
seed parents breeding efficiency will be better with the sions of P. purpureum were evaluated for tolerance to
A5 CMS system, followed by the A4 CMS system, and salinity but no tolerance was detected in this material
lesser with the currently commercial A1 CMS system. (Kulkarni et al. 2006).

Genes Involved in Tolerance to Biotic and Abiotic 13.2.2 Wide Use of Wild Pennisetum
Stress
Species
Despite the drawbacks of hybrids sterility, species
from secondary and tertiary gene pools are also poten- 13.2.2.1 Breeding for Fodder
tial donors of favorable traits (Hanna 1986, 1990; Rao
et al. 2003). Evaluation of wild accessions for useful Genetic improvement of Napier grass itself for
traits, such as resistance and tolerance, is needed for the increase of biomass yield is usually achieved
incorporation of these species in breeding strategies. using ecotypic selection. An extensive breeding and
For instance, P. orientale L. C. Rich (2n ¼ 2x ¼ 18) yield evaluations were performed in Florida during
has been assessed as potential donor for some charac- the 1980s. Two high yielding improved varieties
ters like perenniality, drought tolerance, pest resis- “Merkeron” Napier grass (a vegetatively propagated
tance, and winter hardiness (Dujardin and Hanna F1 clone) and “Mott” Napier grass (a selfed progeny
1987). Trispecific hybrids between pearl millet, of Merkeron Napier grass) have been registered in
P. squamulatum, and napier grass have also been 1989. In tropical grasses, numerous interspecific and
obtained and could be used to transfer germplasm some intergeneric hybrids were produced (see Burson
across species (Dujardin and Hanna 1985). and Young 2001). For Napier grass, in addition to
The traits of agronomic interest identified in wild conventional breeding programs, the interspecific
species are usually transferred to pearl millet via back- hybridization between P. purpureum and pearl millet
13 Pennisetum 247

is an alternative to obtain superior fodder cultivars. effective than tree hedges for reduction of soil erosion
Indeed, P. purpureum, one of the two members of and thereby for yield improvement of maize. It has
the secondary gene pool, is easily crossable with been widely adopted among indigenous grasses as the
P. glaucum, but sterility is predominant in the interspe- more popular in Kenya for soil conservation.
cific triploid (2n ¼ 3x ¼ 21) hybrids. Nevertheless,
these triploid hybrids show high heterosis for fodder
quality and yield, due to introgression of favorable Push–Pull
traits from P. glaucum, such as forage quality. Allohex-
aploids expressing male- and female-fertility have been In Subsaharan African regions, stem borers and para-
produced by means of colchicine chromosome dou- sitic weeds like striga are among the main constraints
bling (Gonzalez and Hanna 1984; Hanna et al. 1984). affecting cereal production. Therefore, research pro-
In addition, Hanna (1990) showed the potentiality for grams on food security concerns have been launched
gene transferring from P. purpureum to P. glaucum for the improvement of farmers’ livelihood. The man-
through a mechanism producing monoploid gametes agement of these two pests is central in these pro-
(see Sotomayor-Rios and Pitman 2001 and Jauhar grams. For instance, ICIPE (International Center of
et al. 2006, for review). Insect Physiology and Ecology) have developed an
Hybrids between pearl millet and wild P. glaucum integrated management system named: Push–Pull in
subsp. monodii accessions from three African Sahelian mitigation. This management technology consists in
countries were tested during 2 years for production of intercropping a fodder legume like Desmodium unci-
dry matter (Hanna 2000). This study and others later natum with maize and planting a perimeter of
confirm that genes for enhancing pearl millet yield are P. purpureum (Niaper grass) around the plot. Stem
present in the wild grassy subspecies. borers are repelled away from the maize crop (Push)
and trapped by the Niaper grass plants (Pull). In addi-
tion, Desmodium roots produce several polyphenolic
13.2.2.2 Miscellaneous Uses of Wild Relatives compounds, some of which stimulated striga seeds’
germination while some others inhibit haustorial for-
BioEnergy mation thereby suppressing striga. The adoption of
this technology by farmers has been tested in Kenya
With progress made on converting cellulosic feed- (Khan et al. 2008). Hari and Jindal (2009) tested the
stocks to ethanol, Napier grass, due to its high biomass efficiency of P. purpureum  P. glaucum as trap crop
producing, could be used for ethanol production for the management of the stem borer Chilo partellus
(Anderson et al. 2008a). Three species (Bermuda on maize. According to their data, Niaper millet
grass, Napier grass, and giant reed) were compared hybrids were preferred for oviposition over maize by
for ethanol production, the results indicate that Ber- the stem borer moths. But subsequently, this material
mudagrass yielded more ethanol compared to napier were found to be poor larval hosts and less eaten by
grass (see Anderson et al. 2008b for more details). borer larvae.

Hedge Vegetative Barriers for Monitoring Soil 13.2.2.3 Pre-breeding Strategies

Erosion in Fields
Bramel-Cox et al. (1986) showed that progeny derived
P. purpureum is also currently used as vegetative from pearl millet  wild species had higher growth
hedge barrier. For instance, it was tested as buffer rates. As mentioned in the Sect. 13.1.3.2, the occur-
strips on row crop production fields (Anderson et al. rence of transgressive phenotypes in F2 P. glaucum 
2008c). The Efficiency of narrow strips of Napier grass P. violaceum population (Poncet et al. 2000) suggests
(P. purpureum) and Vetiver (Vetiveria zizanioides) in that the wild forms can be used to improve pearl millet
reducing runoff and nutrient loss from the field have and be integrated into breeding programs. Hajjar and
been tested in Kenya (Owino et al 2006). In this study Hodgkin (2007) reviewed information on the presence
and others (Mutegi et al. 2008), Napier grass was more of useful genes from crop wild relative (CWR) in
248 T. Robert et al.

cultivars of crops and noted an increase in the rate of ered from genes to eco/agrosystems. As noted by many
cultivars containing genes from CWR, compared to authors (see Purugganan and Fuller 2009 for a review),
the figures previously reported by Prescott-Allen and domestication is one of the most important technologi-
Prescott-Allen (1986, 1988). cal innovations in human history. Humans and Poa-
Potentiality of wild Pennisetum species for genetic ceae family are interdependent. Poaceae family
improvement of pearl millet is well documented and includes about 10,000 species, which provides cereals,
reviewed in Rao et al. (2003) and Jauhar et al. (2006). forages, sugarcane. The basis of this interdependence is
All the above mentioned strategies for the use of to be unraveled both through anthropological studies
genes from wild relatives are often restricted to the and biological studies aiming at understanding the spe-
transfer through backcrossing of genes involved in cial features that put Poaceae species on the trail of
simply inherited characters (e.g., genes encoding for domestication. Thus, domestication process has inspired
resistance to disease or tolerance to stresses). How- many scholars including Charles Darwin. The appropri-
ever, this procedure may lead to genetic erosion ate paradigm for the study of domestication process
because of the large amount of wild relatives collected is necessarily based on a multidisciplinary approach
and the genetic uniformity resulting from backcrosses (genetics, cytogenetics, genomics, physiology, arche-
on the recurrent cultivated parents. Pre-breeding has ology, botany, anthropology, economic botany, agron-
been identified as an appropriate solution to avoid this omy, etc.). The key question is actually to understand
drawback and for the conservation and massive use of the genetic basis of adaptation. Wild relatives of crop
genes from wild relatives. A project aiming at the plants are central to the assessment of this question.
implementation of pre-breeding in pearl millet has Thanks to the dramatic progress in understanding
been developed (Lamy et al. 1994; Sarr et al. 2001). genome organization made possible by molecular
In the frame of an integrated paradigm developed for genetics and genomics, the mechanisms underlying
the study of domestication process in pearl millet, plant adaptation and plasticity can now be dissected.
recombinant populations (P. glaucum  P. violaceum) The ultimate goal is to understand from an evolution-
were created and tested in two African countries. The ary point of view the architecture of key adaptative
levels of introgression were assessed using molecular characters. Comparative QTL mapping and subsequent
markers (allozymes, RLFP, STS, AFLP) and cytoge- molecular investigations leading to QTA and QTN
netics combined with reproductive biology and quan- (individual nucleotide that causes particular pheno-
titative trait segregation. This strategy was efficient in type) suggest parallel molecular evolution in Poaceae
partitioning the recombinant offsprings according to family at some few key loci involved in the domestica-
their levels of introgression with wild genome. Three tion process (e.g., ZmTB1 gene). These studies bear
pools were identified according to the number of intro- future hopes for understanding how some grass mem-
gressed linkage groups. These populations are poten- bers of the Poaceae family were recruited successfully
tially interesting as both conservation units and in the domestication process. This issue could signifi-
breeding material. cantly help to uncloak the genetic architecture of the
useful traits and then to monitor efficiently wide trans-
fer of genes from wild relatives. As discussed in this
chapter, the genus Pennisetum offers favorable condi-
13.3 Conclusion and Future Scope
tions to address this issue. Indeed, the occurrence of the
of Research domesticated P. glaucum in sympatric or parapatric
situations with wild relatives in the Sahelian region
In this chapter, we put emphasis on the domestication is a unique situation for the implementation of the
process as it occurred in the primary gene pool of above mentioned paradigm. Climatic pejoration hardly
P. glaucum. Indeed, it is currently understood that the impacts this region generating, in conjunction with
challenges and hopes for sustainable uses of genetic human activities, accelerated desertification process.
resources are by far dependent on the understanding of Thus, unraveling the genetic basis of adaptation of
the dynamics of biodiversity, i.e., the factors involved pearl millet and its wild relatives to these difficult
in the origin, maintenance, or loss of diversity consid- ecological conditions is a major issue. Indeed, gene
13 Pennisetum 249

flow, farmers’ strategies to monitor the evolution of Amoukou AI, Marchais L (1993) Evidence of a partial repro-
domesticated varieties, and folk uses of wild relatives ductive barrier between wild and cultivated pearl millets
(Pennisetum glaucum). Euphytica 67:19–26
can be addressed on a multidisciplinary basis (anthro- Amouroux-Pezas C (1985) B Chromosomes occurrence in a
pological, genetics, and genomic approaches, for wild Pennisetum form (P. violaceum) : Behaviour and trans-
instance). These studies are under progress and need ferring strategies into domesticated pearl millet inbred lines.
strong support in the frame of ecological rehabilitation PhD Thesis. Paris XI University, France. (Orsay) ( Les
chromosomes B du mil : leur gestion dans une forme spon-
and biodiversity management programs. Conserva- tanée Pennisetum violaceum et leur transfert dans des lig-
tions initiatives for Pennisetum species have been nées cultivées. Thèse docteur 3ème cycle Orsay, France
launched by many international organizations and Anderson WF, Dien BS, Brandon SK, Peterson JD (2008a)
national bodies. Many gene banks hold accessions of Assessment of bermudagrass and bunch grasses as feedstock
for conversion to ethanol. Appl Biochem Biotechnol
Pennisetum species. But the in situ conservation initia- 145:13–21
tives involving farmers’ knowledge and know-how Anderson WF, Casler MD, Baldwin BS (2008b) Improvement
should be prioritized. The on-farm conservation con- of perennial forage species as feedstock for bioenergy. In:
cept is consistent with these initiatives. Vermerris W (ed) Genetic improvement of bioenergy crops.
Springer, Berlin
As for industrial or commercial uses of Pennise- Anderson WF, Hubbard RK, Strickland TC (2008c) Napier
tum species, many initiatives are under progress. grass as a field border and bioenergy source. In: Proceedings
Nevertheless, the improvement in these fields could of joint annual meeting of the Georgia Chapters SWCS,
benefit from the above mentioned assessment of ASABE, and IECA, Athens, GA, USA
Antonovics J, Bradshaw AD (1970) Evolution in closely adja-
genetic basis of adaptation and dissection of the cent plant populations. VIII. Clinal patterns at a mine bound-
architecture of characters using evolutionary geno- ary. Heredity 25:349–362
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Avdulov NP (1931) Karyo-systematische untersuchungen der
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.
Chapter 14
Phleum

Alan V. Stewart, Andrzej J. Joachimiak, and Nicholas W. Ellison

14.1 Introduction northern latitudes of Europe and North America with

approximately 34,000 ton produced each year
(Anonymous 2007). Use of Timothy in the southern
The genus Phleum contains one major perennial for-
hemisphere is minimal with the major user, New Zeal-
age species, Timothy (P. pratense) and approximately
and, using less than 100 ton annually (Charlton and
13 other species. Timothy is commonly used as a
Stewart 2000; AgriQuality 2007).
forage and hay crop in cold temperate regions of the
Outside its original Eurasian range, P. pratense has
world, where it is noted for its winter hardiness and
now expanded to most suitable habitats around the
high forage quality.
world. It is now widespread in North America, where
Phleum species have only rarely been crossed with
it is both a major and valuable forage species and,
species outside this genus and they are not closely
along with many introduced grasses, is of ecological
related to any major crop. Therefore, the story of
concern for displacing native grasses.
wild relatives for Phleum is largely one of species
and genomic diversity within the genus. Although
the distribution and diversity of Phleum is complex,
much of this can be readily explained by its evolution-
ary, migrational and genomic history over the last few 14.3 Morphology and Flowering
hundred thousand years. The understanding of the Behavior
diversity within wild relatives of Timothy has consid-
erable implications for germplasm collection and con-
Timothy is a perennial bunchgrass that grows up to 1.5-
servation as well as for breeding techniques employed.
m tall with flat leaves and is characterized by corms,
bulb-like swellings of stem bases that serve as storage
organs for carbohydrate reserves. New tillers form
14.2 Agricultural Status from axillary buds at the base of the corms, resulting
in a relatively non-aggressive spreading habit. At times
elongated stems trail along the ground and root at the
Phleum is the third most important forage grass genus
nodes, thus exhibiting a semi-stoloniferous habit. This
of the temperate world following Lolium and Festuca.
is particularly apparent in diploid forms from central
Seed production of Timothy makes up over 8% of world
Europe (Kovats 1976), but it is also common in
production of temperate forage species (Bondesen
P. pratense forms as a result of high temperature pre-
2007). Its production and use is concentrated in the
venting reproductive initiation but not internode elon-
gation (Cooper 1958).
Commercial hexaploid Timothy lacks a vernaliza-
A.V. Stewart (*)
tion requirement for flowering unlike many other tem-
PGG Wrightson Seeds, PO Box 175, Lincoln, Christchurch
7640, New Zealand perate grasses. It, therefore, has no strong mechanism
e-mail: [email protected] to prevent reproductive development during summer

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 257
DOI 10.1007/978-3-642-14255-0_14, # Springer-Verlag Berlin Heidelberg 2011
258 A.V. Stewart et al.

and the growing point becomes elevated and vulnera- 14.4.1.2 P. pratense L.
ble to removal by grazing. This makes the plant sus-
ceptible to grazing damage in summer. However, a P. pratense L. is a lowland species represented by a
vernalization response is known in diploid germplasm diploid to octoploid polyploid series
(Cooper and Calder 1964) and some Mediterranean
1. Diploids occur throughout much of Europe and
hexaploid germplasm, but breeders have not intro-
parts of North Africa; P. pratense subsp. bertolonii
duced this feature into commercial cultivars.
(DC.) Bornm.
P. bertolonii DC.
P. nodosum L.
2. Less common tetraploid forms in southern Europe;
P. pratense subsp. pratense.
14.4 Taxonomy 3. Widespread agricultural hexaploid forms; P. pra-
tense subsp. pratense.
4. An octoploid form restricted to southern Italy;
Species of the genus Phleum have been divided into
P. pratense subsp. pratense.
four sections as outlined by Joachimiak 2005 (mod-
ified from Humphries 1980; Dogan 1988, 1991; Dogan
and Us 1996). 14.4.1.3 Phleum echinatum Host.

Phleum echinatum Host. is the most unusual spe-

cies within the section Phleum as it is a winter active
14.4.1 Section Phleum annual grass of eastern Mediterranean mountains.

Today eight “entities” from diploid to octoploid are

recognized in three species, two perennials and one
14.4.2 Section Chilochloa (Beauv.) Dum.
annual (Joachimiak 2005; Kula 2005a; Stewart et al.
2008). In this review, we use the widely accepted
nomenclature of Humphries (1978, 1980) for This contains four perennial and three annual spe-
European species and of Barkworth et al. (2007) for cies representing the polyploid series from diploid to
American species as follows: octoploid.

14.4.2.1 P. phleoides (L.) Karsten,

14.4.1.1 P. alpinum L.
P. phleoides (L.) Karsten, is a perennial grass occurring
P. alpinum L. is an alpine species differentiated into in dry grasslands, often in calcareous soils in parts of
diploid or tetraploid cytotypes with ciliate or glabrous Asia, Europe and Morocco (Fig. 14.1) at altitudes of
awns: 150–2,800 m (Dogan 1991). This species and P. hirsu-
1. A glabrous awned tetraploid; P. alpinum L.
tum are seen as a core species of section Chilochloa
P. commutatum Gaudin. with a circumpolar north- (Joachimiak 2005). Diploid and tetraploid forms occur
ern hemisphere and South American distribution. (Humphries 1980). This species exhibited little agri-
2. A ciliate awned diploid form, known from the cultural potential in New Zealand and was noted as
Rhaetic Alps of Italy and the Balkans; P. alpinum having poor productivity, an open growth habit and an
subsp. rhaeticum Humphries
P. rhaeticum early flowering behavior with few seedheads (Caradus
(Humphries) Rauschert. 1978).
3. A glabrous awned diploid, also known as P. alpi-
num L., which cannot be differentiated morpholog-
ically from the widespread tetraploid; for ease of 14.4.2.2 P. hirsutum Honckeny
presentation we will refer to this by the informal
name “commutatum” following Joachimiak and P. hirsutum Honckeny is a diploid perennial of grass-
Kula (1993). lands, open woods, stony places and mountain pastures
14 Phleum 259

Fig. 14.1 Distribution of P. phleoides (after Conert 1998)

at 900–2,200 m in South and Central Europe from at 1,500–2,000 m in Asia from Russia to Turkey, in
Bulgaria to Italy (Humphries 1980). This species exhib- North Africa in Morocco and in Europe from Bulgaria
ited little agricultural potential in New Zealand (tested and Spain.
as P. michelii All.) and was noted as having poor pro- Subsp. serrulatum (Boiss) Dogan occurs on moun-
ductivity with an early flowering behavior with few tain slopes, in tree plantations, mixed deciduous wood-
seedheads (Caradus 1978). lands on sandy soils at 430–2,800 m, in Asia from Iran
to Turkey and in Europe from Bulgaria to Italy.
This species exhibited little agricultural potential
14.4.2.3 P. arenarium L. in New Zealand and was noted as having poor produc-
tivity with an early flowering behavior with few seed-
P. arenarium L. is a diploid annual on sand dunes, dry heads (Caradus 1978).
limestone verges from 0 to 1,700 m, distributed in
southern and western regions of Europe as well as
14.4.2.5 P. paniculatum Hudson
Morocco (Humphries 1980). This species exhibited
little agricultural potential in New Zealand and was
P. paniculatum Hudson is a tetraploid annual
noted as having poor productivity but had an early
(Humphries 1980), which is commonly divided into
flowering behavior with a prolific number of seed-
two subspecies:
heads (Caradus 1978).
Subsp. ciliatum (Boiss.) Dogan occurs on mountain
slopes, sandy places and dried mud from 30 to 1,800 m,
distributed in Europe from Greece to Britain.
14.4.2.4 P. montanum C Koch
Subsp. paniculatum occurs in fields, dry rocky slopes
and open places, often on limestone, 300–2,440 m, and
P. montanum C Koch is a variable perennial grass of
is distributed in Asia from Iran to Turkey.
dry mountain slopes in southern and eastern Europe
and Asia. Its chromosome numbers are recorded as
2x ¼ 14, 4x ¼ 28, and 6x ¼ 42 and it is divided into 14.4.2.6 P. himalaicum Mez.
two subspecies:
Subsp. montanum occurs on dry mountain slopes, P. himalaicum Mez. is an octoploid (8x ¼ 56) ann-
grassy igneous sandstone slopes, edges of beech forests ual on mountain slopes and steep rocky hillsides,
260 A.V. Stewart et al.

1,300–2,200 m, distributed from the Himalayas of 80 to 1,650 m, distributed from Iran to Britain. The
India through to Afghanistan (Joachimiak 2005). chromosome size of this form was 1.6 pg (studied as
P. graecum Kula 2005b).
Subsp. aegaeum (Vierh.) Greuter occurs on sand
14.4.2.7 P. iranicum Bornm. et Gauba dunes at sea level, distributed from Palestine to
Greece.
P. iranicum Bornm. et Gauba is a perennial with
creeping rhizomes occurring on open slopes, stony
clays from 2,130 to 2,800 m, distributed in Iran. This
species exhibited little agricultural potential in New 14.4.4 Section Maillea (Parl.) Horn
Zealand and was noted as having poor productivity, af Rantzien
and an early flowering behavior with few seedheads
(Caradus 1978). This contains a single annual species P. crypsoides
(d’Urv.) Hackel distributed on sandy soils near the
coast in Cyprus, Italy, Greece, and Crete.
14.4.3 Section Achnodon (Nees) Griseb.

This section contains three diploid annual species. 14.4.5 Cytology and Karyotype

14.4.3.1 P. subulatum (Savi) Asch. et Graelon. Phleum has a basic chromosome number of seven.
Species range from diploid (2n ¼ 14) to octoploid
P. subulatum (Savi) Asch. et Graelon. is a diploid (2n ¼ 56), with the exception of P. echinatum, with
species considered to be a core species for this section a reduced chromosome number of 2n ¼ 10. Karyo-
(Joachimiak 2005). It is divided into two subspecies: types of P. pratense and P. alpinum taxa are slightly
Subsp. subulatum occurs on sandy clay cliffs on asymmetric and composed of medium-sized chromo-
coast, limestone mountain slopes, grasslands, oak somes. The chromosome complement of P. echinatum
and pine forest from 0 to 1,100 m, in the region from is much bigger and highly asymmetric. Heterochro-
Turkey to Israel and Cyprus. matin is mostly located near centromeres and telo-
Subsp. ciliatum (Boiss.) Humphries occurs on meres. Joachimiak (2005) provides a comprehensive
grassy slopes, lawns, stony ground near the coast, on review of the karyotypes of the majority of these
limestone 0–700 m, in Greece and Crete species.
The genomes of species in the different sections
appear to be substantially different since molecular
14.4.3.2 P. boissieri Bornm. probes designed to detect species in section Phleum
failed to detect those of P. montanum in section Chilo-
P. boissieri Bornm. is a diploid occurring in oak scrub, chloa (Cai and Bullen 1994).
rocky limestone slopes, sand dunes, fallow fields
450–1,300 m, from Afghanistan across to Britain
(Dogan 1991). The behavior of B-chromosomes in
14.4.6 Genome Size
this species has been studied by Joachimiak (1982).

Phleum has a relatively small genome size and seems

14.4.3.3 P. exaratum Hochst. particularly prone to developing higher ploidies
(Kellogg and Bennetzen 2004). In the majority of species
P. exaratum Hochst. is a diploid of dry open habitats the basal genome size (1Cx) does not exceed 2 pg
divided into two subspecies: (Sliwinska et al. 2003; Kula 2005a). The only excep-
Subsp. exaratum Hochst. Ex Griseb. occurs in open tion is P. echinatum with 1Cx ¼ 3.64 pg. Sliwinska
places, pine forest, saltflats and calcareous soils from et al. (2003) showed that as expected some DNA mass
14 Phleum 261

is lost with increased ploidy so that tetraploids have – Autotetraploid pratense in France BSBSBNBN
slightly less than expected for twice the diploid, – Allotetraploid pratense in the Italian Alps BNBN
an effect due to the partial loss of heterochromatin RGRG
(Joachimiak 2005). – Tetraploid hybrid of bertolonii and hexaploid pra-
Section Chilochloa contains at least two groups tense BNBNBNRG
based on genome size, P. hirsutum (1.9 pg) and – Common agricultural hexaploid pratense BNBNBN
P. phleoides 1.8 pg are large, while both P. arenarium BNRGRG
and P. montanum (1.4 pg) are smaller (Kula 2005b). – Hexaploid pratense in southern Italy RGRGXXXX
The genome size of P. subulatum (1.9 pg) in section – Hexaploid pratense in Morocco BSBSXXXX
Achnodon is similar to that of P. phleoides and – Octoploid pratense in southern Italy R8R8XXXX
P. hirsutum (Kula 2005b). XX

14.4.7.3 P. echinatum
14.4.7 Genomic Formula
P. echinatum has been assigned EE.
Genomic formulae have been assigned to 22 genomic
forms within section Phleum (Fig. 14.2).
14.4.7.4 Other Sections

The only species from any other section for which it is

14.4.7.1 P. alpinum
possible to assign a partial genomic formula is RARAXX
for P. paniculatum of section Chilochloa.
– Ancestral paleo-diploid rhaeticum in Asia RARA
– Diploid rhaeticum in the Alps RSRS
– Diploid rhaeticum in the Pyrenees RPRP
– Diploid rhaeticum in Italy RIRI
– Diploid rhaeticum in Greece RGRG
14.5 Distribution in Relation
– Diploid “commutatum” in the Carpathian moun- to Historical Glaciations
tains CC and Migrations
– Diploid hybrids of rhaeticum and “commutatum”
RC In order to understand the distribution and relation-
– Tetraploid rhaeticum “commutatum” hybrids, Italy ships among species in section Phleum, it is important
RSRSCC to understand how historical glaciation events have
– Tetraploid rhaeticum “commutatum” hybrids, driven migrations that allowed a range of entities to
Caucasus CCRGRG come into contact and hybridize and create today’s
– Tetraploid alpinum of Europe across to Iceland allopolyploids.
REREXX Using molecular sequences, it has been possible to
– Tetraploid alpinum East Asia, the Americas, to reconstruct the origin of the species within section
Iceland RWRWXX Phleum (Stewart et al. 2008). The chloroplast trnL
intron and the nuclear internal transcribed spacer
(ITS) sequences have provided a valuable methodol-
14.4.7.2 P. pratense ogy for tracking the adaptive radiation of diploid
Phleum species as well as up to two of the genomes
– Diploid subsp. bertolonii in northern Europe BNBN in polyploid forms.
– Diploid subsp. bertolonii in Spain and Portugal The results show that an ancestor of P. alpinum
BSBS subsp. rhaeticum RARA is the progenitor of section
– Diploid subsp. bertolonii in Greece and the Balkans Phleum. The widespread circumpolar allotetraploid
BGBG P. alpinum RRXX is a hybrid of this paleogenome with
262 A.V. Stewart et al.

Deschampsia
cespitosa

P. hirsutum,
P. phleoides,
P. iranicum

Ancestral
rhaeticum
central Asia RARA

P. paniculatum
RARAXX
(1 unknown genome)

ITS mutation event

Chloroplast mutation event

rhaeticum Alps of
Austria, Italy,
alpinum 4x Switzerland RSRS
Japan, America RWRWXX
(1 unknown genome) rhaeticum,
Alps of Italy
and Austria

alpinum 4x
Europe, Iceland REREXX alpinum 4x alpinum 4x
(1 unknown genome) Iceland Taiwan

alpinum 4x paternal
Italian Alps rhaeticum
rhaeticum Pyrenees
echinatum EE
Greece diploid RC and tetraploid RPRP
awns Mediterranean
RGRG RRCC hybrids of become mountains
rhaeticum and glabrous
rhaeticum "commutatum" , maternal
Greece Caucusus, Carpathian and
Altau Mountains

diploid
"commutatum"
CC Austria,
Romania 8x pratense
reduction in Southern Italy
telomeric (3 unknown genomes)
heterochromatin R8R8XXXXXX
and anther length
paternal
diploid reduction in
4x pratense, "commutatum" centromeric
Italy BBRR and Austria, heterochromatin
commercial 6x Switzerland and awn length
pratense
BBBBRR
tetraploid hybrid of
maternal 2x bertolonii BB and
6x commercial
pratense BBBR
Alps Italy
bertolonii BSBS
Portugal, Spain,
bertolonii BNBN and 6x pratense
Northern Europe Morocco
BSBSXXXX
hubbardii
Poland 4x pratense
tetraploid BNBNBNBN Southern France
bertolonii France BNBNBSBS

Fig. 14.2 The development of Phleum species from an Asian ancestor of diploid P. alpinum subsp. rhaeticum RARA showing ITS
and cpDNA mutations, hybridizations and polyploidizations
14 Phleum 263

an unknown genome (probably another Phleum spe- would have had a more continuous distribution at
cies from a different section). The three other diploids, lower altitudes during the most recent glaciation event
P. alpinum form “commutatum” CC, P. echinatum (Thorn 1960).
EE, and P. pratense subsp. bertolonii BB, are all
derived from P. alpinum subsp. rhaeticum. In each
case these have undergone molecular, cytological 14.5.1.1 Diploid Phleum alpinum
and morphological changes as well as changes in
adaptation to environmental conditions. On the basis of morphology and molecular form, dip-
loid P. alpinum can be divided into three groups,
subsp. rhaeticum RR, “commutatum” CC, and their
hybrids RC.
14.5.1 Phleum alpinum

P. alpinum L. occurs in most mountains of Europe, Subsp. rhaeticum RR

northern Asia and North and South America. At 30
latitudes, it occurs at altitudes over 4,000 m but this The diploid subsp. rhaeticum RR is recognized by the
reduces to sea level at 60 . Tetraploids are widespread, presence of ciliate awns, a feature where intermediate
while diploids are restricted to Europe (Fig. 14.3). forms can make this distinction difficult. Subsp. rhae-
P. alpinum is considered to be a glacial relict and ticum is the dominant species in Switzerland, but it

Fig. 14.3 Distribution of P. alpinum complex (Conert 1998)

264 A.V. Stewart et al.

occurs throughout the European Alps RSRS and in Of the 21 hybrids discovered, five were a new tetra-
most central and southern European mountain ranges ploid form.
including the Pyrenees RPRP and the Balkans RGRG.
It usually occurs in the subalpine and alpine belt from
1,000 to 2,500 m in fertile and humid habitats (Zernig 14.5.1.2 Tetraploid Phleum alpinum
2005).
The molecular results show that rhaeticum has On the basis of morphology and molecular form, tet-
migrated out from a base population in the Alps raploid P. alpinum can be divided into two groups, a
along adjacent mountain chains to the Pyrenees, Apen- widespread allotetraploid RRXX, as well as the tetra-
nines and the Balkans. Associated with these migra- ploid hybrids of “commutatum” CC, rhaeticum RR
tions are small changes in the genome as typified by the and their hybrids RRCC or CCRR.
reduction in centromeric heterochromatin observed in
the Greek molecular form (Kula 2005a).
The rhaeticum molecular form present in the Alps Tetraploid Euro-American P. alpinum RRXX
RSRS is the progenitor of all the forms within section
Phleum, “commutatum” CC, bertolonii BB, and poly- This form has the widest distribution of any of the
ploid pratense. The notable exception to this is the Phleum species. Not only is it present on many of the
widespread paleo-allotetraploid P. alpinum RRXX. In mountains of Europe and North Asia, it is the only
particular, the molecular form of rhaeticum from the species of the genus to successfully migrate to both
eastern Alps and Greece identifies itself as an ancestor North and South America (Conert 1998).
of all agricultural hexaploid pratense sharing the The molecular data reveal a central Asian origin
RGRG ITS sequence. with a divergent migration east to coastal Asia, Japan
and to the Americas RWRWXX and west into Europe
REREXX. Once in America it migrated rapidly to
Form “commutatum” South America. A recent divergence of the European
and American forms is supported by the lack of signif-
The diploid CC identified by its glabrous awns is icant karyological differences between them (Kula
found in the Alps and north into Germany and the et al. 2006) and only minor ecotypic differences
Czech Republic, as well as the Carpathian mountains (Heide and Solhaug 2001). Its circumpolar migration
from Poland to Romania and Sweden (Joachimiak and was completed in Iceland where derivatives of both
Kula 1996). Diploid “commutatum” CC originates the European and American molecular forms occur.
from rhaeticum of the Alps RSRS but differs from it However, we cannot discount the possibility that the
by a chloroplast DNA insertion glabrous awns, European forms may have been introduced into Ice-
reduced telomeric heterochromatin (Joachimiak 2005), land with human colonization.
reduced anther length and by growing at higher alti- The formation of its allotetraploid ancestor RRXX
tudes among the snow-bed vegetation (Zernig 2005). is likely to have been in Asia over 300000 years BP,
From the geographic location of molecular derivatives, before it diverged into the two forms. The diploid
it is apparent that there has been a general northern behavior of the species and the difficulty in crossing
and eastern radiation outwards from the Alps towards with P. pratense also suggest an ancient formation of
Germany and the Carpathian mountains of Poland and this allotetraploid (Nördenskiold 1945).
Romania.

Tetraploid Hybrids Between rhaeticum

Hybrids Between rhaeticum and “commutatum” and “commutatum” RRCC, CCRR

Of the European diploid P. alpinum analyzed, approx- We report a new form of tetraploid P. alpinum RRCC
imately a third each were rhaeticum RR, “com- and CCRR based on hybrids of rhaeticum and “com-
mutatum” CC, and their hybrids RC or CR. A high mutatum,” which adds further complexity to the many
frequency of hybridization has occurred over time. P. alpinum forms. Five populations were discovered,
14 Phleum 265

one in the European Alps, two in the Caucasus Moun- 2008). Compared with the hexaploid, it is less com-
tains and two in the subalpine meadows of the Altau mon in northern areas, being absent from the Roma-
Mountains. This suggests these hybrids are part of a nian and East Carpathian regions and more common in
continuing eastward migration, extending at least as the south (Humphries 1980), except for Italy where it
far as Kazakhstan. is uncommon (Cenci et al. 1984). It also occurs in the
mountains of North Africa (Maire 1953).
Løhde (1977) and Foerster (1968) describe taxo-
14.5.2 P. pratense nomic differences between diploid subsp. bertolonii
and hexaploid P. pratense but Løhde concedes that the
only reliable taxonomic differences are chromosome
P. pratense is a lowland species distributed naturally
number and ligule hairiness.
throughout Europe, parts of North Africa and Asia
Subsp. bertolonii BB is derived from P. alpinum
(Fig. 14.4). Hexaploids are used for agricultural pur-
subsp. rhaeticum RR and associated with this deriva-
poses in all cool temperate regions of the world.
tion is the loss of awns, loss of centromeric hetero-
chromatin (Joachimiak 2005), and most importantly, a
14.5.2.1 Diploid subsp. bertolonii change in adaptation from subalpine to lowland con-
ditions.
Diploid subsp. bertolonii occurs throughout Europe Two major molecular forms exist within bertolonii,
often sympatric with hexaploid forms (Perný et al. one restricted to Spain and Portugal, BSBS, and a

Fig. 14.4 Distribution of P. pratense diploids and hexaploids (after Conert 1998)
266 A.V. Stewart et al.

second widespread across northern Europe, BNBN. It hybridization (GISH) studies also suggest the presence
is likely that these have diverged as a result of glacia- of two bertolonii genomes (Joachimiak unpublished)
tion events in Europe. The northern European molec- and on the basis of geographic origin these two
ular form is likely to have reinvaded northern Europe bertolonii genomes are most probably both the north-
after the last glaciation from a refuge in Italy or the ern European form making hexaploid pratense
Balkans. BNBNBNBNRGRG. Chromosome pairing in triploid
A particularly stoloniferous form of bertolonii in plants of hexaploid pratense (7II þ 7I) (Nördenskiold
central Europe has been named Phleum hubbardii 1945) suggests at least minor differences in the struc-
(Kovats 1977, 1980). Its molecular ITS sequence is ture of the bertolonii B and rhaeticum R genomes. The
derived from the northern European bertolonii BNBN, small difference between these two genomes readily
but differs with a mutation at one position. This minor allows a synthesized auto-hexaploid bertolonii to cross
difference and the lack of any significant differences in with natural hexaploid P. pratense (Nördenskiold
karyotype (Kula 2003) suggest that P. hubbardii is not 1957). The uniformity of the molecular profile in agri-
a very well differentiated taxon. cultural P. pratense suggests that the formation of this
hexaploid is probably post-glacial. Its distribution sug-
gests it has expanded throughout Europe from a glacial
14.5.2.2 Tetraploid P. pratense refuge, most likely from the Balkans/Italy glacial ref-
uge (Hewitt 1999).
Tetraploid P. pratense has been reported to occur in Two different hexaploid molecular forms were also
central and northern Italy, France, Belgium, Spain, found in southern Italy RGRGXXXX and Morocco
Poland, Slovakia and Bulgaria (Kozuharov and Petrova BSBSXXXX. Our results show that hexaploid pratense
1991; Uhrı́ková and Králik 2000; Joachimiak 2005). is polyphyletic in origin, having formed at least three
We have found four molecular forms of tetraploid times from different diploid ancestors, a situation very
pratense, two different allotetraploids and two autote- common in polyploid species (Soltis and Soltis 2000).
traploids. Two allotetraploid pratense were discovered
in the Alps, one a hybrid of bertolonii with rhaeticum
BBRR and the other a hybrid of bertolonii with hexa- 14.5.2.4 Octoploid P. pratense
ploid pratense BBBR. One autotetraploid pratense
from southern France has molecular characteristics Octoploid pratense R8R8XXXXXX is reported only
of both northern European and Spanish bertolonii from southern Italy (Cenci et al. 1984). Two samples
BSBSBNBN, while the second French population exhi- with the very short stature of diploid bertolonii have
bits only northern European bertolonii BNBNBNBN the maternal genome derived from rhaeticum R, but it
molecular characteristics. is not possible to determine from our results the origin
of the other three genomes.

14.5.2.3 Hexaploid P. pratense

Hexaploid pratense can be divided into three molecu- 14.5.3 P. echinatum

lar forms. The most common is the agricultural hexa-
ploid with the cytoplasmic molecular pattern of The annual P. echinatum EE occurs in eastern Medi-
bertolonii BB and an ITS molecular pattern from the terranean mountains ranging from Sicily to Crete
Balkans rhaeticum RGRG present from Greece to the (Humphries 1980). Our results show that this species
eastern Alps. Although it is not possible from our has developed from hybridization between two differ-
results to determine the origin of the third genome, ent derivatives of rhaeticum. Furthermore, it is likely
there is cytological and molecular evidence to suggest that it originated from a single hybridization event as
that this third genome is homologous with one of the this species has undergone genetic reconstruction from
two genomes (M€ untzing and Prakken 1940; Levan the genus norm of 14 chromosomes to 10 (Ellestrom
1941; Nördenskiold 1945; Cai and Bullen 1991, and Tijo 1950). Surprisingly, dysploid reduction from
1994; Joachimiak 2005). More recent genomic in situ x ¼ 7 to x ¼ 5 was accompanied with the considerable
14 Phleum 267

increase of genome size in this species. The species Migration also occurred onto the colder mountain
exhibits some features of P. alpinum subsp. rhaeticum, ranges to the north into Germany and to the Carpathian
but has a longer awn and reduced centromeric hetero- Mountains of Poland and Romania but was associated
chromatin (Joachimiak 2005). with micro-evolutionary changes in morphology and
cytology to develop into diploid “commutatum” CC.
The overlap of the range of rhaeticum and “commu-
tatum” has since allowed considerable hybridization
14.5.4 Migration History in Relation so that a swarm of hybrids RC or CR overlaps the
to Glaciation Events range of “commutatum” and part of the rhaeticum
range. Occasional tetraploid hybrids RRCC or CCRR
have developed and have migrated east at least as far
The molecular results show an Asian origin for the as Kazakhstan.
section Phleum and identify two separate migrations Migration of rhaeticum populations back into the
into Europe. lowlands as a result of climate cooling eventually
The first migration into Europe was of an ancestor resulted in the first lowland species of this group,
of diploid P. alpinum subsp. rhaeticum RR. The pen- P. pratense subsp. bertolonii BB. This was also accom-
ultimate Riss glaciation 130000–150000 year BP panied by micro-evolutionary changes in cytology,
provided ample opportunity for this alpine species to morphology and adaptation. As the climate cooled
migrate vast distances through lowland areas to even- during the last glaciation (the W€urm 22000 to 13000
tually become isolated on the Alps RSRS during years BP) this lowland species retreated into southern
subsequent warmer interglacial periods. Subsequent European glacial refugia. Upon warming these subse-
migration along mountain ranges has occurred so quently reinvaded northern Europe from the Balkan/
that today rhaeticum occurs in the Alps RSRS, Pyre- Italy refugia as molecular form BNBN, with a second
nees RPRP, Apennines RIRI, and the Balkans RGRG. molecular form BSBS remaining restricted to the

Fig. 14.5 Generic glacial refugia of southern Europe (shaded areas) (after Hewitt 1999) and potential post-glacial migration route
of diploid ssp. bertolonii and agricultural hexaploid pratense
268 A.V. Stewart et al.

Spanish glacial refuge. Hybridization occurred when although it is unexpected for many people that a spe-
these forms met at the interface in France resulting in a cies so common in high latitude cold temperate zones
recent autotetraploid BSBSBNBN. has originated within southern European glacial ref-
Hybrids formed in the Italian Alps where subsp. ugia and that these areas still retain high genetic
bertolonii and the Balkans rhaeticum overlapped diversity today. Although it has been suggested that
resulting in an allotetraploid pratense BNBNRGRG. It northern Europe is a center of diversity (Guo et al.
is probable that a further hybridization with the adja- 2003), this appears unlikely as it has only been free of
cent northern European subsp. bertolonii BNBN lead to glaciation in the last 12,000 years and any diversity
the formation of the agricultural hexaploid pratense must be recent, or of migratory origin.
BNBNBNBNRGRG. Upon warming in the holocene
these subsequently reinvaded northern Europe from
the Balkan/Italy refugium, a refugium common to a
wide range of European biota (Hewitt 1996, 1999) 14.5.5 Gene Exchange Between Genomes
(Fig. 14.5). and Inheritance
Two further hexaploid forms and an octoploid
occur within glacial refugia, all based on local diploids Hexaploid P. pratense has many examples of tetradi-
but in all cases the remaining genomes are unknown, a somic or hexasomic inheritance (Nördenskiold 1953;
hexaploid in southern Italy RGRGXXXX, another Nielsen and Smith 1959; Nath 1967). This means that
hexaploid in Morocco BSBSXXXX and an octoploid there is considerable gene exchange between genomes
in southern Italy R8R8XXXXXX. and that, while genomes can be identified and an allo-
Hybridization among rhaeticum forms led to the hexaploid genomic formula can be assigned BNBNBN
Mediterranean mountain annual P. echinatum EE BNRGRG, the genomes are not well differentiated. This
with a reconstructed genome of only 10 chromosomes. is consistent with many studies concluding that
This event probably took place prior to the last glacia- P. pratense is largely an autopolyploid (Nielsen and
tion enabling it to spread throughout the Balkan/Italy Nath 1961; Nath 1967; Nördenskiold 1953, 1957,
glacial refuge and to subsequently recolonize the 1960; Wilton and Klebesadel 1973) and that autohex-
mountains of the eastern Mediterranean as the climate aploids developed from diploid P. pratense subsp. ber-
warmed. tolonii are very similar to the agricultural hexaploid
The very widespread allotetraploid P. alpinum P. pratense (Nördenskiold 1949).
formed over 300000 years BP in Asia from hybridiza- From our new molecular knowledge on genomic
tion of an ancestral rhaeticum with another unknown origins of P. pratense hexaploids we now know that all
genome, RARAXX. This form remained in Asia until three genomes are derived from the rhaeticum RSRS
eventually migrating into Europe during the last glaci- genome marooned in Europe around 150,000 years
ation (the W€urm 22000 to 13000 years BP) when con- ago. The differentiation of this subalpine genome
ditions were suitable. At the same time many species RSRS into the Balkans form RGRG and the lowland
including this one were able to migrate into the Amer- derivative BB genome may be much less than 50,000
icas via the Bering/Aleutian route, although probably years. For this reason there is only minor differentia-
not completing their entry until this route became open tion between these genomes and it is to be expected
around 8,000 years ago (Hong et al. 1999; Weber that P. pratense behaves largely as an autopolyploid.
2003). This divergent migration has lead to a diver- On the other hand the widespread paleo-allotetra-
gence in molecular forms, one in northern Europe ploid P. alpinum RRXX had both greater differentia-
REREXX and the other in Japan and the Americas tion between the genomes and a longer period of time,
RWRWXX. The circumpolar migration was completed probably over 300,000 years, to become stabilized as
in Iceland where derivatives of both forms occur. an allotetraploid (Nördenskiold 1945). However other
Dogan (1991) describes the Mediterranean and more recent tetraploid P. alpinum forms based on
western Asia as the center of origin for the genus hybrids of rhaeticum and “commutatum” should
Phleum. This remains true for P. pratense today, behave more like autotetraploids.
14 Phleum 269

14.6 Wild Relatives as Genetic sufficiently winter-hardy for use in more northern
Resources for Phleum pratense European climates. However, such material deserves
special attention as it represents unique genomic hexa-
ploid constitutions, and genetic erosion in these south-
P. pratense genetic resources can be divided into pri- ern regions is occurring at an alarming rate as many in
mary, secondary, tertiary and quaternary resources as situ populations are under threat from climate warm-
follows. ing and human-induced habitat degradation. Gene-
bank material from these southern regions is very
limited and we know of only one sample available
from North Africa and only two from southern Italy.
14.6.1 Primary Gene Pool The senior author has found that the Moroccan
BSBSXXXX and the southern Italian RGRGXXXX
This is the major pool of hexaploid P. pratense germ- both readily cross with commercial hexaploid pra-
plasm used by breeders, consisting of those adapted tense BNBNBNBNRGRG.
cultivars, elite breeding lines and local ecotypes
within the regions where the species is used. This
is almost exclusively the common genomic form
14.6.3 Tertiary Gene Pool
BNBNBNBNRGRG used in northern Europe, northern
Asia and North America and to a much lesser extent
New Zealand. The molecular diversity of the This consists of germplasm derived entirely from the
European lines exhibits a geographic pattern and deri- original P. alpinum subsp. rhaeticum genome RARA
vatives in Asia, Americas and New Zealand can be but of ploidy levels other than hexaploid. P. pratense
related to these (Yang-Dong et al. 2003). The vast diploid BB, tetraploid BBBB and BBRR, octoploid
majority of hexaploid P. pratense accessions in gene- RRXXXXXX as well as diploid rhaeticum RR, “com-
banks, potentially over 99%, would be of this common mutatum” CC, and the diploid and tetraploid hybrids
genomic form. As there is considerable exchange of of rhaeticum and “commutatum” RC and RRCC and
germplasm among breeders in different regions it CCRR. The widespread paleo-allotetraploid P. alpi-
would probably be fair to conclude that this pool of num RRXX would be excluded from this pool and
germplasm has been extensively exploited by bree- classified in the quaternary pool.
ders. Yet by and large it represents only the winter- With the current knowledge of the genomic constitu-
hardy northern genomic form of P. pratense and fails tion of P. pratense and other forms it should be possible
to utilize the material from the center of origin in to either resynthesize P. pratense from different forms
southern regions of Europe and North Africa. of the same genomes, or use genomes from other forms
and ploidy levels of section Phleum for introgression
into P. pratense hexaploids. While there do not appear
to be any major barriers to hybridization between
14.6.2 Secondary Gene Pool Phleum species or forms (Nath 1967), in practice the
major barrier is one of ploidy and/or allopolyploidy.
This pool includes hexaploid germplasm of P. pra- Crosses between diploid bertolonii and hexap-
tense from the center of origin in the Balkans/ loid P. pratense are easy to produce artificially
Italy region of southern Europe, the Mediterranean (Nördenskiold 1945), and although not always tetra-
mountains and North Africa, a region largely outside ploid (Løhde 1977), BNBNBNRG are found in nature
the region of commercial use. It would also include (M€untzing 1935; Foerster 1968, 2005). These readily
other genomic constitutions of hexaploid P. pratense cross back to hexaploid P. pratense.
such as the southern Italian RGRGXXXX and Hybrids between tetraploid P. pratense BNBNRG
Moroccan BSBSXXXX (potentially an autohexaploid RG and the octoploid R8R8XXXXXX have been made
BSBSBSBSBSBS). by the senior author and these form fertile hexaploid
These regions have never been targeted for gene- plants. These plants have been crossed with regular
bank collection because this material is not considered agricultural hexaploids BNBNBNBNRGRG and these
270 A.V. Stewart et al.

also form fertile progeny. Some sterility is apparent former glacial refugia. Even in many of the northern
from low seed yield and from ergot in the seed sam- European agricultural regions older fields are replaced
ples, a factor encouraged by the sterile florets remain- as agriculture intensifies (Weibull et al. 2007). These
ing open for very long periods of time. changes make the ex situ conservation in genebanks
Hybrids of tetraploid and hexaploid forms are pen- and breeders’ seed collections very critical for the
taploid as expected (Nielsen and Nath 1961) and these future.
may be backcrossed to hexaploids quite readily.
The tertiary genepool potentially represents an
enormous pool of unexplored material for breeders,
but collections are necessary. 14.6.6 Germplasm Banks

Today there are approximately 120 cultivars of

14.6.4 Quaternary Gene Pool P. pratense certified under the OECD seed scheme
around the world. There could be potentially another
30 or more outside this scheme as well as an estimated
By definition these would consist of the more difficult- 50 older cultivars no longer used, making probably no
to-cross material such as allotetraploid P. alpinum more than 200 named cultivars of Timothy.
RRXX as well as species in other sections, such as Most of these cultivars are preserved in genebanks
P. phleoides in section Chilochloa (of which there is along with collections from old pastures in the regions
an herbarium sample from Iceland in the University of where Timothy is used commercially. Germplasm
Leiden, National Herbarium, Netherlands) and P. hirsu- collections generally preserve seeds of Phleum under
tum in section Chilochloa (which the senior author has refrigerated conditions and cryopreservation has not
crossed with hexaploid P. pratense to form tetraploids), been carried out to any extent.
and P. subulatum of section Achnodon. The hybrid with There are more than 6,000 accessions of Phleum
P. subulatum formed a male sterile tetraploid morpho- around the world in genebank collections. Of these
logically similar to P. pratense (Myers 1941). over 98% are P. pratense and less than 2% are
There is also a report of a sterile cross between P. alpinum and other species. Major collections occur
tetraploid Dactylis glomerata and hexaploid P. pra- in Europe (>4,800), USA (>550), New Zealand (>300),
tense (Nakazumi et al. 1997). Canada (>80) and Japan (>40). It is impossible to
At this stage the quaternary gene pool offers much determine the number of unrelated accessions due to
less potential for breeders than the secondary and high rates of duplication within and between gene-
tertiary gene pools and collection and crossing efforts banks.
would be much better targeted at these. It is of concern that there are so few P. pratense
subsp. bertolonii and diploid P. alpinum accessions
from Europe as well as so few southern European
hexaploid P. pratense accessions.
14.6.5 The Conservation of Genetic
Resources

The conservation of a full range of genetic resources is

pivotal to future improvements in P. pratense. Ulti- 14.6.7 Development of Core Collections
mately the best method of conservation would be in
situ in the natural and agricultural world. However, in An important process of germplasm conservation is to
many regions Phleum species face an enormous threat prioritize the collection into a core collection to allow
from changing land use as well as climate change. In maintenance of a relatively full range of diversity
particular, these two factors place a serious threat to the using a more manageable number of accessions.
many isolated mountain populations of P. alpinum There are many factors to consider in the develop-
and to P. pratense in the Mediterranean mountains of ment of a core collection. These include morphological,
14 Phleum 271

geographic, edaphic and molecular diversity as well as 14.7 Cytological, Genomic and
an extreme variation of key individual traits. Casler Molecular Resources in Phleum
(2001) showed very strong relationships between geo-
graphic origin of germplasm and a series of morpho-
logical traits and he was able to develop a core There have been numerous cytological studies in
collection on this basis. Phleum, largely in an effort to understand the relation-
Now that we have more understanding of the geno- ship between the genomes in P. pratense, for example
mic constitutions and diversity, as well as the geo- as described by Joachimiak (2005). The B-chromo-
graphic variation due to glacial driven migrations, it somes have also been studied (Bosemark 1967; Fröst
should be possible to assemble a more representative 1969; Joachimiak 1982, 1986). There have been no
core collection. However, to do this it will require the attempts to develop chromosome substitution or dele-
collection of Phleum resources in the glacial refugia of tion lines.
southern Europe and North Africa. These forms are Anther culture methodology has been developed
seriously lacking in any collection. and triploid and hexaploid regenerants have been
obtained (Abdullah et al. 1994).
Molecular techniques have been developed for
phylogenetic and genomic identification (Stewart
et al. 2008). Cai et al. (2003) developed a set of simple
14.6.8 Endophytic Fungi sequence repeat (SSR) markers and a team in Finland
announced its intention in 2006 (Manninen et al. 2006)
Many grasses host choke forming endophytic fungi of to use a candidate gene approach, bulked segregant
the genus Epichlöe or their non-choking asexual deri- analysis and gene expression chips to find modern
vatives Neotyphodium. The sexual stroma of Epichlöe genetic tools applicable to Timothy breeding. This
cause “choke disease,” which prevents seedheads has resulted in a publication on the development of
emerging. Non-choke inducing asexual Neotypodium markers for feeding quality and gray snow mould
endophytes occur in many grasses such a perennial resistance (Tanhuanp€a€a et al. 2007, 2008).
ryegrass (Lolium perenne) and tall fescue (Festuca A study on genetic resources of Nordic Timothy is
arundinacea) and they provide the host with advan- underway in Norway using molecular techniques
tages such as improved insect resistance and grazing (Fjellheim et al. 2007).
deterrence (Schardl and Phillips 1997). Although no genetically modified Timothy cultivar
Timothy hosts a partial choke forming Epichlöe has been developed, Timothy has been included in a
typhina endophyte and in this situation infected plants number of patents for genes for disease resistance,
suffer from less infection with the leaf disease Clados- herbicide tolerance and allergy studies.
porium phlei (Seto et al. 2005). In Finland, 33% of plants
were found to be infected with endophyte (Saikkonen
et al. 2000).
In Patagonia P. alpinum is known to host the non-
14.8 Recommendations for Future
choking asexual Neotyphodium tembladerae (Moon
et al. 2004; Gentile et al. 2005). This provides infected Actions
plants with an advantage over uninfected plants in
some circumstances (Cabral et al. 2007). In this situa- The current understanding of the genomic constitution
tion the infection with Neotyphodium represents a within Phleum should allow breeders to utilize the
symbiotic relationship whereby the plant gains protec- genetic resources more effectively than previously. It
tion from insects and overgrazing while the endophyte should now be possible to resynthesize new forms and
obtains nutrition. The full relevance of endophyte to introgress a much wider range of diversity into
fungi in Phleum is not known. However, it is important P. pratense than has occurred naturally. However,
that endophytes are collected and studied. Germplasm many of the genetic resources of wild relatives are
collections will need to monitor and maintain endo- under threat from climate warming and human
phytes where they are present. induced habitat degradation (t’Mannetje 2007).
272 A.V. Stewart et al.

There is an urgent need to collect hexaploid Cai Q, Bullen MR (1994) Analysis of genome specific sequences
P. pratense germplasm from Mediterranean mountain in Phleum species: identification and use for study of genomic
relationships. Theor Appl Genet 88:831–837
glacial refuge areas as well as a wide range of geneti- Cai HW, Yuyama N, Tamaki H, Yoshizawa A (2003) Isolation
cally diverse diploid, tetraploid and octoploid P. pra- and characterization of simple sequence repeat markers in
tense and of the readily crossable forms of P. alpinum. the hexaploid forage grass timothy (Phleum pratense L.).
These include diploid subsp. rhaeticum and “commu- Theor Appl Genet 107:1337–1349
Caradus JR (1978) Plant introduction trials. Performance of
tatum” as well as their diploid and tetraploid hybrids. Timothy cultivars and lines in New Zealand as spaced plants.
These collections should be integrated into core NZ J Exp Agric 6:11–17
collections to maximize molecular diversity of the Casler MD (2001) Patterns of variation in a collection of Timo-
available genomes. thy accessions. Crop Sci 41:1616–1624
Cenci CA, Pegiati MT, Falistocco E (1984) Phleum pratense
It is also important that each of the major regions (Gramineae): chromosomal and biometric analysis of Italian
where Timothy is used maintains strong functional field populations. Willdenowia 14:343–353
breeding programs to allow adequate cultivar develop- Charlton JFL, Stewart AV (2000) Timothy, the plant and its use
ment, germplasm collection, introgressions of wild on New Zealand farms. Proc NZ Grassl Assoc 62:147–153
Conert HJ (1998) Phleum. In: Hegi G (ed) Illustrierte Flora von
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Cooper JP (1958) The effect of temperature and photoperiod on
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port of PGG Wrightson Seeds and AgResearch, and the numer- spp.). J Br Grassl Soc 13:81–91
ous people who supplied germplasm. Cooper JP, Calder DM (1964) The inductive requirements for
flowering of some temperate grasses. Grass Forage Sci
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lemma in Phleum, Pseudophleum and Rhizocephalus (Gra-
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Chapter 15
Setaria

Henri Darmency and Jack Dekker

15.1 Introduction focuses on foxtail millet only. It is generally classified

into three races: maxima, indica and moharia (Prasada
Rao et al. 1987). Races maxima and indica are typi-
Foxtail millet, Setaria italica (L.) Beauv., is a small
cally used as a cereal, while race moharia is used as
grain C4 cereal of the Paniceae tribe of Poaceae family
forage.
(Fig. 15.1). It is one of the few economically important
As far as we know, the use of wild Setaria in
species of Setaria. It has long been important in China
breeding programs to improve foxtail millet is very
and India for human consumption. It is also grown in
scarce. In contrast, wild Setaria were invaluable
small quantities throughout Eurasia for some tradi-
resources to gain better knowledge of the genome
tional uses and for feeding birds, and in Europe and
organization of the crop. Exploiting wild and weedy
America for hay and silage. Grains have been discov-
Setaria species has not gained the interest of crop
ered in Neolithic relics (circa 8,000 years ago) in
breeders for several reasons. Perhaps, the fact that
various places of China, along the upper and middle
most of the research and achievements have been
valleys of the Yellow river and the Yangtze River (Li
carried out in China, India, and Japan led to the release
and Wu 1996; Nasu et al. 2007) and in Europe (Naciri
of the corresponding reports written in local languages
and Belliard 1987). A small number of Setaria species
and thus unidentified in recent worldwide databases
are closely related and constitute a narrow gene pool
(Gu 1987; Li 1995). Another reason of the rare use of
potentially useful for foxtail millet genetic improve-
the wild germplasm could be the structure of the
ment. Besides S. italica, other Setaria are grown as
wild–weed–crop complex providing enough genetic
crop or forage (De Wet et al. 1979; Austin 2006). It
diversity through “off-types” to satisfy new breeding
includes Setaria pumila partially domesticated and
plans. In addition, the huge diversity of landraces
cultivated in mixed stand with other millets in India
collected in China (23,400; Li and Wu 1996), India
(Kimata et al. 2000), S. sphacelata is used as wild
(1,951; Seetharam 1998) or Japan (NIAS 2009) made
cereal in Africa and planted as a tropical pasture spe-
available a quantity of interesting material to modern
cies (Jank et al. 2007), and S. parviflora is used as a
breeders, which resulted in poor interest in wild
cereal and is found in the pre-historic remains in
Setaria. Collection and characterization of landraces
Central America (Austin 2006). The present chapter
were carried out through China (Li and Wu 1996),
India (Upadhyaya et al. 2008), Russia (Varadinov
1986), France (Nguyen Van and Pernès 1985), and
Japan (Nakayama et al. 2005). Finally, the crop foxtail
millet is not traded on world grain exchanges, while
H. Darmency (*) it remains an important grain crop for indigenous,
UMR 1210 Biologie et Gestion des Adventices, Institut National
often poorer, populations in Eurasia, Africa and the
de la Recherche Agronomique, 17 rue Sully, BP 86510, 21065
Dijon, France Americas, and is therefore not of interest to main-
e-mail: [email protected] stream crop improvement projects.

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 275
DOI 10.1007/978-3-642-14255-0_15, # Springer-Verlag Berlin Heidelberg 2011
276 H. Darmency and J. Dekker

Fig. 15.1 Mature infrutescences (ears) of foxtail millet and green foxtail

15.2 The Complex of Species of parts of the world and is of significant agricultural
Foxtail Millet importance as crops and also as noxious weeds
(Rominger 1962; Prasada Rao et al. 1987). The taxon-
omy of the genus is very complex, and an accurate
15.2.1 A Brief Glance classification has been confounded by the high degree
of overlapping morphological characters both within
Worldwide there are 125 Setaria species divided and between species, and the diverse polyploidy levels
among several subgenera (Hubbard 1915; Rominger within the genus. One of the most coherent taxonomic
1962). The genus, Setaria P. Beauv. (the foxtails), is treatments was given by Rominger (1962). There is
widely distributed in tropical, subtropical, and temperate evidence that the genus Setaria is actually divided into
15 Setaria 277

two groups, or clades (Doust and Kellogg 2002). This continuous and overlapping genetic variation, the evi-
reassessment of the genus would place S. pumila and dence of interfertility, and the weedy origins of the
S. parviflora into a common (S. pumila) clade, and crop (see Sect. 15.6.1) greatly support the hypothesis
S. italica, S. viridis, S. faberi and S. verticillata in of a common “biological species” status, namely
another (S. viridis). S. viridis, with two subspecies: spp. italica (foxtail
Observations drawn from karyological studies and millet), and spp. viridis (green foxtail). Beside a long
interspecific hybridization suggest that the complex history of nomenclatural changes, including that both
of species of foxtail millet is organized in three gene taxa be considered as subspecies of S. italica, which
pools (Fig. 15.2). Closest to S. italica is its putative wild implied the primacy of the crop over that of its wild
ancestor, S. viridis (L.) P. Beauv., the green foxtail, Setaria ancestors, the correct binomial to be used now
which forms the primary gene pool at a diploid level maintains the two taxa at the species level, S. viridis,
with the A genome (Li et al. 1945; Harlan and De Wet and S. italica.
1971; Darmency et al. 1987b; Benabdelmouna et al. Present nomenclatural authorities are presented in
2001a). The secondary gene pool corresponds to the Table 15.1 for the complex of species of foxtail millet.
species carrying the B genome. It includes the diploid Besides, there are many nomenclaturally designated
S. adhaerans (Forsskal) Chiov. (bristly grass), and the forms or racial taxonomies below the subspecies level
tetraploid species S. verticillata (L.) P. Beauv. (bristly that have been proposed based on morphology rather
foxtail) and S. faberi F. Hermann (giant foxtail). A than genetic distance (Rominger 1962; Douglas et al.
tertiary gene pool could tentatively contain S. pumila 1985). For instance, S. viridis ssp. pycnocoma should
(Poiret) Roemer & Schultes (yellow foxtail) in addition not be considered as a fixed narrow taxon since numer-
to many other wild species with unidentified genomic ous weedy forms may originate independently through
composition, but no link has been established yet. All natural crossing, as indicated in Sect. 15.3.3.2.
these species are annual, typically weedy, and origi-
nated from the temperate regions of the northern hemi-
sphere. Reviews of the weedy Setaria species-group Table 15.1 Karyological data of Eurasian taxa of the complex
(Dekker 2003, 2004), as well as three of individual of species of the foxtail millet
species have been published previously in the Biology Species Most frequent Genome
ploidy level
of Canadian Weeds series (S. pumila and S. verticillata,
S. italica (L.) P. Beauv. 2n ¼ 2x ¼ 18 AA
Steel et al. 1983; S. viridis, Douglas et al. 1985; S. viridis (L.) P. Beauv. 2n ¼ 2x ¼ 18 AA
S. faberi, Nurse et al. 2009). S. viridis spp. pycnocoma 2n ¼ 2x ¼ 18 AA
(Steudel) Tzveleva
S. adhaerans (Forsskal) Chiov. 2n ¼ 2x ¼ 18 BB
S. adhaerans var. antrorsa 2n ¼ 2x ¼ 18 BB
15.2.2 Taxonomy (A. Braun) H. Scholz
Spontaneous F1 hybrid S. viridis 2n ¼ 2x ¼ 18 AB
 S. adhaerans
The genus Setaria belongs to the tribe Paniceae, sub- S. verticillata (L.) P. Beauv. 2n ¼ 4x ¼ 36e,f AABB
family Panicoideae and family Poaceae. Linnaeus S. verticillata var ambigua 2n ¼ 4x ¼ 36 AABB
(1753) was the first to recognize foxtail millet and (Guss.) Parl.d
S. viridis as two independent species, Panicum itali- Spontaneous F1 hybrid 3x ¼ 27 AAB
cum (foxtail millet) and P. viride (S. viridis). However, S. verticillata  S. italica
S. faberi F. Hermann 2n ¼ 4x ¼ 36 AABB
numerous observations and experiments on their S. pumila (Poiret) Roemer & 2n ¼ 4x ¼ 36g,h,i ?
Schultesc
S. parviflora (Poir.) Kerguélend 2n ¼ 36 j ?
S. adhaerens
S. italica
Frequent synonymous: aS. viridis var. major, bS. verticilliformis,
c
S. verticillata ? S. lutescens and S. glauca, dS. geniculata. Variant 2n numbers:
S. viridis
e
2n ¼ 18 (Pohl 1962; Osada 1989), f2n ¼ 54 (Khosla and
S. faberi Sharma 1973; Singh and Gupta 1977; Osada 1989), g2n ¼ 18
(de Wet 1954; Wang et al. 2007), h2n ¼ 72 (Brown 1948;
Primary gene pool Secondary gene pool Tertiary gene pool Rominger 1962; Khosla and Sharma 1973; Wang et al. 2007),
Genome A Genomes A and B
i
aneuploidy, hypertetraploidy n ¼ 22, n ¼ 19 þ 03B (Khosla
and Sharma 1973); j2n ¼ 72 (Brown 1948; Rominger 1962;
Fig. 15.2 Putative gene pools of foxtail millet Osada 1989; Wang et al. 2007)
278 H. Darmency and J. Dekker

Darmency (2004) discussed all the conditions that to be conspecific. The analysis of ribosomal genes’
have been met to establish a dynamic and evolving sequences confirmed the closest relationship of S. ita-
system known as a “wild–weed–crop complex” and lica and S. viridis accessions from the same geograph-
comprising S. viridis, S. viridis ssp. pycnocoma (as the ical region (Benabdelmouna et al. 2001b). Wang et al.
descendants of interspecific hybrids) and S. italica. (1998) compared both intra and interspecific restric-
It is likely that varieties such as robusta-alba and tion fragment length polymorphism (RFLP) based
robusta-purpurea described in America belong to maps of S. italica and found that these maps were
that dynamics. Indeed, early morphological and chem- very similar in length, show the same order of the
ical methods indicated that “robust” foxtails are more RFLP markers and also the same genetic distances
closely related to the giant green foxtail than to the between the loci. This strongly indicates that there
green foxtail (Williams and Schreiber 1976). In addi- have been no major chromosomal rearrangements
tion, all these subspecies and varieties are completely between the genomes of S. italica and S. viridis.
embedded within the S. viridis range of variation in a Finally, a genomic-based chromosome analysis, geno-
multiple isozyme loci analysis (Wang et al. 1995a). mic in situ hybridization (GISH), showed that the
Another case of taxonomic trouble arises with genomes of S. viridis and S. italica were closely
S. adhaerens and S. verticillata, both species involving related and could not be separated, while there was a
the majority of vouchers with retrorsely barbed bris- clear absence of homology between S. italica and
tles, but a few display antrorse barbed bristles, which S. adhaerans (Benabdelmouna et al. 2001a). Whether
lead to various taxa levels (for instance var. ambigua S. italica and S. viridis should be considered as con-
¼ S. verticilliformis) although it is a simple genetic specific or as subspecies is a matter of taxonomy rules.
attribute. The different ploidy levels also, because they The only justification for keeping the two taxa as
are not apparent in the phenotype, add trouble in the separate species is the domestication syndrome that
correspondence between the botanical descriptions makes morphology, growth and agronomic use of
and the actual genetic nature. foxtail millet very different in both taxa.
The GISH techniques also provided a powerful tool
to revise the phylogeny of the secondary gene pool
of the foxtail millet complex. S. adhaerens was con-
15.2.3 Phylogeny firmed to belong to that secondary gene pool, S. faberi
was confirmed as allotetraploid (after the hypothesis
Harlan and de Wet (1971) suggested that S. italica and of Li et al. 1942), but S. verticillata appeared to
its assumed wild ancestor S. viridis are taxonomically share the same allotetraploid genome as S. faberi
the same species. Indeed, several studies showed that (Benabdelmouna et al. 2001a). Thus, the fundamental
a limited number of differential traits have been difference between the two allotetraploids, S. faberi
involved in domestication (Li et al. 1945; Darmency and S. verticillata, seems not to be linked to their geno-
and Pernès 1987). Such a strong link is also supported mic composition. Perhaps, the difference between the
by results of comparative studies between S. italica two allotetraploid species could be a matter of geo-
and S. viridis, which showed that the storage proteins graphical origin or genetic difference among parents
and isozyme differentiation between the two types was for a few genes encoding morphological traits. The
regional rather than taxonomic: S. italica and S. viridis comparison of the sequences of the ribosomal genes
from the same geographic region were more closely adds evidence of the phylogenetic relationships
related than S. italica accessions from different between the diploid and the allotetraploid Setaria
regions (De Wet et al. 1979; Jusuf and Pernès 1985; (Benabdelmouna et al. 2001b).
Gao and Chen 1988). More recently, molecular data The origins of S. parviflora (Poir.) Kerguélen (syn.
also support the close relationship of the cultivated S. geniculata P. Beauv.) are of particular note.
species S. italica and the wild species S. viridis. S. parviflora, a perennial, is the only weedy foxtail
Using random amplified polymorphic DNA (RAPD) native to the New World, and very closely resembles
analysis among and within species, Li et al. (1998) S. pumila, an annual species of Eurasian origin
concluded that S. italica is more closely related to (Rominger 1962; Wang et al. 1995b). S. parviflora
S. viridis than to other species of Setaria and appears and S. pumila are frequently misidentified by weed
15 Setaria 279

managers. The primary morphological differences clear discrimination between the A and the B genomes
between S. parviflora and S. pumila are found in the was efficient (Benabdelmouna et al. 2001a). A similar
spikelet length and lower floret, palea, and the pres- pattern was observed between S. italica and S. adhae-
ence of short rhizomes (Hitchcock 1971). S. parviflora rens, but no difference was detected between S. italica
has a high degree of sterilization in the fascicle, unlike and S. viridis, thus confirming their genetic proximity.
any other species native to North America, but similar This technique was also applied on polyploid Setaria
to tropical Setaria species. It has been suggested species and enabled to confirm the assumed AABB
that the cause of this enigma was a very ancient dis- allotetraploid nature of S. faberi (Li et al. 1942) and to
persal event westward from the Old to New World demonstrate that both S. verticillata and S. verticillata
(Rominger 1962). var. ambigua were also allotetraploids of the same
genomes AABB (Benabdelmouna et al. 2001a). Hexa-
ploid accessions of S. verticillata have not been
included in this study so that the question remains
15.3 Genetics Attributes open about a third genome or partial autoploidy,
which could explain the abnormal meiosis observed in
these plants (Gupta and Yashvir 1973). Interestingly,
15.3.1 Cytogenetics and Karyotypes S. queenslandica, a species originating from Australia,
where both S. viridis and S. italica were imported,
The genome size of Setaria is one of the smaller one of showed to be the first recorded autotetraploid composed
cereals, with rice (Devos et al. 1998). S. italica and of genome AAAA (Wang et al. 2009b). The polyploid
S. viridis have the same DNA content, 2C ¼ 0.51 pg S. pumila was shown to bear an unknown genomic
(Le Thierry d’Ennequin et al. 1998). The most fre- composition, which is not closely related either to
quently reported number base of chromosomes ever genome A or to genome B (Benabdelmouna et al.
reported is x ¼ 9 (Singh and Gupta 1977). Setaria 2001a), but no study was carried out to investigate a
species of the foxtail millet complex includes the poly- possible homology to some part of genomes A and B,
ploid series of 2x (S. italica, S. viridis, S. adhaerens), 4x which should be helpful to determine if S. pumila could
(S. faberi, S. pumila, S. parviflora, S. verticillata), 6x be used in breeding foxtail millet. A third, different,
(S. verticillata), and 8x (S. pumila, S. parviflora) possi- genome, designated as genome C, was recently discov-
bilities (Table 15.1). The tetraploid level contains the ered in S. grisebachii that originated from Mexico, but
majority of counts recorded for S. verticillata and there is no information about its potential crossing to
S. pumila. Diploid counts of S. verticillata could be foxtail millet as yet (Wang et al. 2009b). The character-
attributed to the diploid S. adhaerens with similar mor- ization of Setaria species originating from Africa and
phology (Wang et al. 2009b). However, the availability America could bring new data about the genetic prox-
of chromosome races in the same taxon raises important imity within the genus. For instance, the close similarity
questions about the taxonomy and speciation processes, between the annual S. pumila of the Old world and the
in particular about the reproductive isolation of these S. parviflora of the New World leads us to expect some
different chromosome races. For instances, samples possible relationships and crossing potential in a foreign
recognized as probably S. verticillata were in fact spon- tertiary gene pool (Wang et al. 1995b).
taneous hybrids between S. italica and S. verticillata,
confirmed at 3x ¼ 27 chromosomes, so that they can
serve as bridge between the ploidy levels (Poirier-
Hamon and Pernès 1986). Aneuploidy in the genus 15.3.2 Mating System
has been reported, but is considered insignificant.
GISH, a technique of painting chromosomes accord- All the Setaria discussed in this chapter are pur-
ing to genomic affinity, showed clear differentiation ported to be highly self-fertilized. The weedy foxtails
between the A genome set of chromosome of S. viridis are primarily a self-pollinated species (Pohl 1951;
and the B set of S. adhaerens. Metaphase of hand Mulligan and Findlay 1970). Wind pollination (ane-
crossed hybrids between these two species had one set mophily) is the mode in those rare circumstances of
of 9 chromosomes of each parent, which confirmed that outcrossing (Pohl 1951), with pollen probably moving
280 H. Darmency and J. Dekker

at most a few dozen meters (Wang et al. 1997). The resistant and susceptible S. faberi and found a single
panicle has many very short branches (fascicles), each codominant allele endowing the resistance.
bearing a number of branchlets and consisting of sev- Asexual reproduction is not a common mode of
eral spikelets and a cluster of setae (bristles). S. pumila reproduction in weedy foxtails. The only weedy fox-
panicle morphology differs from that of the secondary tail relying on vegetative asexual reproduction is
gene pool: only a single spikelet is found in each S. parviflora, a perennial species with short, branched,
S. pumila fascicle. A spikelet comprises two flowers; knotty rhizomes (Rominger 1962). Apomixy in
one is hermaphrodite, the other is sterile for S. viridis, S. viridis has been noted (Mulligan and Findlay
S. faberi and S. verticillata. In S. pumila, the lower 1970), and has also been suggested for S. pumila and
floret has no pistil but three well developed anthers, S. verticillata (Steel et al. 1983). Apomixis has also
which open 3–7 days later than the first floret been reported in other Setaria species within the
(Willweber-Kishimoto 1962). Although the anthers Setaria subgenus (Emery 1957; Chapman 1992).
are visible at flowering, and then feathery stigmates Emery (1957) reports both sexual (autogamy) and
also exert from the glumes, the fertilization principally apomictic seed production in S. leucopila, S. machros-
occurs within the flower. Stigmates maturity is tachya and S. texana, a highly unusual condition.
generally synchronous with anther dehiscence, which S. faberi and S. pumila tillers readily root in soil
results in a high probability of self-pollination. Some when cut, separated from the plant, and buried in
rare cases of protogyny, or in contrast, of protandry, moist soil, an important trait allowing weedy foxtails
can be observed according to the genotype and the to reestablish themselves after cultivation and mowing
environmental conditions, but in both cases the more (Santleman et al. 1963; Schreiber 1965).
proximate pollen belongs to the other spikelets of the
same panicle, thus leading again to self-pollination.
Enclosing one single panicle of S. viridis and S. viridis
spp. pycnocoma in a bag causes no more than 5% of 15.3.3 Interspecific Hybridization
empty spikelets, which could be due to the bag and the
greenhouse effects (Darmency and Pernès 1985). The 15.3.3.1 Hand Crossing
former experiments to report on quantitative estimates
were carried out for foxtail millet using red pigmenta- Beside spontaneous crosses reported or analyzed in
tion of the stem as a marker. When sowed in rows some instances, most of the direct knowledge on the
0.60 m apart, outcrossing was 0.59%, but it could be hybridizations relies on hand crossing experiments in
up to 2.26% in the case of mixed planting (Takahashi controlled conditions, as described in Sect. 15.7.1.
and Hoshino 1934). Thick bristled strains showed Hybridization between foxtail millet and all the other
lower percentage of natural crossing than the rough species of the group were successful, but to different
bristled, which could predict lower outcrossing in extent: it was rather easy with S. viridis (Kihara
wild Setaria because they are generally more bristly. and Kishimoto 1942; Li et al. 1945; Darmency and
Till-Bottraud et al. (1992) found 0.74% outcrossing Pernès 1985), but it was apparently less evident with
for S. viridis plants spaced every 0.25 m. Similarly, a S. adhaerens, S. faberi, and S. verticillata. In addition,
selfing rate exceeding 99% was reported in Jasieniuk further viable progeny was not always obtained,
et al. (1994) for S. viridis. Using a dominant herbicide- which, however, depends on the efforts made for the
resistance marker, Volenberg and Stoltenberg (2002) number of crosses carried out and the genotypes used.
found an outcrossing rate ranging from 0 to 2.4% for Benabdelmouna et al. (2001a) obtained an average of
S. faberi planted at 0.36 m interval. There was very one hybrid per emasculated spike of S. viridis when
little report of intraspecific crosses to study the genetic the pollen donor belongs to S. adhaerens. These inter-
nature of the traits of the wild Setaria. In two cases, specific hybrids were weak, grew slowly, and pro-
they concern herbicide resistance. Jasieniuk et al. duced very few flowers, most of them were sterile.
(1994) crossed trifluralin-resistant and susceptible The reciprocal cross gave no hybrid, but from a lesser
S. viridis plants, which showed that the resistance number of spike. Li et al. (1942), after many attempts,
was under the control of a recessive nuclear gene. obtained two seeds of the S. faberi  S. italica cross.
Volenberg and Stoltenberg (2002) crossed fluazifop These hybrids looked like S. faberi but they were
15 Setaria 281

almost sterile. Poirier-Hamon and Pernès (1986) it is likely that the contribution of one hybrid to the
obtained a few sterile hybrids between S. verticillata seed production of a field population would be about
and S. italica, but a colchicine treatment got a very 30 times less than for a green foxtail. Hybridization
variable progeny. Although attempted, no natural or studies showed that hybrids between green foxtail and
artificial crosses between S. viridis and S. pumila suc- foxtail millet had three times fewer tillers and spikes
ceeded. Willweber-Kishimoto (1962) successfully (Darmency et al. 1987a), and four times less spikelet
pollinated both S. italica and S. viridis with S. faberi fertility for the main spike (Darmency and Pernès 1985;
and hybrid seed was formed. Willweber-Kishimoto Wang et al. 2001), and 12 times less fertility for
(1962) successfully pollinated S. pumila (n ¼ 18) secondary tillers (Wang et al. 2001) than their green
with S. faberi (n ¼ 18), no hybrid seed were formed. foxtail parents. Similar results were found by Li et al.
(1945) and reported by Gu (1987). Hybrids using the
giant green foxtail, S. viridis ssp. pycnocoma, had less
15.3.3.2 Spontaneous Crosses spikelet sterility and were rather close to the cultivated
type, thus indicating a closer proximity to the domes-
In experimental plots, as much as 3% outcrossing was ticated species (Darmency and Pernès 1985). This
recorded in experiments where the two species were closer proximity was confirmed by lower variance of
grown together (De Wet et al. 1979). Lower frequen- the traits measured on the F2 compared to that of a cross
cies were generally found in most experiments, values with a typical S. viridis (Darmency et al. 1987b).
differing according to the cultivar, the ratio of target Some spontaneous hybrids, or descendants of
versus pollen donor, the respective location of target hybrids, were directly identified in the field in few
plants in the experimental design, and which species instances. Sterile hybrids between S. italica and
was the target (Darmency et al. 1987b, 1992). In S. verticillata were observed repeatedly in millet field
conditions that mimic a foxtail millet field, seeds in France (Poirier-Hamon and Pernès 1986). Fertile
produced by S. viridis growing between the rows hybrids between S. italica and S. viridis also occurred
contained 0.2% hybrids, while only 0.002% of in both directions in the same region, as determined by
the grains produced by the crop were hybrids chloroplast DNA analysis (Till-Bottraud et al. 1992).
(Till-Bottraud et al. 1992). This unequal balance is Stace (1975) found a naturally occurring S. viridis 
due to the taller habit of the crop that produces a lot S. verticillata hybrid called S.  ambigua Guss
of pollen grains directly above the wild plants, while (2n ¼ 36; Clayton 1980) (syn. S.  ambigua Guss
the pollen of the shorter S. viridis remains close to the emend. Koyama; S.  decipiens Schimp. (Osada
soil even with air turbulence. The rate of hybrids 1989)) throughout southern Europe. Finally, the occur-
decreases with the distance to the plot of foxtail millet rence of allotetraploids of AB genomes indicates that
used as pollen donor (Wang et al. 2001). Under com- spontaneous interspecific crosses were successful in a
mercial field conditions, the rate of hybrid produced remote past and have resulted in fertile progeny. In
by S. viridis is still lower: 0.039, 0.007 and 0.002% particular, the relatively homogeneous population
inside the field, 5 and 20 m away, respectively (Shi allozyme data suggest this polyploidization event in S.
et al. 2008). faberi was a relatively recent evolutionary event (Wang
Since both S. italica and its wild relatives are very et al. 1995b). A comprehensive picture of the relation-
variable in morphology, it is quite normal that ships among the Setaria species of the foxtail millet
botanists rarely described hybrid forms. For instance, complex has been presented in Darmency (2004).
shape and growth habit of hybrids between S. viridis
and S. italica correspond to the gross description of
S. viridis spp. pycnocoma that is often still considered 15.4 Phenotypic Variations
a genuine taxon (Darmency et al. 1987b; Darmency
2004). In the wild, such hybrids could suffer fitness
penalty due to a mix of antagonistic wild and domes-
15.4.1 Morphological Variants
ticated characters, lack of seed dormancy, needs of
more resources, etc. Although no direct estimate of S. viridis varies greatly in morphology and growth
the relative fitness of hybrids has ever been carried out, characteristics. A number of morphological variants
282 H. Darmency and J. Dekker

of S. viridis have been reported, including named (see dormancy below). Similarly, the growth kinetics
varieties: S. viridis var. viridis; S. viridis var. weinmanni parameters have been seldom taken into account in the
(R. & S.) Brand.; S. viridis var. ambigua; S.viridis description of the variability, but they can have major
var. major (Gaud.) Posp., giant green; S. viridis var. impact on the competitive and reproductive output of
robusta-alba Schreiber, robust white; S. viridis var. the variants. Thus, experiments in growth cabinet sug-
robusta-purpurea Schreiber, robust purple; S. viridis gest higher potential for competition of the giant type
var. pycnocoma (Steud.) Henrard & Nakai; and S. viridis spp. pycnocoma at the seedling and juvenile
S. viridis var. pachystachys (Franch & Savat.) Makino stage (Darmency et al. 1999).
& Nemoto (Hubbard 1915; Schreiber and Oliver 1971;
Kawano and Miyake 1983). The taxonomic validity of
formally recognized morphological varieties within
S. viridis varieties has been questioned because they
do not differ isozymatically from typical forms of
15.4.2 Physiological Variants
S. viridis (Wang et al. 1995a). It was observed that
these varieties show no genetic differentiation from The very rapid increase in frequency of herbicide
the common type, and are completely embedded in resistant variants in populations is the most apparent
common S. viridis, S. viridis var. viridis allozyme behavioral variation among Setaria phenotypes
principal component analysis (PCA) plots. Confusion observed in management situations in the last 25
with identification is highlighted by the observation years. Cases of resistance to dalapon (Santelmann
that presence or absence of hairs on upper leaf surface, and Meade 1961), dinitroanilines, photosystem II,
normally thought of as diagnostic for S. faberi, are ACCase and ALS inhibitors were reported in Canada,
found in both S. viridis (spp. pycnocoma, giant green) USA and Europe (see references in Heap 2009).
and S. faberi (green giant; Wang et al. 1995a, b). Besides mutations (see Sect. 15.9.3), the mechanisms
However, S. parviflora and S. pumila, natives to of resistance present in weedy foxtail populations
two different continents, were found to be so similar include exclusion and metabolism (e.g., Thornhill
morphologically that the 24 accessions studied by and Dekker 1993; Wang and Dekker 1995). They
Wang et al. (1995b) were initially identified as resulted in several marked differences with respect to
S. pumila. Allozyme data proved to be diagnostic the wild type, for instance chloroplast anatomy and
in the identification of these cryptically different functioning (Gasquez and Compoint 1981; Ricroch
species. Wide morphological and physiological varia- et al. 1987), seed germination (Reschly et al. 1996;
tions among accessions of yellow and S. faberi were Tranel and Dekker 2002), growth and reproduction
observed (Santelmann and Meade 1961). (Wang et al. 2010), although not in all the cases
Abnormal forked, divided or segmented panicles (Wiederholt and Stoltenberg 1996).
in S. viridis have been observed, although these traits Physiological variation in dormancy and germina-
apparently are not heritable. Reproductive develop- bility induced in seed during embryogenesis is one of
mental mutants have been observed in which spikelets the most important phenotypic traits leading to colo-
proliferate as green scales, named S. viridis, var. vivi- nizing ability and is discussed in detail below (see
para (Bertol.) Parl. (Dore and McNeill 1980). Seed Sect. 15.6.1.4 on seed dormancy below; Dekker et al.
(spikelet) morphology and shape vary widely among 1996).
individual foxtail species and genotypes. The degree Variation in drought tolerance among the foxtails
of rugosity of the lemma is a valuable taxonomic has been observed (Blackshaw et al. 1981; Manthey
diagnostic character. Rugosity varies from smooth and Nalawaja 1987, 1982; Taylorson 1986). Poten-
and shiny in foxtail millet, to finely ridged in S. viridis, tially salt tolerant genotypes of S. viridis, var. pachys-
to very coarse rugose seed in other foxtail species tachys have been observed along the seacoasts of
(Rominger 1962). This rugosity may play an important Japan (Kawano and Miyake 1983; Jack Dekker per-
role in interactions with the soil environment deter- sonal observation 1992, 2000, data not reported). Salt
mining their behavior in soil seed banks, especially at tolerance exists in other Setaria species (Chapman
the soil particle-seed interface affecting the uptake of 1992). These studies indicate that there may be an
water and gases into the foxtail seed placental pore intimate physiological and morphological relationship
15 Setaria 283

between drought tolerance, salt tolerance and seed culms per plant. They have small (5–20-cm long),
dormancy. This is most apparent in extreme habitats erect or slightly nodding panicles with short (1–3-cm
where foxtail millet remains an important crop and long) fascicle branches on which the spikelets are
cultivated cereal (e.g., Central Asia including Afghanistan, arranged in clusters (de Wet et al. 1979; Prasada Rao
India, sub-Saharan Africa). “Primitive” variants of et al. 1987). The setae are well developed. They differ
foxtail millet (e.g., race moharia) are grown in mar- from weedy variants by having spikelets that persist
ginal areas of Asia (e.g., Afghanistan) and are well after maturity, aiding harvest. Variants of the race
adapted to high elevation, semi-arid conditions on maxima are 45–100-cm tall, 1-8 (av. 2) unbranched
poor soil (de Wet et al. 1979). These variants yield a culms, and large, pendulous panicles (to 35-cm long,
crop in areas with as little as 125 mm of rainfall per 8-cm wide) with elongated secondary (fascicle)
annum. branches (2–6-cm long) forming lobed clusters (de
Wet et al. 1979; Prasada Rao et al. 1987). Race max-
ima is highly variable and totalizes the main part of
landrace accession in genetic resources bank. Variants
of race indica are intermediate to the other two races,
15.4.3 Geographic Variants 1-25 (av. 3) culms, but are unique due to panicles
6–30 cm long whose fascicle branches are loosely
Locally adapted populations of Setaria occur through- arranged along the main axis, and their occurrence in
out its range of distribution, yet almost nothing is India and Pakistan. In addition, one may add among
known of this type of adaptation or how this local the domesticated variant the giant green foxtail,
adaptation occurs. Wang et al. (1995a, b) observed S. viridis spp. pycnocoma because it is a side-product
north–south genotypic variation in North American of the domestication and believed to be a hybridi-
populations of S. viridis and S. parviflora. Differences zation product of foxtail millet and S. viridis (see
in S. pumila seed dormancy and germination require- Sect. 15.3.3.2). This variant is robust (1.5–2.5 m
ments were found among five S. pumila accessions height), with larger panicles (to 20 cm), compared to
from California, Iowa, Pennsylvania, Connecticut, other S. viridis variants. It also has larger, lobed pani-
and Massachusetts (Schoner et al. 1978; Norris and cle branch fascicles (to 4 cm).
Schoner 1980). A more complete presentation of
global geographic variation is presented in the section
on population genetic structure below (Sect. 15.5.2).

15.4.5 Plasticity of Weedy Setaria

One of the most striking observations of foxtail behav-

15.4.4 Domesticated Foxtail Millet
ior is the utilization of phenotypic heterogeneity
Variants within a single genotype or individual, rather than
genetic diversity among the individuals of a popula-
Hubbard (1915) found foxtail millet to be exceedingly tion, as a means of exploiting a locality (Scheiner
variable, and describes in detail three subspecies, five 1993; Wang et al. 1995a, b). This phenotypic hetero-
varieties, six subvarieties, and six formae; as well as geneity takes the form of phenotypic plasticity and
many different cultural names for the different var- somatic polymorphism in many of its most important
iants. “Primitive” races of foxtail millet resemble giant traits, especially during reproduction. For instance, the
S. viridis inflorescences, while more “highly evolved” number of fertile spikelet per fascicle within each
(domesticated) races possess extremely large panicles giant or green foxtail panicle is variable and more
(de Wet et al. 1979). A more recent classification of plastic in response to environmental conditions
foxtail millet recognizes two or three different mor- (Clark and Pohl 1996; Doust and Kellogg 2006). A
phological races based on plant and panicle structure large individual variability on a range of 1–20 was
(de Wet et al. 1979; Prasada Rao et al. 1987). Variants found for the size and reproductive output among
of race moharia are 25–100-cm tall, with 5-52 (av. 9) the progeny of a single green foxtail plant in a maize
284 H. Darmency and J. Dekker

field, mainly dependent on earliness of emergence S. adhaerens needs a warmer climate than S. viridis
(Darmency et al. 1999). Most of the phenotype space that in turn needs a warmer climate than S. verticillata.
of the foxtails has not been characterized. Only For instance, S. verticillata is found all over France,
the most obvious behaviors and morphologies have while S. viridis is recorded in the southern half part of
been described (e.g., plant size and habit, color mor- France, and S. adhaerens in the Mediterranean border
phology, abundance of bristles, herbicide resistance), only (Tela-Botanica 2009). In contrast, foxtail millet
and these are usually the characters that are most cultivars are adapted to a wide range of climatic con-
apparent in management situations. ditions, including in northern latitudes (North China,
Mongolia, Russia) where its wild ancestor S. viridis is
seldom found.

15.5 Biology and Habitat of the Wild

Setaria 15.5.2 Population Genetic Structure

15.5.1 Habitat and Distribution The pattern of genetic diversity within an individual
weedy Setaria sp. is characterized by unusually low
The wild Setaria of the foxtail millet complex are well intrapopulation genetic diversity, and unusually high
known weeds and colonize waste places (Dekker genetic diversity between populations, compared to
2003, 2004). The biology of S. viridis was summarized an “average” plant (Wang et al. 1995a, b). These
by Douglas et al. (1985), that of S. pumila and two patterns of population genetic structure appear to
S. verticillata by Steel et al. (1983), and that of typify introduced, self-pollinated weeds that are able
S. faberi by Nurse et al. (2009). Frequently more to rapidly adapt to local conditions after invasion
than one Setaria species coexist together in a locality; and colonization. Although relative genetic diversity
possibly allowing a more complete exploitation of within each of the several Setaria species is very low,
resources left available by human disturbance and differences between homogeneous populations are
management. Several weedy foxtail species often high, indicating a strong tendency for local adaptation
coexist in a single field (most commonly with S. vir- by a single genotype. Nearly all populations analyzed in
idis), each exploiting a slightly different “opportunity America consisted of a single multilocus genotype,
space” or niche. Their presence is not reported in while more diversity could be found within European
natural types of plant community. Since S. viridis and Chinese populations. Genetic bottlenecks asso-
may be the pre-agricultural wild colonizer that humans ciated with founder events may have strongly con-
first domesticated, it is likely that its habitat turned to tributed to that genetic structure. The founder effect
be completely cultivated or managed by human activ- has been observed in S. viridis to a certain degree.
ities. This probably applies for the other species of the S. viridis accessions from North America have reduced
secondary gene pool. In addition, gene flow within allelic richness compared to those of Eurasia. Genetic
the wild–crop–weed complex probably has changed drift probably has occurred in S. viridis, as indicated by
the original genetics of the present S. viridis, such the many fixed alleles in North American accessions.
that there are no longer any wild progenitors left in Multiple introductions of S. viridis, in the absence
the world capable to establish in other communities. of local adaptation, should have produced a random,
Discovering wild specimens participating stably or mosaic pattern of geographic distribution among North
repeatedly in natural communities could be invaluable American accessions. Instead, a strong intracontinental
to enlarge the environmental scale of habitat adapta- differentiation is observed in S. viridis populations, both
tion of the germplasm resources of foxtail millet. in Eurasia and North America (Jusuf and Pernès 1985;
Wild allies of foxtail millet in Eurasia are distri- Wang et al. 1995a). S. viridis populations in North
buted in subtropical to temperate areas, and they are America are genetically differentiated into northern
found under similar climates in other places of the and southern groups separated on either side of a line
world where they have been introduced by man. How- at about 43.5 N latitude. The northern type is less
ever, the species do not exactly match together: variable than southern type. This regional divergence
15 Setaria 285

suggests that natural selection has partitioned S. viridis North America, S. parviflora accessions were strongly
along a north–south gradient. These observations imply differentiated into southern and northern groups at
that the present patterning among S. viridis populations about the Kansas-Oklahoma border (37 N latitude);
in North America is the consequence of multiple intro- indicating greater genetic differentiation within
ductions into the New World followed by local adapta- North American populations than between North
tion and regional differentiation. American and Eurasian populations.
S. pumila populations are genetically clustered into S. faberi contains virtually no allozyme variation.
overlapping Asian, European, and North American Fifty of the fifty-one accessions surveyed by Wang
groups (Wang et al. 1995b). S. pumila populations et al. (1995b) were fixed for the same multilocus
from the native range (Eurasia) contain greater genetic genotype.
diversity and a higher number of unique alleles than
those from the introduced range (North America).
Within Eurasia, Asian populations have greater
genetic diversity than those from Europe, indicating 15.6 Role of Wild Setaria in Elucidation
that S. pumila originated in Asia, not Europe. These of Origin and Evolution of Foxtail
observations indicate there have been numerous intro- Millet
ductions of S. pumila from Eurasia to North America,
the majority from Asia. This pattern may also explain
15.6.1 Analysis of the Domesticated
the enigma of the origins of S. pumila and S. parvi-
flora. The pattern of S. pumila genetic variability in Traits
North American was unexpected: nearly the entire
diversity of this species appears to be encompassed 15.6.1.1 Seed Shedding
by accessions from Iowa, whereas populations col-
lected from other North American locations were On the basis of visual estimates of two classes (shed-
nearly monomorphic for the same multilocus genotype ding or not), Li et al. (1945) showed that seed shedding
(Wang et al. 1995b). In this respect, it is significant or is controlled by two pairs of complementary genes,
coincidental that this pattern was repeated in the diver- with shedding being dominant, and a 9:7 ratio in F2
sity data for S. viridis, also a native of Eurasia (Wang populations. However, the frequencies obtained in the
et al. 1995a). Iowa possesses a surprising Setaria F3 were significantly different from the expected ones.
genetic diversity: all five weedy Setaria species are Other studies showed different ratios, from 14:2 to
present. Typically two or more Setaria species occur 11:5, and around 35% of non-shedding F2 segregating
in the same field at the same time. Iowa is the center in F3 (Darmency and Pernès 1987). Finally, a full
of the north–south agro-ecological gradient in North quantitative approach estimating seed shedding as
America, perhaps leading to greater environmental the proportion of seed with attached glumes allowed
heterogeneity. to calculate a minimum number of four genes involved
Despite originating on different continents, the (Darmency et al. 1987a), probably two major genes as
genetic diversity patterns for S. parviflora parallel observed above and two minor genes. Several quanti-
those for S. pumila and S. viridis: greater genetic tative trait loci (QTLs) related to seed shattering, stem
diversity occurs in accessions from the New World height, node number and length of panicle were iden-
compared to those from the introduced range (Eurasia) tified using an F2 population (Kun et al. 2008).
(Wang et al. 1995b). This most likely reflects genetic
bottlenecks associated with sampling a limited number
of founding propagules and the history of multiple 15.6.1.2 Inflorescence Architecture
introductions from the Americas to Eurasia. The pop-
ulation genetic structure of S. parviflora consists of The morphology of Setaria inflorescences consists of
three nearly distinct clusters, groups from Eurasia, organs and tissues organized in hierarchical nested
northern United States, and southern United States. structural sets: tiller culm, panicle, fascicle, spikelet
Accessions from Eurasia and North America are and floret (Dekker et al. 1996). The domesticated
approximately equally diverse genetically. Within species has more primary fascicle (branch) number
286 H. Darmency and J. Dekker

and more spikelet number than its wild ancestor, and seedling emergence; Dekker 1999). There is evi-
which negatively correlates with primary branch den- dence that soil water, temperature and oxygen play a
sity. No classical genetic interpretation has been unifying role in regulating both the global bio-geo-
given. Eleven QTLs of five linkage groups were sig- graphic distribution of weedy Setaria spp., as well as
nificantly associated to these traits (Doust et al. 2005). the responses of individual seeds in a soil microsite
Several candidate genes identified to have similar (Dekker 2000; Dekker et al. 2001; Dekker and
action in maize and rice were proposed to control Hargrove 2002). The morphology of weedy Setaria
these wild versus crop differences. spp. seeds provides clues about which environmental
factors limit germination and maintain dormancy.
The weedy Setaria spp. seed interior (embryo, endo-
sperm, and aleurone layer) is enveloped by several
15.6.1.3 Tillering
layers controlling its behavior, notably the hull
(lemma, palea) and the caryopsis coat (Rost 1973;
The profuseness of tillering due to the wild parent of the
Dekker et al. 1996; Dekker and Luschei 2009). The
cross between S. viridis and S. italica makes it quite
caryopsis coat is water- and gas-tight, and continuous
difficult to classify the progeny. On a visual scale, low
except at the narrow placental pore opening on the
tillering seems to be somewhat recessive and high til-
basal end of the seed. The mature weedy Setaria spp.
lering quantitatively dominant. Earlier works suggested
seed is capable of freely imbibing water and dis-
that only one co-dominant gene was involved, meaning
solved gases, but entry is restricted and regulated by
that 3/4th of the F2 progeny have various degrees of
the placental pore tissues, notably membrane control
profuseness of tillering, and 1/4th have few tillers
by the transfer aleurone cell layer (Rost and Lersten
resembling the crop parent (Li et al. 1945). In a quanti-
1970). This morphology strongly suggests that seed
tative analysis of the parents, of the F1, F2 and F3
germination is restricted by water availability in
populations, a broad heritability estimate of h2 ¼ 0.65
the soil, and by the amount of oxygen dissolved in
was calculated (Darmency et al. 1987a). A QTL
water reaching the inside of the seed interior to fuel
approach allowed the identification of four QTL mar-
metabolism. Thus, the signal stimulating Setaria
kers associated with tillering difference between
behavior in an individual seed is oxygen mass per
S. italica and S. viridis, and four associated with axillary
water volume of symplast (caryopsis) per time (e.g.,
branching: candidate genes could be researched at
hour, day): mass O2 volume H2O1 time1 seed1
corresponding places on the gene map of other cereals
(oxy-hydro time; Dekker et al. 2001; Dekker and
(Doust et al. 2004). A further research increased the
Atchison 2003).
number of QTLs associated to the phenotypic distribu-
tion of the traits in F3 families, but some of these QTLs
were also associated to phenotypic variation according
to the environment (Doust and Kellogg 2006).
15.6.1.5 Grain Size

There is no more than one single grain or seed per

15.6.1.4 Germination spikelet. This seed is indurate, usually transversely
wrinkled, lemma and palea (hull) of similar marking
No inheritance study was directly dedicated to ger- and texture, which tightly encloses the caryopsis
mination and seed dormancy, probably because seed within at maturity (see dormancy regulation below
germination in weedy Setaria is controlled by many for caryopsis morphology). Spikelet length for Setaria
gene complexes involved in the formation and varia- spp. varies both within and between species (Rominger
tion in seed structures. The wide geographic range of 1962). S. viridis seeds are 1.8–2.4-mm long (var. giant
adaptation, heterogeneity in dormancy phenotypes, S. viridis, 1.8–2.2 mm; var. robust purple and robust
and genotypic diversity raise the question of what white, 2.0–2.4 mm), 1.0–1.3-mm wide, and weigh
mechanisms in weedy Setaria spp. seeds drive seed 0.71–0.87 mg (var. giant S. viridis, 1.08 mg; var.
behaviors (e.g., induction of dormancy, after-ripen- pachystachys 0.87 mg; var. robust purple 1.27 mg;
ing, germination, induction of summer dormancy, var. robust white 1.39 mg). S. faberi seeds are
15 Setaria 287

2.5–3.0 mm in length, and weigh between 2.00 and 15.6.2 Application of Morphotaxonomy,
2.169 mg (Schreiber and Oliver 1971; Kawano and Chemotaxonomy, Biochemical
Miyake 1983). S. pumila seeds are 2.5–3.4-mm long,
and Molecular Markers
1.5–2.2-mm wide, 1.0–1.5-mm thick, and weigh
between 2.7 and 4.5 mg (Stevens 1932; Povilaitis
1956; Schreiber and Oliver 1971; Kawano and Miyake The evolution and domestication history of S. italica
1983; Steel et al. 1983). S. verticillata spikelets are have been studied using different characters such as
1.8–2.3-mm long, 1.0–1.5-mm wide, 0.6–1.3-mm phenol color reaction, esterase isozymes, hybrid
thick, and weigh between 1.1 and 1.3 mg (Steel et al. pollen sterility, morphological data, storage protein,
1983). Foxtail millet of maxima race are 2.8–3.5 mg. RFLP, RAPD and amplified fragment length poly-
Quantitative analysis showed that grain size was morphism (AFLP) molecular markers, and transpo-
highly heritable (broad heritability h2 ¼ 0.84), but son DNA sequence. A few to several S. viridis were
it was impossible to propose any genetic system generally included in the studies that comprised large
(Darmency et al. 1987b). geographical samples of landraces and cultivars. It
has been suggested that a unique domestication
center in China was followed by local adaptation
(Nakayama et al. 1999; Le Thierry d’Ennequin
15.6.1.6 Grain Quality et al. 2000). Alternately, up to three domestication
centers have been suggested in China, Europe and a
Two grain attributes appear to be tightly associated to geographical area ranging from Afghanistan to Leba-
the wild Setaria status and are rarely found among non (Kawase and Sakamoto 1982, 1987; Jusuf and
the crop cultivars, probably because they confer bitter Pernès 1985; Darmency and Pernès 1987; Li et al.
taste to the flour: the presence of polyphenoloxidases 1995, 1998; Fukunaga et al. 1997; Schontz and
in the seed coat, and the presence of phenolic pig- Rether 1999; Hirano et al. 2008). A different picture
ments in the pericarp. The phenol color reaction is provided by the study of the heterogeneity of
is controlled by a single gene, with the positive copia-like retrotransposons. Their sequencing from
(i.e., the wild) phenotype being dominant (Kawase five species revealed high sequence heterogeneity
and Sakamoto 1982; Darmency and Pernès 1987; even between clones from a single species (Benab-
Till-Bottraud and Brabant 1990). As for pericarp delmouna and Darmency 2003). Since the degree of
pigmentation, F1 hybrids of interspecific crosses sequence divergence is linked generally to phyloge-
have the crop phenotype (unpigmented pericarp). netic relationships, it could have been expected that
Various F2 segregation ratios were observed in bottlenecks of diversity occurring along with domes-
different studies, but all the authors propose a single tication of the cultivated S. italica would have
gene for pigmentation with its action augmented, resulted in a much lower genetic variability in the
modified, or suppressed by the action of a second cultivated crop than in its wild ancestor S. viridis and
gene (Li et al. 1945; Darmency and Pernès 1987; allied species. However, the sequence divergence
Till-Bottraud and Brabant 1990). was found to be as high between the two cultivars
of S. italica as among the four wild Setaria species. It
is possible that the high level of divergence revealed
in the cultivated species resulted from numerous
15.6.1.7 Miscellaneous introgressions from related wild species. Other stud-
ies suggested that genetic differentiation occurred
Other information not directly associated to the after domestication and that the lack of structure of
domestication process was also obtained from the the wild gene pool might be a consequence of gene
interspecific crosses. For instance, the stem base pig- flow between the crop and its wild relatives
mentation (one dominant gene) and molecular markers (Nakayama et al. 1999; Le Thierry d’Ennequin
including isozymes and SSR and also QTLs were used et al. 2000). Alternatively, multiple independent
to identify linkage groups (Darmency and Pernès domestication events, from different S. viridis acces-
1987; Kun et al. 2008). sions, could have occurred, thus generating copia
288 H. Darmency and J. Dekker

sequence variability at the very origin of the crop. 15.7.2 Genes and Genome Sequencing
The variability and differences among ribosomal
DNA gene sequences of foxtail millet landraces sup-
A few DNA sequences are already available. They
port such a multiple, independent, domestication
correspond to the few genomics resources developed
(Fukunaga et al. 2006).
in studies including wild Setaria and are available at
EMBL and DDBJ GenBanks. They include ribosomal
DNA genes (Benabdelmouna et al. 2001b), ACCase
gene (Délye et al. 2002), copia-like retrotransposons
15.7 Role of Wild Setaria in Molecular
(Benabdelmouna and Darmency 2003), a and b-tubulin
Genetic Studies genes (Délye et al. 2004), and ribosomal intergenic
spacer subrepeats (Fukunaga et al. 2006).
15.7.1 Development of Cytogenetic The genome of S. italica is being sequenced.
Stocks and Genetic Maps Together with a crop cultivar, an accession of S. viridis
will be sequenced too and compared to that of the
domesticated plant (Doust et al. 2009). It will facilitate
In parallel to constructing a RFLP-based map from
comparative genomic analysis, assignment of domes-
an intraspecific S. italica F2, Wang et al. (1998) con-
tication traits along the chromosome, thus allowing
structed another map from an interspecific S. italica 
better knowledge of the genome organization and con-
S. viridis F2, in order to reveal wider polymorphism.
stituting a helpful tool to manage introgression when
As expected, the second map showed much more
desirable genes will have to be introgressed from wild
polymorphism (75% relative to 44% polymorphic
relatives.
loci). The map based on the interspecific cross was
subsequently used as a reference for comparative stud-
ies, in particular to investigate the synteny with rice
and provide researchers with large pools of known 15.8 Methods for Hybridization
markers for gene identification (Devos et al. 1998). and Introgression of Traits
The map comprises nine linkage groups, which were from Wild Setaria
assigned to the corresponding nine chromosomes
using trisomic lines (Wang et al. 1999). One of the
first outcome of this genetic approach was to allow 15.8.1 Hybridization
QTL analyses to determine regions containing candi-
dates genes in maize or rice, such as tb1 (teosinte Anther emasculation by hand is possible, but it is a
branched 1) and ba1 (barren stalk 1) for the control careful, boring and time-consuming method. In many
of tillering and branching (Doust and Kellogg 2006). cases, emasculation was performed by dipping the
In addition, because of the plant plasticity of the wild entire inflorescence just starting to bloom in hot
parent according to the growth conditions, QTL water at 42 C for 20 min (Li et al. 1935). The inflores-
regions variably associated to a given trait could cence was then enclosed in a glassine bag along with
be both involved in its genetic control and its pheno- an inflorescence of the male parent, and the bag was
type expression, which potentially provides new shaken several times a day to allow the pollen reaching
insight in the genome organization. This map from the the castrated flowers. A second round of the emascu-
S. italica  S. viridis F2 is now enriched with simple lation technique 1 or 2 days later can allow up to 60%
sequence repeat (SSR) markers (Jia et al. 2009). A of cross-hybridization, but the hybrid status of the
green foxtail bacterial artificial chromosome (BAC) released seeds must be confirmed because of abortion
library is under construction (X Diao personal com- or destruction of all the pollen grains of the inflores-
munication). Trisomic lines and other aneuploids to cence used as the female cannot be warranted. Differ-
identify linkage groups were obtained by crossing ent morphological or molecular markers are necessary
colchicine-induced autotetraploid S. italica crossed to confirm the hybridity, for instance the dominant red
to normal line (Wang et al. 1999); wild tetraploid was collar pigmentation easily observed at the two-leaf
never successfully used. stage (Darmency and Pernès 1985), or isozymes
15 Setaria 289

markers (Wang and Darmency 1996), or panicle char- 15.9.1 Male Sterility
acteristics (size, anther color, bristle color; Li et al.
1945). When male sterile lines of foxtail millet were
A cytoplasmic male sterile (CMS) line was obtained in
made available, hybridization became very simple by
the progeny of backcrossed plants after crossing a
enclosing one ear of each parent in the same paper bag
S. verticillata with an autotetraploid S. italica (Zhu et al.
(Wang and Darmency 1997), but no such male sterile
1991). However, the research of restorer genes among
plants were found among the wild relatives.
the wild collections to setup restorer lines has not been
documented yet. Nuclear encoded male sterile genes are
15.8.2 Introgression now used in millet breeding, but they apparently belong
to crosses between geographically distant landrace acces-
sions (China and Australia) whose pedigree could have,
Mass selection among the huge variability created in
perhaps, included wild relatives.
the F2 progeny of interspecific hybrids is a first pos-
sibility to detect favorable genetic recombination.
Multivariate analyses showed that the differing char-
acters between the crop and the wild millet seem to be 15.9.2 Tetraploidy
distributed along three independent groups (1) the
plant tillering, (2) the seed size and shedding, and (3)
the size of the vegetative and reproductive organs. Numerous crop species are polyploid so that attempts
Other traits, such as flowering time, behave indepen- were performed to obtain tetraploid cultivars exhib-
dently (Darmency et al. 1987a, b). Since important iting higher productivity. However, autotetraploids
characters to differentiate the wild and the cultivated resulted in poor values compared to diploids, and we
millet seems to be grouped, a rapid return to the are not aware of any tetraploid commercial cultivar.
cultivated type can be achieved in the breeding plans In a few instances, a wild species was involved in a
while new independent traits are identified and used. breeding plan in order to stabilize the valuable
This fact has an important outcome on crop breeding characteristics of the tetraploid (small plant and high
as it opens the way for a larger use of wild plants in thousand-grain weight) and to counterbalance the defi-
improving the agronomic quality of foxtail millet. ciencies (in particular a reduced fertility), or simply to
High genetic variation remained within F3 families explore the potential outcome of interspecific breeding
for most characters, which may provide further gain or serve as a bridge to use wild related polyploids. For
in further selection (Zangre and Darmency 1993). instance, an autotetraploid S. italica was created to
However, if autogamy allows easy selection of F2 cross with S. yunnanensis, a tetraploid wild relative.
and F3, backcrossing is a more difficult process. Back- The resulting hybrids were sterile, but some progeny
crossing to the same parent can be done easily in the at 2n ¼ 36 could be obtained through in vitro culture
case of a male sterile line (Wang et al. 2009a). Other- of immature inflorescences (Zhou et al. 1988). Simi-
wise, accurate hand crossing involving very few flow- larly, crosses have been done between an autotetra-
ers and availability of differential parental markers are ploid S. italica line and S. faberi, a wild tetraploid
necessary. Alternatively, screening suitable plants species that already presents large seeds (Luo et al.
among selfed F2s retaining a given marker and then 1993). Alternatively, selection of autotetraploid could
backcrossing them to the cultivar can allow a rapid be performed by colchicine induction after the produc-
return to the cultivated type within only two backcross tion of an interspecific hybrid between S. viridis and
generations (Naciri et al. 1992). S. italica (Ahanchede et al. 2004).

15.9 Role in Crop Improvement

15.9.3 Herbicide Resistance
Very little achievements of foxtail millet breeding using
wild relatives have been reported. They include cases of Research for herbicide resistance (or tolerance) is a
male sterility, tetraploidy and herbicide resistance. very recent goal in plant breeding (Darmency 2003).
290 H. Darmency and J. Dekker

In the case of foxtail millet, weed control remains one 15.9.3.2 Trifluralin Resistance
of the main problems to be solved as no selective
herbicide has been developed for it because it does Several populations of green foxtail evolved resistance
not represent a market large enough to justify the to dinitroanilines, a group of chemicals that inhibits
research expenses. Thus, no satisfactory weed control cell division. The resistant plants were seven times
herbicide program exists, while hand control is more more resistant to trifluralin and caused trouble in
and more expensive and less and less attractive for wheat fields (Beckie and Morrison 1993). The
farmers. Herbicide resistant millet cultivars were resistance was inherited as a recessive nuclear gene
designed to overcome this situation. (Jasieniuk et al. 1994), but a strong bias was observed
in the progeny of the interspecific hybrids, with on
average 14% homozygous resistant instead of 25%
15.9.3.1 Triazine Resistance (Wang et al. 1996b). The discovery of the mutations
responsible for the resistance, a Thr239–Ile and a
Triazine is a herbicide class that inhibits the electron Leu136–Phe mutation of the gene encoding a2-tubulin
transfer at the photosystem II level. In Europe, atrazine (Délye et al. 2004), allowed to check that only one
at low dose was often used for weed control in foxtail structural gene was involved. The bias was inherited in
millet, but its effect on weeds does not last enough the next generations, but after further crosses and
to maintain an efficient protection during the whole generations, a normal inheritance pattern could be
growth season. A few populations of green foxtail observed in some lines, which suggested the possibil-
resistant to 1,000 times the field doses of atrazine ity of a linkage between the a2-tubulin and a modifier
were found in France (Gasquez and Compoint 1981), gene (Tian et al. 2006). It was inferred from the
and subsequently in other Setaria species and other alignment of rice and millet genetic maps that the
countries (Heap 2009). Since no resistant cultivar was a2-tubulin gene belongs to the linkage group IX that
observed among a world collection of foxtail millet, showed distorted segregation also in an RFLP study
the resistant green foxtail was used in order to intro- (Wang et al. 1998). Resistant lines were further
gress the resistance trait. Only hybrids from the green selected, but because the intensity of the resistance is
foxtail used as the female parent were confirmed to be not high enough to warrant good weed control in the
resistant, and all their F2 progeny by self-pollination field, registered cultivars displaying this property
were resistant, which indicated maternal inheritance could hardly have been developed. However, it is a
(Darmency and Pernès 1985). Further studies showed helpful tool to maintain pure male sterile lines used
that the Ser264–Gly mutation of the chloroplast psbA to produce hybrid seeds as it allows discriminating
gene was the molecular basis of atrazine resistance in homozygous recessive resistant from heterozygous
Setaria (Tian and Darmency 2006). This mutation at susceptible descendants developed from uncontrolled
the photosystem II, however, was shown to incur a pollination (Wang et al. 1996a). An allele-specific
high fitness cost. When resistant BC2 were compared polymerase chain reaction (PCR) routine protocol
to the original susceptible cultivar, both had similar was set up to allow quick discrimination of the differ-
developmental characteristics, but the rate of photo- ent alleles (Délye et al. 2005).
synthesis (CO2 fixation) was lower for the resistant at
27 C while remaining not different than that of the
susceptible plants at 17 C (Ricroch et al. 1987). Chlo- 15.9.3.3 Sethoxydim Resistance
rophyll fluorescence analysis showed that electron
transfer in photosystem II was slower in the resistant Several populations of green foxtail evolved resistance
plants, thus representing a limiting factor at tempera- to sethoxydim in Canada (Heap and Morrison 1996), a
tures allowing higher biological activity. When both herbicide of the cyclohexanedione group that blocks
plant materials were grown in the field, this resulted in fatty acid biosynthesis in the Gramineae. Crosses were
a 22% grain yield reduction for the resistant plants carried out with fertile and male sterile millet lines.
(Darmency and Pernès 1989). Nevertheless, high Results in F1, F2 and BC1 showed that the resistant
yield lines could be derived from these crosses (Ji progeny was 700 times more resistant than the suscep-
et al. 2006; Shi et al. 2008). tible cultivars and that a single, completely dominant,
15 Setaria 291

nuclear gene controlled the resistance (Wang and of uses of foxtail millet (Kimata et al. 1998). This
Darmency 1997). Materials derived from these crosses diversity of use, the need to get a new taste or a new
were used in a breeding program to release a series of color for food, the increasing importance of plants as
resistant lines, of which some (e.g., variety SR3522) sources of therapeutic compounds and dietary supple-
were registered as elite cultivar at the Seed Supervis- ments, could encourage searching for new quality
ing Bureau of Zhangjiakou in China (2005). The resis- attributes among wild relatives. Because the native
tance was demonstrated to be due to a point mutation gene pool for Setaria is very large its potential
in the carboxyltransferase domain of the nuclear gene is vast, especially as a crop in poor, dry and salty
encoding plastidic ACCase isoform, a Ile1781–Leu environments.
residue substitution (Délye et al. 2002). It was not
clear whether the resistant allele has fitness cost
because it was possibly linked to genes boosting juve-
nile growth, which resulted in being advantageous in 15.10 Conclusion
stressing field conditions (Wang et al. 2009a). Close
AFLP markers are now available to facilitate selection The Setaria (foxtail) wild–crop–weed species-group is
(Niu et al. 2002). one of the most successful terrestrial plant complexes
on earth. The genotypic and phenotypic biodiversity of
this species-group has conferred on it the ability to
15.9.3.4 Other Sources of Resistance invade, colonize, adapt and endure in a wide range of
disturbed habitats in temperate, tropical, and subtropi-
Further sources of differential selectivity of herbicide cal regions. Over 100 Setaria species were distributed
were demonstrated to occur in various wild Setaria throughout the world before the advent of agriculture.
for dalapon and butylate (Santelmann and Meade With crop domestication and land cultivation about
1961; Oliver and Schreiber 1971), and for PSII inhi- 10,000 years ago, wild, pre-agricultural, colonizing
bitors (Thornhill and Dekker 1993; Wang and Dekker foxtails could adapt and thrive in the newly created
1995). Perhaps, other mutant alleles, expressing dif- agro-ecosystems. The success of the foxtails is due to
ferent pattern of cross-resistance, could be found the intentional and unintentional consequences of
in the giant foxtail resistant to ACCase inhibitors weed management for over 10,000 years. Therefore,
(Stoltenberg and Wiederholt 1995; Volenberg and the future of the foxtail complex is to continuously
Stoltenberg 2002). High resistance against imidazoli- adapt to the management systems of human-imposed
none and sulfonylurea herbicides, two groups of mole- selection forces, and also to changing climate and soil
cules that block the biosynthesis of branched-chain conditions. Evolution of a complex seed dormancy
amino acids, was found in several populations of system has given the foxtails the ability to precisely
green, yellow and giant foxtail (Volenberg et al. time seedling emergence on time scales from hours to
2001; Heap 2009) in Canada and USA. In the case decades, providing them with the ability for enduring
of S. viridis, the resistance was demonstrated to be occupation of a locality. Foxtail soil seed banks are the
due to several mutations at the acetolactate synthase source of all future annual weed infestations in a
genes: Ser653–Thr, Ser653–Asn, Ser653–Ile, and locality, spreading the risk of continued development
Gly654–Asp (Laplante et al. 2009). Crosses between over time, a bet-hedging strategy balancing mortality
male sterile millet and plants of one of these IMI- against future fitness. Phenotypic plasticity in growth
resistant populations were undertaken in China and development has allowed the foxtails to optimize
(T. Wang personal communication). the match between the immediate resources available
and vegetative size and reproductive output, avoiding
pre-mature mortality of the individual or its parts. The
foxtails are endowed with the ability to tolerate high
15.9.4 Other Desirable Agricultural Traits concentrations of salt in the soil, low soil water, till-
age, and extremes in temperature. Because of their
Beside the use as a cereal for food, grain to feed ability to resist herbicides and adapt to most advanced
poultry and spring forage for cattle, there are varieties and powerful farming practices, they cannot be
292 H. Darmency and J. Dekker

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Chapter 16
Zoysia

Shin-ichi Tsuruta, Makoto Kobayashi, and Masumi Ebina

16.1 Basic Botany of the Species Zoysia. Because the five Japanese species have the
and Conservation Initiatives same chromosome number (2n ¼ 4x ¼ 40) and there
is no chromosomal barrier of interspecific crossing,
intermediate types are found along species’ borders
The members of the genus Zoysia Willd. (family in their natural habitats in Japan and have allotetra-
Poaceae, subfamily Chloridoideae, tribe Zoysieae) are ploidy (Forbs 1952; Yaneshita et al. 1999). Pistils are
mat-forming perennial grasses, consisting of about ten mature for 7–10 days prior to stamens, thus alloga-
species, which are indigenous to the Pacific Rim mous crossing is considered to be the preferred mode
(Clayton and Renvoize 1986; Engelke and Anderson of reproduction. The genus is self-compatible, how-
2003). The native distribution of the genus extends ever, and has very low canopy height and compara-
from New Zealand to the Hokkaido Island of Japan, tively heavy pollen, so the ratio of allogamous
including Polynesia and Southeast Asian coastal crossing in natural habitats is open to question.
countries (Engelke and Anderson 2003). Two of the Z. japonica is distributed in Japan, Korea, and
species, Z. japonica and Z. machrostachya, are scat- eastern China. The species is perennial and forms
tered in the some parts of Hokkaido (Ishida 1990; large mats in sunny fields or is cultivated as lawn,
Osada 1993). Hokkaido has a subarctic climate, and with long creeping rhizomes. Culms are 5–15-cm
the genus Zoysia is one of the C4 grasses most well tall. Leaf blades are 3–10-cm long and 2–5 (or
adapted to cold climatic regions (Hatch and White 6) mm wide, flat, with no ligule, but with long hairs
2004). Therefore, the genus Zoysia is one of the most at the mouths of sheaths. Spikelets are obliquely ovate,
salt and cold tolerant C4 grass species in the family about 3-mm long (two to three times that of the
Poaceae. breadth), and one-flowered; they consist of the upper
Three commercially important species are recog- glume and lemma and lack the lower glume and palea.
nized in the genus Zoysia and referred to as zoysia The upper glume is coriaceous, smooth, glossy, and
grass, Z. japonica Steud., Z. matrella (L.) Merrill, and keeled on the back, with five faint, barely visible
Z. tenuifolia Will. ex Trin. At least five species of the nerves; the lemma is papery, slightly shorter than the
genus, including the three commercially important glume, one-nerved, and sharply keeled. Anthers are
species, are native to the Japanese islands (Fig. 16.1), 1.5-mm long (Osada 1993).
and they exhibit morphological and ecological differ- Along the southern ends of Tanegashima and Yaku-
ences (Ishida 1990). Therefore, Japan is considered to shima Islands in Japan, the biogeographical line of
be one of the origins of the diversity of the genus Watashe is recognized at a latitude 30 N. Z. japonica
is found north of this latitude. Also, Blakiston’s line is
M. Ebina (*) recognized between Honshu and Hokkaido Islands at
National Institute of Livestock and Grassland Science, Forage latitude 40 N (Tamate et al. 1998). Because the distri-
Plant Breeding and Biotechnology, 768 Sembonmatsu, bution of Z. japonica on Hokkaido is limited and
Nasushiobara, Tochigi 329-2793, Japan scattered, the northern edge of the species’ original
e-mail: [email protected]
Shin-ichi Tsuruta and Makoto Kobayashi contributed equally natural habitat is considered to be Honshu (Ishida
and should be viewed as first authors. 1990). Z. japonica adapts to low-input sustainable

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 297
DOI 10.1007/978-3-642-14255-0_16, # Springer-Verlag Berlin Heidelberg 2011
298 S. Tsuruta et al.

Fig. 16.1 The five species of Zoysia indigenous to Japan: Z. japonica, Z. matrella, Z. tenuifolia, Z. macrostachya, and Z. sinica

grazing fields and is commonly used in the borders of paleas are usually absent (Osada 1993; Engelke and
paddy fields (Yamamoto et al. 1988; Otani et al. 1999; Anderson 2003).
Ogura et al. 2001). The other four species, native to Z. tenuifolia is a low-growing rhizomatous peren-
Japan, have strong salt tolerance, and only Z. japonica nial found in subtropical and tropical regions and is
has a weak tolerance to salt (Oishi and Ebina 2005). widely distributed in Japan (southern Kyushu and
Z. japonica is distributed naturally in mountainous Okinawa Islands), Taiwan, southern China, Indonesia,
areas, along riversides, and in coastal area (Otani Micronesia, New Guinea, Madagascar, and India. It is
et al. 1999). It has a moderate to weak shade tolerance adapted to coastal zones buffeted by high winds and
and subsists in soils ranging from infertile sands to clays waves that deliver substantial amounts of salt. Leaf
(Ishida 1990). Growth is better in soils that are weakly blades are completely involute and acerate. Racemes
alkaline, but this species tolerates acidic soils as well have fewer than 12 spikelets (Osada 1993; Engelke
(Ishida 1990). The hard seed easily germinates in the and Anderson 2003).
manure of ruminant animals (Takahashi et al. 1995). Z. macrostachya Franch. et Savat. is a creeping
Z. matrella is morphologically intermediate perennial that grows only on sandy soil along the
between Z. japonica and Z. tenuifolia and is distri- seashore. Culms are tufted, erect on the nodes of
buted in the southeastern part of Honshu Island, creeping rhizomes, and 15–40-cm tall. Leaf blades
southern region of South Korea, and China. It is a are 3–8-cm long and 2–5-mm wide, involute when
low-growing, rhizomatous perennial with culms up dry, and spiny at the apex. Racemes are dense and
to 5-cm tall. Leaf blades are glabrous, up to 7-cm spike-like, 3–4-cm long, and 5–7-mm wide. The lower
long and <3-mm wide, and involute. Racemes are half of each raceme is enclosed by the uppermost leaf
1–4-cm long and have 10 to 40 spikelets. The spikelets sheath. Spikelets are 6–8-mm long and 2–4-mm wide
are 2.1–3.2-mm long, each with one floret that is (Osada 1993). The rhizome is too prostrate to create a
smaller than but quite similar to that of Z. japonica. mat. Therefore, no commercial use has been reported.
First glumes are usually absent. Second glumes The species is distributed widely from the Ryukyus to
have indurate bases, and basal margins are connate Honshu, Hokkaido, and southern region of South
and awnless. Lemmas are 1.6–2.5-mm long, and the Korea, but the natural habitats are very narrow strips
16 Zoysia 299

along the seashore. Natural hybrids with Z. japonica was selected for its ability to form a dense turf and the
are found. Z. macrostachya has a unique trait of rapid establishment of turf and grazing fields.
mature seeds that can be peeled easily. Several
attempts have been made to introduce this trait to
16.2 Salt Tolerance Mechanisms
construct a new cultivar of seed-propagating zoysia
grass. This trait appears to be controlled by very com-
and Phylogenetic Relationships
plex quantitative trait loci (QTL), however, no signifi-
cant segregation occurs in F2 progeny. Therefore, In their natural habitats, zoysia grasses (except
breeding efforts for constructing seed-propagating Z. japonica) have strong salt tolerance. Because they
zoysia grass are still underway. have a low canopy but moderate to weak shade tole-
Zoysia grass collection and release in the US were rance, in the humid Pacific Rim zoysia grasses prefer
described in detail by Engelke and Anderson (2003). costal habitats, where no other plant can survive. The
The first organized collection expedition supported salt tolerance among the coastal species of zoysia grass
by the US Department of Agriculture (USDA) was is associated with shoot saline ion exclusion via leaf
undertaken in 1982 by Murray and Engelke. From salt glands and the overall density of leaf salt glands
the Pacific Rim countries of Japan, Korea, Taiwan, (Marcum et al. 1998). In addition to these morphologi-
and the Philippines, approximately 700 accessions cal characteristics for salt tolerance, Oishi and Ebina
from five of Zoysia species were collected. The origi- (2005) reported biochemical salt protection from the
nal collection is maintained as vegetative propagules osmotic balancers glycine betaine and polyamine. The
in greenhouses at Texas A&M University (Engelke gene coding for the enzyme involved in the synthesis of
and Anderson 2003). The evaluation of the original glycine betaine (BADH) was isolated and the enzymatic
accessions and continuous breeding efforts have been properties were determined. The BADH enzyme from
maintained to improve zoysia grass, particularly the the salt-tolerant species Z. tenuifolia has bifunctional
traits of seed propagation, turf density, drought resis- enzyme activities: it functions as betaine aldehyde
tance, salinity tolerance, cold hardiness, and insect dehydrogenase to accumulate betaine aldehyde and as
resistance (Engelke and Anderson 2003). aminoaldehyde dehydrogenase to accumulate polya-
With the aim of identifying zoysia grasses well mines. The betaine aldehyde is a comparatively small
suited for low-input sustainable grazing, two collec- molecule that maintains osmotic balance (Nuccio et al.
tions of the Japanese ecotypes have been performed. 1999), and the polyamine has a huge molecular weight
The first collection occurred on all the Japanese islands and acts as an osmoprotectant (Trossat et al. 1997).
in the 1980s and included all five of the species native In addition to the elucidation of the salt-tolerance
to Japan. Materials from this collection were evaluated mechanisms, isozyme patterns and genomewide poly-
at the Japanese National Agricultural Research Center morphisms also have been studied to estimate phylo-
for the Tohoku Region and conserved in the Japanese genetic relatedness and trace the evolutionary path of
National Livestock Breeding Center (Fukuoka 2000; zoysia grass (Table 16.1). Yaneshita et al. (1997) used
Fukuoka et al. 2009). Several ecotypes were selected the restriction fragment length polymorphism (RFLP)
for good turf quality, and one of them has been markers to estimate the genetic variation among five
released as “Tsukuba Green.” The second collection species of Zoysia collected from natural populations
was organized by the Japanese Ministry of Agriculture, in Japan by analyzing nuclear and chloroplast DNA.
Forestry and Fisheries and focused intensively on Based on the analysis of genetic variation by RFLP of
grasses for low-input sustainable grazing. More than ribosomal and chloroplast DNA of five Zoysia species,
1,200 accessions of Z. japonica were collected and Akiyoshi et al. (1998) reported that salt tolerance of
evaluated (Ebina et al. 2000a), and these are main- zoysia grass originated from the nuclear DNA of
tained at the Japanese National Institute of Livestock Z. matrella and was not maternally inherited. Ampli-
and Grassland Science, supported by the Genebank of fied fragment length polymorphism (AFLP) variation
the National Institute of Agrobiological Science in zoysia grass also has been investigated, and genetic
in Japan. Z. japonica “Asagake” was selected for its diversity and its geographical variation among natural
ability to rapidly establish grazing fields, and “Asamoe” populations have been characterized using AFLP
300 S. Tsuruta et al.

Table 16.1 Molecular marker-based isozyme and DNA profiling studies in zoysia grasses
Target Marker system No. of markers No of accession Species References
Protein Isozyme 1 24 Zj Yamada and Fukuoka (1984)
Isozyme 2 182 Zm, Zs Weng (2002)
Isozyme 2 131 Zm, Zt, Zs Weng et al. (2007)
Nuclear RFLP 20 17 Zj, Zm, Zt, Zs, Zmac Yaneshita et al. (1993, 1997)
RAPD 12 131 Zm, Zt, Zs Weng et al. (2007)
AFLP 4 46 Zj, Zm, Zt, Zs, Zmac Ebina et al. (2000a)
AFLP 10 20 Zj Hong et al. (2008)
SSR 12 24 Zj, Zm, Zt, Zs, Zmac Tsuruta et al. (2005)
SSR 12 41 Zj, Zm, Zt Hashiguchi et al. (2007)
SSR 30 20 Zj Ma et al. (2007)
Chloroplast RFLP 10 17 Zj, Zm, Zt, Zs, Zmac Yaneshita et al. (1997)
Sequence 6 3 Zj, Zm, Zt Tsuruta et al. (2008)
SSR 21 3 Zj, Zm, Zt Tsuruta et al. (2008)
Species: Zj: Z. japonica, Zm: Z. matrella, Zt: Z. tenuifolia, Zs: Z. sinica, Zmac: Z. macrostachya

markers (Fig. 16.2). Recently, random amplified poly- temperatures than the other species. Z. matrella and Z.
morphic DNA (RAPD) analysis (Weng et al. 2007), tenuifolia are native to coastal areas in tropical and
simple sequence repeat (SSR) or microsatellite analy- subtropical regions and have poor cold tolerance, but
sis (Tsuruta et al. 2005; Ma et al. 2007), and sequence their tolerance of drought, heat, and salinity is excellent.
variation of chloroplast DNA (Tsuruta et al. 2008) The three species of Zoysia are allotetraploids, with
were applied to clarify the phylogenetic relatedness a chromosome number of 2n ¼ 4x ¼ 40, and they can
among Zoysia species. However, the phylogeny and generate interspecific hybrids in all combinations
taxonomy of this genus remain unclear, and leaf width, (Forbs 1952; Arumuganathan et al. 1999). The species
traditionally used as an indicator for species discrimi- are also protogynous hermaphroditic plants with both
nation, shows continuous variation among species, a pistil and three stamens in a single flower. The pistil
making species identification difficult. More recently, matures 7–10 days earlier than stamens, and the green
microsatellite markers have been developed based or yellow lemma turns dark brown at the tip as the
on chloroplast DNA sequences of Zoysia (Tsuruta pistil matures (Honda and Kono 1963). The difference
et al. 2008), and phylogenetic clarification utilizing in maturation stage between the pistil and stamens
these molecular markers from the organellar genome promotes outcrossing, although plants can be artifi-
is now in progress. cially self-pollinated at the stage when both the pistil
and stamens are mature because of the lack of physio-
logical self-incompatibility mechanisms (Asano and
Aoki 1998).
16.3 Classical and Molecular Genetic
1. Interspecific hybrids of Zoysia have proven to be
Studies highly effective in the genetic analysis of commer-
cially important traits, such as environmental adapt-
Among the members of the genus Zoysia, Z. japonica, ability. Genetic linkage maps of Zoysia have been
Z. matrella, and Z. tenuifolia are used commonly in constructed on the basis of RFLP (Yaneshita et al.
parks and as sports turf (Ishida 1990; Engelke and 1999), AFLP (Ebina et al. 1999; Cai et al. 2004), and
Anderson 2003). Also, in the coastal and cultivated microsatellite markers (Cai et al. 2005) using the
volcanic mountainous areas of Japan, Zoysia species progeny obtained from interspecific hybrids of Z.
have been used successfully for livestock grazing japonica and Z. matrella. Yaneshita et al. (1999)
(Ishida 1990; Akamine et al. 2005). The natural popula- and Cai et al. (2004, 2005) used the self-pollinated
tions have different distributions, reflecting the genus’ progenies of the ecotype or F1 clone of Z. japonica 
environmental adaptability. Z. japonica is adapted to Z. matrella hybrids as the mapping populations,
warm climates and is more resistant to low whereas Ebina et al. (1999) employed F1 progenies
16 Zoysia 301

SIMILARITY
0.6 0.65 0.7 0.75 0.8 0.85 0.9 0.95 1.0

16 Z. macrantha I
26
24 Z. sinica var . sinica II
25
17
21
III
19 Z. macrostachya
18
20
48
45 IV
Z. japonica
46
47
2
15
3
10
11
V
6 Z. matrella
5
1
8
9
4
12
Z. matrella
13
36
38
Z. tenuifolia VI
35
37
14 Z. matrella
43
44
41 Z. sinica var. nipponica
40
39
7 Z. matrella
31
33
Z. tenuifolia
34
VII
32
42
22 Z. sinica var. nipponica
23
30
29
Z. tenuifolia
28
27

Fig. 16.2 Phenogram of the 46 accessions including five species of Zoysia based on the unweighted pair-group method average
using 206 bands obtained with four AFLP markers

produced from crossing between Z. japonica and bands were obtained from the maternal parent Z.
Z. matrella ecotypes (Fig. 16.3). In the linkage anal- japonica, and most of them did not segregate in the
ysis using the F1 population, 267 polymorphic bands F1 population, indicating that the maternal ecotype
from the pollen parent Z. matrella were observed by used in this study had higher homozygosity com-
using 30 AFLP primer combinations, of which 252 pared to the pollen parent Z. matrella. This accession
bands (94.4%) were segregated at a 1:1 ratio (chi- of Z. japonica was collected from an extremely
square test, P >0.01). In contrast, 41 polymorphic isolated, high-elevation natural population in
302 S. Tsuruta et al.

Fig. 16.3 The parents of the F1 progeny. Accession P29 is an ecotype that is typical of Z. japonica but lacks anthocyanin in the
runner. Accession P272 is a Z. matrella ecotype with a narrow leaf

Japan, suggesting that the high homozygosity in this ingredients. However, the characterization and identi-
accession was associated with geographical factors. fication of major genes governing such qualitative
Using 252 markers derived from 30 primer combina- traits can also be accomplished in Zoysia.
tions, Ebina et al. (1999) successfully constructed the
AFLP linkage map of zoysia grass with 32 linkage
groups covering a total of 1,600 cM, with the average
distance between markers being 6.4 cM. Further- 16.4 Crop Improvement Using
more, the F2 population, consisting of 93 individuals Traditional and Advanced Tools
and produced by selfing the individuals in the F1
segregating population, was analyzed using 159
Zoysia grasses have been used as economically
AFLP markers derived from 24 primer combina-
and environmentally desirable landscape and forage
tions. The linkage map consisted of 27 linkage
grasses. In Japan, the use of zoysia grass was recorded
groups with a total map distance of 1,418 cM and
in the Sakuteiki, a Japanese gardening book published
an average distance of 8.9 cM between markers.
in 1156, and its commercial use began in the 1700s
Segregation of the main characteristics of zoysia (Kitamura 1970). However, the selection and breeding
grass was observed in the F1 and F2 populations (Ebina of zoysia grass is a relatively recent phenomenon.
et al. 2000b). Pigmentation in the stolon from antho- Zoysia grass was introduced to the US from Japan
cyanin, which was absent in the maternal parent in 1902 (Meyer and Funk 1989). Z. japonica “Meyer”
Z. japonica, segregated at a 3:1 ratio for the AFLP was the first variety developed jointly by the USDA
markers, indicating that the trait is determined by a and the US Golf Association. “Meyer” was originally
single major dominant gene. The anthocyanin locus selected from a population of plants grown from seed
was found to be closely linked to AFLP markers in 1941 and was released in 1951 (Grau and Radko
B4-182 and E5-42 (Fig. 16.4). Because Zoysia species 1951), and it is the most widely used zoysia grass
are vegetatively propagated by stolons and rhizomes, cultivar in the US. In 1955 “Emerald,” selected
F2 seeds should be easily obtained by self-fertilization from an interspecific hybrid between Z. japonica and
of propagated F1 clones. Therefore, genetic Z. tenuifolia, was released by the USDA and the
approaches using large-scale mapping populations Georgia Agricultural Experiment Station (Forbs
are available to identify and isolate the major genes 1962). It combines the fine texture of Z. tenuifolia
governing important traits in zoysia grass. Recently, with the cold tolerance and faster rate of spread of
the availability of the inclusion of anthocyanin in the Z. japonica. Subsequently, some commercial cultivars
abovementioned F1 and F2 populations has stimulated of zoysia grass such as “Midwest” (released in 1963),
an interest in the study of their potential as functional “El-Toro” (released in 1986), and “Belair” (released
16 Zoysia 303

Dis Marker Distance from

1998). Z. matrella “Winter Carpet” and “Winter
cM name ANTC locus Field” were released by Nagatomi et al. (1993, 1998)
in 1995 and 1996. These cultivars were first produced
Group 1 by gamma-irradiation mutagenesis in the sod; they
B1-112
exhibit a completely distinct character: retaining
green leaves under the same seasonal conditions in
28.2 which other varieties of Z. matrella lose their green
leaves. Thus far, 41 zoysia grass varieties have been
A4-92 released in Japan. Most cultivated zoysia grasses have
11.3
A4-80
been improved through conventional breeding meth-
1.2
1.7 A8-96 ods such as clonal selection, hybridization, and muta-
5.5 E2-105
10.6 D6-65 genesis. Because zoysia grasses show great natural
A7-65 variation, these will likely be the main methods used
6.1
A2-341
12.8
for the development of new varieties of zoysia grass.
D2-151 As noted above, DNA-based molecular markers
5.3
D1-236 such as RFLPs, AFLPs, and SSRs have been deve-
9.3
C4-172 loped and are being exploited increasingly in the con-
struction of genetic linkage maps of zoysia grass.
In the near future, breeders may be assisted by DNA
analysis, which confirms the degree of genetic varia-
tion in zoysia grass. Ebina et al. (2000b) have con-
75.4 ducted a genetic linkage analysis of several important
traits such as leaf width, salt tolerance, and freezing
hardiness using F1 and F2 populations derived from an
interspecific hybrid of Z. japonica and Z. matrella.
Leaf width is used to discriminate among Zoysia spe-
E5-42 12.9 cM
cies and is one of the most visible determinants of turf
12.9
0.0 cM quality (Kitamura 1970; Turgeon 1996). According to
5.4 ANTC
4.2 B4-182 5.2 cM QTL analysis of several morphological traits, leaf
B4-157 width is controlled by two major QTLs and one
minor QTL. The major QTLs, which are located in
38.8 linkage groups 7 and 15, have significant effects, and
the region around the D2-443, AFLP marker has an
especially large effect (Fig. 16.5a; Table 16.2). These
E4-139 results suggest that the genetic factors controlling leaf
width in zoysia grass might be relatively simple,
Fig. 16.4 Linkage group 1, which contains the anthocyanin
although most morphological traits for turf species
locus (ANTC) and tightly linked AFLP markers show continuous phenotypic variation and are con-
trolled by QTLs. In addition, a linkage peak at a
similar position to that for leaf width has been identi-
in 1987) have been selected from collected genetic fied with leaf length, runner length, and the length of
resources and populations of plants grown from seed. the stem at heading, suggesting that these traits are at
Additional cultivars based on zoysia grass collections least partially controlled by the same QTL.
have been developed and released since the 1990s Although salt tolerance segregated in the F1 popu-
(Asano and Aoki 1998). By the 1980s, the breeding lation, QTLs with LOD scores higher than 3.0 were
of zoysia grass had started in Japan as well. “Miyako,” not detected (Ebina et al. 2000b). Two QTLs with
a naturally interspecific hybrid between Z. japonica LOD scores greater than 2.0 were detected in linkage
and Z. matrella that was originally selected, was the groups 15 and 28, respectively (Fig. 16.5b, Table 16.2).
first cultivar registered in Japan (Asano and Aoki In addition, a peak identified with leaf width is at a
304 S. Tsuruta et al.

Fig. 16.5 LOD scores from the QTL analysis of leaf width (a) and salt tolerance (b) of the F1 population derived from the cross
between Z. japonica and Z. matrella

similar position in linkage group 15 as a peak for

salinity tolerance. In a study of barley, Mano and
Table 16.2 Location of QTLs affecting leaf width and salt Takeda (1997) reported that the QTLs for the most
tolerance effective abscisic acid response, which is correlated
Character Peak marker Locus LOD Contribution ratio
with leaf growth, were located very close to those for
Leaf width D2-433 7 3.55 0.266
salt tolerance. Therefore, genetic linkage between
E5-97 7 3.37 0.246
salinity tolerance and leaf width may be a characteris-
D6-278 7 3.15 0.236
B6-256 15 2.62 0.208 tic common among members of the Poaceae. This
A3-165 7 2.62 0.201 finding should assist in the elucidation of salt tolerance
A3-59 7 2.62 0.201 mechanisms in Zoysia and related genera.
Salt tolerance A6-383 15 –2.08 –0.163 Zoysia grass is one of the most tolerant of freezing
H2-158 28 –2.04 –0.160 among the C4 turf grasses. Freezing tolerance is often
B6-256 15 –1.78 –0.146 evaluated by measuring electrolyte leakage or re-
B3-202 28 –1.33 –0.104
growth of plant tissues after freezing. A QTL for
D3-172 15 –1.23 –0.098
B5-202 28 –1.19 –0.094 freezing tolerance of zoysia grass was detected in
linkage group 25 in the F2 population (Table 16.3),
16 Zoysia 305

Table 16.3 Location of QTLs affecting freezing tolerance problems, genetic engineering represents a new avenue
Peak marker Locus LOD Contribution ratio for the rapid production of unique varieties of Zoysia.
B1-227 25 –6.49 –0.294
E2-177 6 –1.64 –0.084
E1-478 24 –1.24 –0.065
B4-124 18 1.18 0.060 16.5 Controlling Weeds in Zoysia Grass

Weed invasion is a potentially great problem when

but no significant QTL was detected in the F1 popula- establishing and maintaining turf. Because zoysia
tion (Ebina et al. 2000b). This result suggests that grass is slow to propagate and establish, early weed
freezing tolerance in zoysia grass is conferred by a control is especially important. Weed control methods
dominant gene, which was homozygous in the female for turf grasses, including zoysia grass, can be divided
parent, Z. japonica. The QTL was located in the region into two categories: cultural and chemical.
of B1-227 in linkage group 25, with a 0.294 contri- Cultural management of weeds in zoysia grass
bution rate. includes the use of mowing, fertilization, and irriga-
In zoysia grass, morphological characters that tion (Turgeon 1996). In addition, varieties of zoysia
can be assessed visually are important factors in deter- grass that are genetically resistant to biotic stress can
mining turf quality, and QTLs for tolerance of envi- be utilized for weed control. Busey (1982) reported
ronmental stressors such as salinity and freezing are that genotypes of zoysia grass with lower densities of
genetic elements that may be widely distributed stinging nematode infestation also had less abundant
among species. Zoysia grass is widely used in many weed populations.
situations, such as sports fields and golf courses, as Since the discovery of the selective toxicity of
well as for livestock grazing. Information on the asso- synthetic organic herbicides such as 2,4-dichlorophe-
ciation between genetic markers and important traits noxyacetic acid (Marth and Mitchell 1944), there has
will be available for the diversification and advance- been considerable research on chemical management
ment of Zoysia varieties in the near future. of weeds in turf grass. Herbicides are applied prior or
Transgenic approaches to genetic manipulation subsequent to the emergence of target weed species;
offer the opportunity to generate unique genetic varia- these are called pre- and post-emergence herbicides,
tion. Plant regeneration systems for zoysia grass have respectively. Although both pre- and post-emergence
been established from protoplasts (Asano 1989; Khayri herbicides effectively control weeds in zoysia grass
et al. 1989; Inokuma et al. 1996) and embryogenic (Johnson 1996a, b), the use of pre-emergence herbi-
calluses (Bae et al. 2001). Recently, transgenic zoysia cides is time consuming and expensive and often
grass plants were produced by polyethylene glycol- affects the ability to reestablish zoysia grass. Conse-
mediated (Inokuma et al. 1998) and Agrobacterium- quently, post-emergence herbicides are often used
mediated transformation (Toyama et al. 2003; Ge et al. to remove weeds. Post-emergence herbicides are of
2006; Zhang et al. 2007). Rahman et al. (2003) estab- two types: non-selective and selective. A non-
lished a plant regeneration system from calluses selective herbicide provides control of most plants,
derived from the apical meristems of seedlings and whereas a selective herbicide is designed to control a
successfully produced transgenic zoysia grass plants specific type of plant, usually annual grasses or broad-
using Agrobacterium-mediated transformation of leaf weeds. Currently, there are no herbicides with
embryogenic calluses (Fig. 16.6). In addition, some adequate selectivity for controlling most perennial
useful genes have been introduced into zoysia grasses grasses. Therefore, non-selective herbicides must be
by Agrobacterium-mediated transformation to used for controlling these weeds (Turgeon 1996).
improve their resistance to a variety of biotic and Herbicide-tolerant zoysia grass also has been
abiotic stressors (Li et al. 2006; Zhang et al. 2007). developed by Agrobacterium-mediated transformation
The transgenic plants can be grown by vegetative (Toyama et al. 2003). All Zoysia species are sexually
propagation of stolons and rhizomes. Although there compatible, and thus it is essential to consider the
is some concern that large-scale releases of transgenic environmental impact of using transgenic Zoysia. In
plants may cause serious ecological and environmental confined and unconfined test fields, Bae et al. (2008)
306 S. Tsuruta et al.

Fig. 16.6 In vitro apical meristematic culture and genetic callus induced from apical meristems of regenerated plants via
transformation of Z. japonica mediated by Agrobacterium tume- seed-derived callus; (e) plants regenerated from callus; (f, g)
faciens. (a) Highly regenerative embryogenic callus induced transient GUS expression on hygromycin-resistant callus after
from seed in Z. japonica; (b) initiation of plant regeneration; infection with A. tumefaciens; GUS expression in leaves of
(c) established regenerated plant from callus; (d) embryogenic transgenic (h) and control plant (i)
16 Zoysia 307

assessed the potential environmental and biodiversity In addition to maintaining the genetic resources of
effects by investigating major traits equivalence, the Zoysia, a robust means of varietal identification is also
ability to cross-pollinate, and gene flow between trans- required for the effective management of collected
genic zoysia grasses containing a herbicide resistance genetic resources and the assessment of turf quality.
gene and wild type grasses. The transgenic zoysia Traditional varietal identification of zoysia grass has
grass showed tolerance to Basta, a broad-spectrum been performed primarily by examining morphologi-
glufosinate-based herbicide, whereas the wild type cal characteristics; however, this is often difficult with
grasses stopped growing and died. Bae et al. (2008) closely related cultivars due to ambiguous differences
concluded that the transgenic zoysia grass would not and phenotypic modifications caused by environmen-
pose a significant risk when cultivated outside of the tal factors (Yaneshita et al. 1997). In addition, the ease
test field, because unintended cross-pollination with with which zoysia grass propagates vegetatively adds
and gene flow from transgenic zoysia grass were not another burden with regard to variety protection. Con-
detected in the neighboring weed species they exam- sequently, the enforcement of plant variety protection
ined, but these were observed in wild type zoysia regulations is difficult, and an efficient and effective
grass. Weeds in zoysia grass could be effectively man- method is necessary for the enforcement of plant bree-
aged by utilizing such an herbicide resistant transgenic ders’ rights. Because zoysia grass is primarily propa-
plant. gated by stolons and rhizomes, there should be no
genotypic differences within a given cultivar. There-
fore, the development of molecular markers would be
a valuable tool for plant protection as well as a vehicle
for facilitating breeding improvements and estimating
16.6 Recommendations for Future genetic relatedness among various zoysia grass geno-
Actions types. Several DNA-based molecular markers, inclu-
ding RFLP (Yaneshita et al. 1997), RAPD (Weng et al.
Germplasm is the basic material for plant breeding, 2007), AFLP (Ebina et al. 2000b), and microsatellites
and the most important foundation for a breeding (Tsuruta et al. 2005; Ma et al. 2007), have been deve-
program is an extensive collection of genetically loped for the estimation of genetic variation in zoysia
diverse germplasm. Characterizing and recording all grass. These markers can be useful for evaluating
the details of the plant material in a collection is also phylogenetic relatedness objectively, performing
important to its subsequent management and use. genetic analyses of commercially important traits, pro-
Thus, a germplasm collection is becoming increas- moting the development of new varieties, and ensuring
ingly important in the breeding and all phases of the protection of zoysia grass.
biological research of zoysia grasses. However, zoysia
grass ecotypes are gradually disappearing due to
changes in their natural habitats. In New Zealand, for
example, habitats of endemic species including Zoysia References
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.
Index

b-Glucuronidase, 8 Angleton grass, 92

2C-value, 154 Anguina, 165
Anther culture, 169
A Anthocyanin, 302
Abiotic Apomicts
stress, 58 facultative apomict, 97, 99, 102, 204, 212
tolerance, 190 obligate apomict, 97
Abscisic acid (ABA), 6, 26, 304 Apomictic, 37, 138, 142, 182, 198, 208, 227
Abscission, 239 gene, 47
Accumulation/loss (A/L), 228 clone, 143
Adaptability, 139, 203, 300 pseudogamous obligate apomict, 211
Adaptation, 64, 82–83, 155, 267 trait, 245
Adaptive polymorphism, 100 Apomixis, 47, 97, 99, 100, 201, 245
Adaptive radiation, 82 aposporous apomixis, 190, 206
Agamic, 90 diplosporous apomixis, 138, 187
complex, 97, 203 facultative apomixis, 208
Agrobacterium, 7, 26, 105, 159, 169 gene, 49
transformation, 305 Apomixis-related gene region (ASGR), 184
Agroforestry, 211 Apospory, 47, 199, 209
Agronomic traits, 127, 204 Apospory-specific genomic region (ASGR), 47, 48,
Agrostis 227, 245
A. alba, 3 Arabidopsis, 9, 245
A. canina, 2 Aromatic, 96
A. capillaris, 2, 3 Asexual revolution, 50
A. gigantea, 3 Association mapping, 84
A. palustris, 2 Autoallooctoploid, 19
A. scabra, 4 Autogamous, 205
A. stolonifera, 2, 9 Autogamy, 280
Allergy, 271 Autoploidy, 279
Allogamous, 22, 158 Autopolyploid, 123, 138, 201
Allohexaploid, 223 Autotetraploid, 79, 96, 183, 268
Allopolyploidization, 123 Aveneae, 16
Allopolyploidy, 3, 98 Aveneae tribe, 1
segmental, 138
Allotetraploid, 19, 22, 23, 114, 144, 209, 225 B
segmental allotetraplod, 38, 211 Backcrossing, 157, 199, 227
Allotetrasomic, 47 Bacterial artificial chromosome (BAC), 157, 170, 288
Allozyme, 282 clone, 47
Amphidiploid, 25, 157 contig, 185
Amplified fragment length polymorphism (AFLP), 3, 19–20, 42, library, 47
66, 125, 140, 157, 168, 183, 201, 237, 287, 299 Bahiagrass, 208
Anamorph, 128 Bayesian analysis, 234
Ancestral species, 123–124 B-chromosome, 227
Androgenesis, 157 Bentgrass
Aneuploid, 31, 96, 222 colonial bentgrass, 2
Aneuploidy, 39 creeping bentgrass, 1, 9

C. Kole (ed.), Wild Crop Relatives: Genomic and Breeding Resources, Millets and Grasses, 311
DOI 10.1007/978-3-642-14255-0, # Springer-Verlag Berlin Heidelberg 2011
312 Index

Bentgrass (cont.) Burrgrass

highland bentgrass, 3 American burrgrass, 36
Idaho bentgrass, 3 hillside burrgrass, 33
redtop bentgrass, 3 large burrgrass, 33
velvet bentgrass, 2
Bermudagrass C
African bermudagrass, 57 Candidate gene, 271, 286
coastal bermudagrass, 68 Capillipedium, 97
Biodiversity, 147, 248 Capim-pojuca, 211
Bioenergy, 58 Carbon isotope discrimination (CID), 43
Bioethanol, 192 C-banding, 22
Biofuel, 147 C3 species, 178
Biolistics, 8, 159 C4 cereal, 275
Biomass, 56, 105, 192, 246 C4 grasses, 178, 297
Biosystematics, 96 C4 photosynthesis, 136, 178, 180
Biotic stress, 42, 305 C4 species, 178
Biotype, 204 cDNA
Birdwood grass, 36 clone, 48
Blast disease, 127 library, 43, 140, 144, 159, 160, 170, 191
BLASTX, 145 microarray, 48
Bluegrass Cenchrus, 31
gulf bluegrass, 92 C. argimonioides, 32
king bluegrass, 95 C. biflorus, 32
nadi bluegrass, 92 C. brownii, 33
Queensland bluegrass, 95 C. calicalatus, 33
Bothriochloa, 97 C. ciliaris, 33, 42, 48
Bridging species, 100 C. distichophyllus, 33
Brome C. echinatus, 33
Alaska brome, 25 C. elymoides, 34
Arizona brome, 25 C. gracillimus, 34
California brome, 25 C. incertus, 34
Mountain brome, 25 C. longispinus, 34
Russian brome, 18–20 C. melanostachyus, 34
Sitka brome, 25 C. mitis, 35
Bromegrass C. multiflorus, 35
arctic bromegrass, 21 C. myosuroides, 35
erect bromegrass, 21 C. palmeri, 35
field bromegrass, 23 C. pennisetiformis, 35
meadow bromegrass, 20–21 C. pilosus, 35
Bromus C. platyacanthus, 36
B. arizonicus, 16 C. prieurii, 36
B. arvensis, 23 C. robustus, 36
B. auleticus, 16 C. setiger, 36
B. biebersteinii, 16 C. somalensis, 36
B. carinatus, 16, 25 C. spinifex, 36
B. catharticus, 15, 16 C. tribuloides, 36
B. erectus, 16, 21 Center of origin, 208
B. inermis, 15, 16, 20, 26 Centromere, 227
B. mango, 16 Chiasma, 227
B. marginatus, 16, 25 Chilochloa, 258
B. mollis, 16 Chloridoid, 130
B. pumpellianus, 21 Chloridoideae, 113, 135, 297
B. riparius, 16 Chloroplast, 66–67, 103, 168, 197, 264
B. rubens, 16 DNA (cpDNA), 18, 23, 123, 299
B. sitchensis, 15, 25 gene, 180
B. valdivianus, 16 genome, 26, 74
B. variegatus, 21 Chlorosis, 4
Brunswick grass, 212 Chromatin bridge, 138
Buffelgrass, 47, 135 Chromosome
cloncurry buffelgrass, 35 deletion, 271
Bulked segregant analysis (BSA), 236, 271 doubling, 96, 138, 211
Index 313

elimination, 210 D. pallidum, 95

organization, 221 D. panchaganiense, 95
pairing, 18, 98, 210 D. queenslandicum, 95
rearrangement, 200, 222 D. sericeum, 95
substitution, 271 D. setosum, 95
Cladosporium phlei, 271 D. tenue, 95
Clustering, 168 D. tenuiculum, 95
Coelachyrum, 49 D. woodrowii, 95
Colchicine, 183 Digestibility, 68, 143
Cold Dilatata complex, 202
hardiness, 299 Diploid, 17, 74
tolerance, 210 Disomic, 47
Colinearity, 126, 200 DNA
Colonization, 284 amplification fingerprinting (DAF), 65–66
Comparative genetic mapping, 126, 159 content, 58
Comparative mapping, 243 Data Bank of Japan (DDBJ), 288
Compilospecies, 90 insertion, 264
Conservation, 180, 192, 270 sequence, 27, 125, 208, 224, 300
Conserved intron scanning primers (CISP), 145 Dollar spot, 7
Cool-season forage, 170 Domestication, 41, 145–146, 230, 238
Cool-season grass, 4, 159 history, 287
Core collection, 62, 270 syndrome, 238, 278
Corynebacterium, 165 Dormancy, 80, 108, 281
Cross-pollination, 66, 307 Doubled haploid, 169, 230
Crown rust, 155, 159 Downy mildew, 107
Cryopreservation, 63, 270 Drought, 42, 192
Cynodon hardy, 211
C. aethiopicus, 54 resistance, 299
C. arcuatus, 54, 55 stress, 139, 160
C. barberi, 54, 55 tolerance, 59, 103, 139, 158, 246, 282
C. dactylon, 53, 54 tolerant, 3
C. incompletus, 54 Dry matter (DM), 139
C. nlemfuensis, 54 Duodecaploid, 25
C. plectostachyus, 54 Dysploidy, 224–227, 266
C. transvaalensis, 54
C. x magennisii, 54 E
Cytogenetic diversity, 96 Ecosystem, 147, 155
Cytokinin, 6 Ecotype, 97, 155, 177, 299
Cytoplasmic male sterility (CMS), 10, 167, 246 Edaphic race, 82
Cytotypes, 65, 181, 183, 206, 258 Elephant grass, 222
Eleusine, 113
D E. africana, 117
Dactylis, 270 E. coracana, 114
D. glomerata, 73 E. floccifolia, 118
Dallisgrass, 203–205 E. indica, 118
prostrate dallisgrass, 205 E. intermedia, 118
Dead spot, 6 E. jaegeri, 118
Desmodium, 247 E. kigeziensis, 118–119
Dichanthium E. multiflora, 119–120
D. affine, 91 E. tristachya, 120
D. annulatum, 90, 91 Embryo
D. aristatum, 92 rescue, 144
D. armatum, 92 sac, 184, 209
D. caricosum, 92 Embryogenesis, 169, 210
D. compressum, 92 Embryology, 98
D. fecundum, 92 Endophyte, 27, 170, 271
D. foveolatum, 95 Endophytic, 170–171
D. humilius, 95 fungi, 27, 271
D. maccannii, 95 Epichlöe, 271
D. mucronulatum, 95 Epigenetic, 189
D. oliganthum, 95 Eragrostis, 130
314 Index

Eragrostis (cont.) differentiation, 168

E. aethiopiaca, 137 expression, 169
E. bicolor, 137 expression chip, 271
E. curvula, 136, 143 flow, 100, 109, 217, 235, 238, 284
E. japonica, 146 ontology, 145
E. lehmanniana, 136, 146 pool
E. pilosa, 136 primary, 83, 221, 246, 269, 277
E. scotelliana, 146 quaternary, 84, 270
E. tef, 138 secondary, 83, 269, 277
E. tremula, 146 tertiary, 37, 83, 246, 269, 277
Ergot disease, 107 wild, 235
Ergovaline, 171 restorer gene, 289
Ethanol, 247 technology, 143–144, 158
Euploid, 96 GeneBank, 48, 49, 77, 83, 124, 128, 155, 171, 270
European Molecular Biology Laboratory (EMBL), 288 Genetic
database, 48 background, 143
Evolution, 75, 221 bottleneck, 284
progressive, 100 differentiation, 66, 237
Evolutionary distance, 234
center, 65 diversity, 19–20, 26–27, 62, 65, 66, 126, 155, 168, 209,
event, 65 233, 284, 299
lag, 79 drift, 238, 284
reconstruction, 225 engineering, 305
system, 212 erosion, 61
Expressed sequence tag (EST), 2, 48, 84, 126, 140, instability, 157
145, 168 isolation, 66
sequence, 187 linkage map, 141, 201
map, 43, 125, 126, 159, 236, 238, 288
F polymorphism, 158
Fertility, 99, 121 relatedness, 66
Fescue screening, 127
meadow fescue, 155 similarity, 140
tall fescue, 158 structure, 168, 221, 283
Festuca, 73 transformation, 8, 105, 143
Festuca Gene Index (FaGI), 160 variation, 66, 79, 140, 155, 158
Festuca, 153 Genome, 19, 206
Festulolium, 157 donor, 121
F1 hybrid, 157 evolution, 154
Finger millet, 113 mapping, 26
Flavone, 77 sequence, 124
Flavonoid, 78 sequencing, 288
Flow-cytometry, 145, 182 size, 16, 57–58, 83, 221, 260
Flowering behavior, 257, 258 Genomic, 126, 238
Fluorescence in situ hybridization (FISH), 19, 38, 47, 158, 207 constitution, 25
Fodder, 145–146 DNA, 102
Forage, 56, 73, 160 organization, 238
Foxtail Genomic in situ hybridization (GISH), 19, 121, 158, 245,
bristly foxtail, 277 266, 278
giant foxtail, 277 Genotype matrix mapping (GMM), 190
giant green foxtail, 281 Genotyping, 160
millet, 275 Germination, 286
F1 population, 301 Germplasm, 27, 85, 155, 180, 269, 307
F2 population, 302 bank, 49
Freezing, 54 collection, 60–63
hardiness, 303 conservation, 60–63
tolerance, 158 Germplasm Resource Information Network (GRIN), 33, 49,
62, 100, 139, 190
G Giemsa C-banding, 19
Gametophytic, 183 Gigantism, 239
Gandhel, 95 Glacial relict, 263
Gene Glaciation, 75, 261–263
Index 315

Glyphosate, 129 Intraspecific hybridization, 203

Golf course, 155, 302 Intraspecific variation, 138
Goosegrass, 129 Introgression, 20, 64, 235, 269, 289
Grain hybrid, 81
quality, 287 map, 157
size, 286–287 Introgressive hybridization, 97
Gramineae, 96, 153 Intron, 158
GrassBase, 175 Invasion, 6, 284, 305
Grazing, 118, 135, 155 Invasive, 17, 147
Guineagrass, 183 species, 191
weed, 41
H Invasiveness, 160
Haploid, 221 Inversion, 200
Heat In vitro culture, 143
shock protein (HSP), 4 In vitro dry matter digestibility (IVDMD), 142
stress tolerance, 4 Isoenzyme, 43, 234
Herbicide, 305 Isozyme, 102, 299
resistance, 26, 129, 282, 289
K
tolerance, 165, 271
Karyotype, 19, 84–85, 137, 221, 260
Heritability, 287
Kleingrass, 135
Heterochromatin, 260
Heterosis, 245 L
Heterozygosity, 80, 190 Lamarckia aurea, 75
Hexaploid, 19, 24, 57, 82, 97, 181, 204, 257 Landrace, 287
Homoeologous pairing, 157 Landscape, 302
Homology, 138, 204 Lasiurus, 49
Housekeeping gene, 169 Leaf spot, 42
Hybridity, 157 Lignin, 102
Hybridization, 54, 79, 124, 130, 160, 167, 212, 238, 268, biosynthesis, 143–144, 159
288–289 Linkage, 221
Hybrid group (LG), 140, 141, 183, 287, 302
breeding, 246 mapping, 144
trispecific hybrid, 246 map, 126, 141, 183, 300
Hygromycin, 169 Lolium, 49, 73, 153
L. canariense, 166
I L. edwardii, 166
Ideogram, 38, 185 L. multiflorum, 165
Inbred line, 167 L. perenne, 157, 165
Indigenous grasses, 247 L. persicum, 165
Inflorescence, 117 L. remotum, 165
architecture, 285–286 L. rigidum, 165
Inheritance, 85, 169 L. subulatum, 166
digenic inheritance, 240 L. temulentum, 165
maternal inheritance, 97 Love grass
monogenic inheritance, 240 boer love grass, 139
Insect resistance, 299 Lehmann love grass, 135
Insertion/deletion (INDEL), 145 stinking love grass, 147
Interecotypic hybridization, 80 weeping love grass, 135
Intergeneric hybrid, 96, 157, 158
Intergenetic hybridization, 9, 38, 158 M
Internal transcribed spacer (ITS), 18, 74, 123, 136, Macrosynteny, 227
167, 224, 261 Magnaporte, 42
International Crops Research Institute for the Semi-Arid Maillea, 260
Tropics (ICRISAT), 128 Major gene, 302
Inter-simple sequence repeat (ISSR), 42, 123, 141–142, Male sterility, 289
208 Manodialdehyde (MDA), 43
Interspecific Mapping, 26, 48
crossing, 59, 287, 297 population, 159, 183, 201, 302
hybridization, 3, 10, 60, 197, 230, 246, 280–281 Marker-assisted selection (MAS), 84, 128
hybrid, 19, 67, 96, 212, 300 Marvel grass, 91
316 Index

Mating system, 279 P. coloratum, 175

Megaspore, 98 P. dichotomiflorum, 175
Meiotic instability, 138 P. maximum, 182
Microcolinearity, 126 P. miliaceum, 175
Microsatellites, 125, 170, 186, 203, 300 P. repens, 175
Microsporogenesis, 98 P. virgatum, 175
Microsynteny, 245 Paraphyletic, 39, 226
Migration, 75, 261–263 Parthenogenesis, 200, 209
Model species, 167 Particle bombardment, 159
Molecular Paspalum
breeding, 160 P. atratum, 211
derivative, 264 P. compressifolium, 212
descent, 74 P. conspersum, 206
diversity, 269 P. coryphaeum, 207
form, 264 P. dasypleurum, 205
mapping, 48 P. denticulatum, 206
profile, 266 P. dilatatum, 201, 202
Monoculture, 146 P. distichum, 210
Monoembryony, 190 P. equitans, 208
Monophyletic, 39, 103, 208 P. exaltatum, 207
Monoploid, 221, 247 P. glaucescens, 212
Monosomic substitution line, 157 P. guenoarum, 211
mRNA, 201 P. haumanii, 207
Mutant, 169 P. intermedium, 202
Mutation, 62, 74, 102, 266 P. juergensii, 202, 207
P. malachophyllum, 201
N P. nicorae, 212
NADP-dependent malate dehydrogenase (NMDH), 105 P. notatum, 200, 202, 208
Napier grass, 246 P. paniculatum, 207
National Plant Germplasm System (NPGS), 62 P. pauciciliatum, 205
National Semi-Arid Resources Research Institute P. plicatulum, 211
(NaSARRI), 128 P. procurrens, 201
Natural selection, 124 P. rufum, 206
Neotyphodium, 271 P. scrobiculatum, 212
Nicotinamide adenine dinucleotide phosphate (NADP), P. simplex, 200, 201
105, 197 P. subciliatum, 209–210
Nuclear DNA, 18, 182, 299 P. urvillei, 205
P. usterii, 201
O P. vaginatum, 202, 210
Ochthochloa, 124 P. virgatum, 206
Octoploid, 181, 182, 258, 266 Pearl millet, 232, 238
Off-type, 275 Pedoturbation, 146
O2 inhibition, 178 Pennisetum, 32, 39, 184
Ophiosphaerella, 6 P. glaucum, 39, 221
Orphan P. mezianum, 223
crop, 130 P. ramosum, 223
species, 145 Pentaploid, 57, 203
Oryza sativa, 245 Peroxidation, 5
Outbreeder, 167 Phenogram, 187
Outcrossing, 237, 279 Phenotypic
Overdominance, 242 heterogeneity, 283
Overexpression, 9 plasticity, 283
Overgrazing, 271 selection, 209
variation, 242
P Phenotyping, 160, 169, 237
Palatability, 157 Phleum, 73, 257
Paleo-allotetraploid, 264 P. alpinum, 258, 261, 263–265
Paniceae, 31 P. arenarium, 259
Panicoideae, 31 P. echinatum, 258, 261, 266–267
Panicum, 49 P. himalaicum, 259–260
P. capillare, 175 P. hirsutum, 258–259
Index 317

P. iranicum, 260 Refugia, 80, 168, 267

P. montanum, 259 Regeneration, 210, 305
P. paniculatum, 259 Relative water content (RWC), 139
P. phleoides, 258 Reporter gene, 9
P. pratense, 257, 258, 265, 266 Resistance (R), 7
Photosynthesis, 178, 290 Restorer, 289
Phylogenetic, 96 Restriction fragment length polymorphism (RFLP), 4, 123, 140,
affinity, 78 157, 167, 186, 199, 234, 278, 299
analysis, 144 R-gene-like sequence (RGL), 7
divergence, 221 Rhizobacteria, 2
relatedness, 187, 300 Ribosomal DNA (rDNA), 121, 123, 185, 223
relationship, 39, 278 RNA interference (RNAi), 109
tree, 123 Ryegrass
Phylogeny, 178, 180, 198, 203, 224, 278 darnel ryegrass, 166
Phylogram, 40 Italian ryegrass, 160, 165
Physical mapping, 185 perennial, 271
Phytoremediation, 130 rigid, 153
PlantGBD, 169
Plasmid artificial chromosome (PAC), 157 S
Pleiotropic, 199 Saccharomyces, 143
Ploidy, 66 Salinity, 108
level, 138, 154, 181, 217 tolerance, 299
Poa, 136, 153 Salt tolerance, 6, 42, 130, 210, 282
P. annua, 6 Sandbur
Poaceae, 1, 31, 135, 297 agrimony sandbur, 32
Pollen, 99 big sandbur, 35
flow, 238 Coastal sandbur, 34, 36
Polyembryo, 183 hedhog sandbur, 33
Polyembryony, 99 Indian sandbur, 32
Polyethylene glycol, 305 Slender sandbur, 34
Polyhaploid, 97 Sclerotinia, 7
Polymerase chain reaction (PCR), 102, 290 Secale cereale, 228
Polymorphic information content (PIC), 170, 187 Seed
Polyploidization, 123–124, 145, 202, 281 bank, 237
Polyploidy, 39, 82, 96, 97, 137, 138, 201 flow, 238
Polyspory, 183 shedding, 285
Pooideae, 1 Segregation, 47, 102, 222
Population structure, 126 distortion, 199, 229
Power of discrimination (PD), 187 Mendelian, 240
Pre-breeding, 247–248 Selection, 237
Principle coordinate analysis (PCA), 187 natural, 237
Protandry, 280 Selectivity, 305
Protogyny, 280 Self-compatibility, 138, 167, 187, 208, 297
Protoplast, 8 Self-fertility, 167, 175, 209
Pseudo-F2, 169 Self-incompatibility, 66, 155, 175, 190, 206
Pseudogamous, 89, 100 Self-sterile, 158
Pseudogamy, 200, 209 Senescence, 4
Pseudomonas, 2 Sequence characterized amplified region (SCAR), 3, 43
Pseudo-testcross, 159 Sequencing, 124, 145
Pyricularia, 42 Setaria, 32, 39, 275
S. adhaerans, 278
Q S. faberi, 277
Quantitative trait loci (QTL), 4, 47, 142, 144, 159, 190, 236, 285, 299 S. geniculata, 278
expression-QTL, 243 S. italica, 277
S. leucopila, 280
R S. machrostachya, 280
Random amplified polymorphic DNA (RAPD), 3, 102, 123, S. parviflora, 277
140, 158, 167, 186, 201, 278, 300 S. pumila, 277
Real-time quantitative PCR (RT-PCR), 169 S. texana, 280
Reciprocal cross, 280 S. verticillata, 277
Recombination, 102 S. verticilliformis, 278
318 Index

Setaria (cont.) Trifluralin, 290

S. viridis, 277 Trifolium, 49
S. yunnanensis, 289 Triploid, 97
Sethoxydim, 290–291 Tripsacum, 189
Sexual Trisomic, 288
diploid, 211 Turf, 56, 160
tetraploid, 211 Turfgrass, 208
Shattering, 168, 238
Sheath blight, 107 U
Shibra, 236, 237 US Department of Agriculture (USDA), 33, 49, 62, 100, 120,
Simple sequence repeat (SSR), 17, 84, 125, 140, 157, 168, 186, 139, 186, 299
233, 271, 288, 300 US Department of Agriculture-Agricultural Research Service
Simplex (USDA-ARS), 62, 84
loci, 183
marker, 183 V
Single nucleotide polymorphism (SNP), 84, 145, 157 Vasey grass, 205
Somatic, 160 Vernalization, 2, 167, 169
hybrid, 160 Vulnerable species, 96
polymorphism, 283
Sorghum, 191 W
Southern hybridization, 169 Water-logging, 210
Speciation, 96, 279 Weed, 305
Stargrass, 59 aggressive, 244
Substitution, 180 Weediness, 26–27
Supernumerary chromosome, 227 Wheat streak mosaic virus (WSMS), 128
Switchgrass, 186, 190 Wild
Synteny, 130, 159 alleles, 244
form, 237
T phenotype, 236
Tall fescue, 139 population, 236
Tetraploid, 4, 37, 54, 75, 79, 118, 181, 197, 258, 266, 289 progenitor, 239
sympatric tetraploid, 78 relative, 246
Tetrasomic, 21 Winter, 23
Threatened species, 92 hardiness, 42, 56, 246
Tillering, 286
Timothy, 257
Tonic, 146 X
Transcript, 4 Xeromorphic, 80
derived fragment, 160
Transformation, 7, 85, 105, 143, 159, 169, 305 Z
Transgene, 143 Zea mays, 228
Transgenesis, 158, 159, 169 Zoysia, 297
Transgenic, 143–144, 305 Z. japonica, 297
cultivar, 85 Z. macrostachya, 298
plant, 8, 85, 158 Z. matrella, 298
Transgressive, 242 Z. tenuifolia, 298
Triazine, 290 Zoysieae, 297