See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/326319103

Foliar epidermal micromorphology and its taxonomic implications in some

selected species of Athyriaceae

Article  in  Microscopy Research and Technique · April 2018

DOI: 10.1002/jemt.23055

CITATIONS READS

51 1,520

9 authors, including:

Syed Nasar Shah Mushtaq Ahmad

Quaid-i-Azam University Athabasca University
36 PUBLICATIONS   851 CITATIONS    343 PUBLICATIONS   7,835 CITATIONS   

SEE PROFILE SEE PROFILE

Muhammad Zafar Abdul Razzaq

Quaid-i-Azam University University of Central Punjab
352 PUBLICATIONS   7,331 CITATIONS    58 PUBLICATIONS   410 CITATIONS   

SEE PROFILE SEE PROFILE

Some of the authors of this publication are also working on these related projects:

Ferns and their allies in Pakistan View project

Male contraceptives View project

All content following this page was uploaded by Fazal Ullah on 11 July 2018.

The user has requested enhancement of the downloaded file.

Received: 3 February 2018 | Revised: 13 April 2018 | Accepted: 26 April 2018

DOI: 10.1002/jemt.23055

RESEARCH ARTICLE

Foliar epidermal micromorphology and its taxonomic

implications in some selected species of Athyriaceae

Syed Nasar Shah1 | Mushtaq Ahmad1 | Muhammad Zafar1 |

Abdul Razzaq2 | Khafsa Malik1 | Neelam Rashid1 | Fazal Ullah1 |

Majid Iqbal1 | Wajid Zaman1,3,4

1
Department of Plant Sciences, Quaid- i-
Azam University Islamabad, 45320, Pakistan
Abstract
2
Center of Plant Biodiversity, University of For the robust identification of taxonomically complex fern family like Athyriaceae, light and scan-
Peshawar, Peshawar, Pakistan ning electron microscopy is significance implications. This article present first microscopic
3
State Key Laboratory of Systematic and investigation of foliar micromorphology of 3 genera and 10 species belonging to Athyriaceae
Evolutionary Botany, Institute of Botany, namely, Athyrium, Deparia, and Diplazium were collected from different localities in Malakand Divi-
Chinese Academy of Sciences, Beijing, China
sion, Northern Pakistan. In present study we compare foliar micromorphology of all 10 species
4
University of Chinese Academy of Sciences,
using standard protocols of light microcopy (LM) and scanning electron microscopy. Qualitative
Beijing, China
micromorphological variations in shape of epidermal cells, anticlinal wall pattern, stomatal type and
Correspondence shape, stomatal pore shape, guard cells shape, and trichomes types were studied. In addition, some
Syed Nasar Shah, Department of Plant quantitative characters were also studied and data were statistically analyzed in epidermal cell size,
Sciences, Quaid- i- Azam University
stomatal size, stomatal pore size, stomatal density, and stomatal index. The pivotal result of study
Islamabad, 45320, Pakistan.
Email: [email protected] include; shape of epidermal cell in all species is irregular on both abaxial and adaxial surfaces. The
and anticlinal walls are sinuous in most of the species but some species have irregular lobed and
Wajid Zaman, Department of Plant
broadly lobed wall. Leaves are hypostomatic in all studied species. Two main categories of stomatal
Sciences, Quaid- i- Azam University
Islamabad, 45320, Pakistan.
type were found: polocytic and anomocytic. Unicellular nonglandular trichomes were observed in
Email: [email protected] only one species Athyrium mackinnoni. The variation in foliar micromorphological characters
between the genera and within the species was useful in identification and classification and have
Review Editor: Prof. George Perry
potential taxonomic significance for species differentiation. An identification key using micromor-
phological characters are provided to distinguish genera and species.

KEYWORDS
Athyriaceae, foliar epidermis, LM, Pakistan, SEM, taxonomic keys

1 | INTRODUCTION family has been variously circumscribed and its limits are still uncertain
(Hasebe et al., 1995; Liu, Chiou, & Kato, 2011; Wang, Chen, Zhang, Lu,
Athyriaceae a leptosporangiate fern family comprises 3 genera and 650 & Zhao, 2002; Wei, Schneider, & Zhang, 2013). In Pakistan, the family
species worldwide in which two genera Athyrium and Diplazium con- is represented by 23 species belonging to 3 genera and it is commonly
tributed nearly 85% species and third genus is Deparia estimated to distributed in Himalayan and Northern region of Pakistan (Nakaike &
contain 15% species (Schneider et al., 2016; Wei et al., 2013). The fam- Malik, 1992; Stewart, 1967; Stewart, Ali, & Nasir, 1972). In present
ily is characterized by: creeping rhizomes, ascending, or erect, scales at study, we studied 10 species belonging to 3 genera namely, Athyrium,
apices; these usually noncalthrate, glabrous, glandular, or ciliate, Diplazium and Deparia.
petioles with two elongate or crescent-shaped vascular bundles facing Athyrium represent one of the species rich and one of the most
one another, blades monomorphic; rarely dimorphic, veins pinnate or confused genus of Athyriaceae about 230 species are known world-
forking; free, sori abaxial; round J-shaped, or linear with reinform, wide distributed in southwest China, Japan, North America, and tropi-
monolete, perine winged, ridged or spiny (Smith et al., 2006). The cal Asia (Roux, 2009; Schneider et al., 2016; Tryon & Tryon, 2012;

Microsc Res Tech. 2018;1–12. wileyonlinelibrary.com/journal/jemt V

C 2018 Wiley Periodicals, Inc. | 1
2 | SHAH ET AL.

Wang et al., 2002). In Pakistan, the genus is represented by 13 species. taxonomic characters (Carlquist & Schneider, 1997, 2000, 2001; Large,
Generic circumscriptions within Athyrioid ferns has been a subject of 1989; Nester, 1985; Schneider & Carlquist, 1998; Suseela & Devi, 1998;
strong controversy since establishment and many related genera Tryon & Lugardon, 2012). Variations in the leaf micromorphological char-
included in and separated over time (Christensen, 1906; Copeland, acters are very useful in the taxonomic study of many ferns species
1947; Fraser-Jenkins, 2006; Wei & Zhang, 2016). (Chuang & Liu, 2003; Nayar, 1962; Shao, Lu, & Shang, 2011; Van Cot-
Diplazium is the most species rich lineage of the Athyriaceae and them, 1970; Wang, Deng, Li, Deng, & Lu, 2009; Wang & Lu, 2010;
contain about 350 known species (Christensen, 1917; Christenhusz, Wylie, 1949). Epidermal and anticlinal wall pattern provided many impor-
Zhang, & Schneider, 2011; Schneider et al., 2016; Tryon & Tryon, tant taxonomic delimitation in ferns (Van Cotthem, 1970). Epidermal and
1982). Most species are distributed in tropical and subtropical climates stomatal characters are taxonomically very important due to its variabili-
but few species have been found in cold temperate. In Pakistan, the 4 ty. Structure of stomata reported in many species of ferns shows a
species of this genus are reported and distributed in Northern areas remarkable taxonomic significance (Chuang & Liu, 2003; Pant & Khare,
along with Kashmir (Stewart, 1967; Stewart et al., 1972). The generic 1971; Sen & Hennipman, 1981; Viane & Cotthem, 1977; Ziegler, 1987).
delimitation of Diplazium is a subject of strong controversy among pter- Stomata size and frequency is reported to delimit the genera of certain
idologist since long time (Arana, Mynssen, & Ponce, 2017; Ching, 1964; ferns species (Lee & Oh, 1988). Trichomes are reported by various work-
Kato, 1979; Tryon & Tryon, 1982; Wei et al., 2013). ers are useful in taxonomic study of many ferns species (Barthlott,
Deparia contain 70 species distributed in tropical and cold temper-

Wiersch, Colić, & Koch, 2009; Krings, Kellogg, Kerp, & Taylor, 2003;
ate forest in most part of old world (Kato, 1979; Kato & Darnaedi, Nayar, 1956).
1988; Kuo et al., 2018; Schneider et al., 2016). In Pakistan the genus is Despite anatomy of many aspects of Athyriaceae foliar micromor-
represented by 5 species. The three genera can be distinguished mor- phological investigation are very few in the literature (Mali & Yi, 1997;
phologically: Athyrium species are characterized by having two strands Zhang, Ye, Liu, & Jing, 2011). No studied are practically existent on the
of vascular bundles in the petioles; the petiole bases are enlarged and foliar epidermal anatomy and systematics of Athyriaceae in Pakistan.
have conspicuous elaborated aerosphores, continuous grooves from Thus, the study stands as first taxonomic investigation of 3 genera and
rachis to costae and have J or horse shaped sori (Ching, 1964; Kato, 10 species of Athyriaceae namely, Athyrium, Diplazium, and Deparia.
1977, 1979; Rothfels et al., 2012). Most species of Deparia can be The present work aims to study the foliar epidermal anatomy of some
distinguished from Athyrium and Diplazium by fronds architecture and Pakistani Athyriaceae species using LM and SEM techniques to provide
segmentation in fronds and presence of articulate multicellular hairs anatomical data of those species which has not been studied before to
(Kato, 1977). Deparia is usually distinguished from Diplazium by its cos- test the effectiveness of foliar micromorphology of epidermal cell and
tae and rachis grooves. Deparia has grooves on adaxial side of costae other stomatal feature for taxonomic segregation.
that are interrupted at the junction with the rachis, but in Diplazium the
costal grooves are continuous with that of the rachis (Sano, Takamiya,
2 | MATERIALS AND METHODS
Kurita, Ito, & Hasebe, 2000).
Anatomical observation of the Athyriaceae have been made by
2.1 | Plant collection, identification, and herbarium
several workers including Ogura (1921), Bir (1969), and Hayata (1927).
deposition
Ogura (1921) studied the stelar anatomy of athyrioid ferns and found
that they are all dictyostelic type. In 1971, Bir described the stelar Plant materials used in this study were collected by the authors from 3
structure of rhizome and stipe of Himalayan Athyrium as being similar Districts of Malakand division during April 2016 to March 2017.
to that of Diplazium. According to the study of Kato (1972) on Athyria- Malakand division situated in the Northern area of Pakistan,
ceae, the vascular structures of rhizomes, stipes and rachis of Athyrium, Khyber PakhtunKhwa province and lies between 348- 22ʺ to 348-
Diplazium and their relatives are very similar to those of Acystopteris, 41ʺ North latitudes and 718-37ʺ to 728-14ʺ East longitudes. Plant
Gymnocarpium, onoclea and Woodsia. Beside these treachery elements species collected (Figure 1) within altitudinal range of 100–4,000 m.
is used for taxonomic purpose in this family (Xu & Wang, 2009) sto- Preliminary identification of the species was performed by comparing
mata types (Yan-Ping & Jian-Meng, 2012) vascular bundles (Karafit, species with authentic herbarium specimens housed in Herbarium of
Rothwell, Stockey, & Nishida, 2006) and the petiole anatomy (Hernan- Pakistan (ISL) QAU, Islamabad and National Herbarium of Pakistan
dez-Hernandez, Terrazas, Mehltreter, & Angeles, 2012) have been used (NARC) Islamabad. In addition the species were confirmed based on
for the taxonomic purpose in Athyriaceae. However, on the basis of characters mentioned in original description of negioubring country
morphological and few earlier anatomical studies the grouping of the flora of China and those used by previous authors (Nakaike & Malik,
taxa looks very artificial some of the phylogenetic studies were per- 1992, 1993; Stewart, 1967; Stewart et al., 1972).
formed which can possibly help in the delimitation of genera and spe- The collected specimens were dried, pressed, labeled and mounted
cies within Athyriaceae (Chunxiang, Shugang, Xiaoyan, & Qun, 2011; on herbarium sheets. The vouchers specimens were deposited to Her-
Liu et al., 2011; Sano et al., 2000; Wang, Xie, & Zhao, 2004; Wei et al., barium of Pakistan (ISL) Quaid-i-Azam University Islamabad. 10 species
2013, 2018). belonging to 3 genera were selected for microscopic investigation. The
The microscopic investigation and micromorphology has been samples include 4 species of Athyrium 4 species of Deparia and 2 spe-
widely used in the systematics of ferns and providing many useful cies of Diplazium. A list of selected species of Pakistani Athyriaceae
SHAH ET AL. | 3

FIGURE 1 Distribution of Athyriaceae species in Malakand and taxon sampling localities [Color figure can be viewed at wileyonlinelibrary.
com]

used for micromorphology habitat, vouchers and other information are Azam University Islamabad. For epidermal cell morphology terminology
presented in Table 1. Distribution of Athyriaceae species in Malakand of Van Cotthem (1970) was followed. Stomatal terminology was based
and taxon sampling localities are shown in Figure 4. The nomenclature on the work of Sen and De (1992).
and classification of Athyriaceae used in this work is those adopted by
Smith et al. (2008) and Christenhusz et al. (2011). 2.3 | Scanning electron microscopy (SEM)
For SEM observation dried leaves were used. Sections from both upper
2.2 | Light microscopy (LM)
and lower surface were fixed on double sided tape mounted on tubs.
For light microscopic investigation samples were prepared by methods The specimens were sputter coated with gold palladium and then
of Van Cotthem (1970) with some little modification. Small epidermal observed under SEM (Model JEOL JSM 25910) installed in the Central
pieces of upper and lower surface of leaf was peeled off and cleared in Resource Laboratory (CRL) Department of Physics University of Pesha-
10% HNO3 (Nitric acid) solution in water until the peeled off com- war. Photographs were taken by using Polaroid PN 665 film.
pletely transparent. Then, rinsed with running water for 1 hr and
washed in distilled water, clearing was completed in 40% of NaOCl
2.4 | Quantitative analysis
solution. By this treatment the leaf pieces quickly become transparent,
so that the upper and lower epidermis of a part of the leaf can be stud- The recorded characters were analyzed using different quantitative
ied in one single preparation. The epidermal peel was mounted in glyc- measures i.e. No of epidermal cell, epidermal cell size (L 3 W), stomata
erin gel. To check the consistency of epidermal characters 4 to 5 leaves size (L 3 W), stomatal pore size (L 3 W), trichomes size (L 3 W), sto-
sample were taken from each species and at least 5 to 7 slides in some matal index and stomatal density. Statistical data analysis was per-
plants up to 10 slides were prepared from leaf surface. The epidermal formed in SPSS software (ver 16). The quantitative characters are
peels were examined by Nikon and Meiji (Japan) light microscope. represented as: minimum- maximum (mean) 6 standard error, for
Micrographs were taken by using LEICA- DM-1000 light microscope instance 17 – 30 (23) 6 5. The stomatal index was calculated as
with fitted camera of Meiji infinity DK-5000. Permanent slides were described by Salisbury (1928, 1932). S. I5 E 1S S 3100; S.I 5 Stomatal
deposited to Plant Systematic and Biodiversity Laboratory Quaid-i- index, S5 No of stomata per unit area E5 No of epidermal cell per unit
4 | SHAH ET AL.

T AB LE 1 Selected species of Pakistani Athyriaceae used for micromorphology: habitat, vouchers and other information

Taxon Name Vouchers Habitat Locality Altitude Coordinates Collector name

Athyrium atkinsonii SNS-2558 Growing in floor of Takkay Banda, 30, 00 m 34.7992098 N S.Nasar shah and
Bedd. coniferous forest. Shangla 72.5713758 E Wajid Zaman

Athyrium attenuatum SNS-1023 Growing terrestrial on Toray Banda, 2,700–3,300 m 34.9052178 S.Nasar shah Wajid
(Wall. Ex. C.B. mountain slopes. Shangla N72.6461528 E Zaman
Clark) Tagawa

Athyrium mackinnoni SNS-1123 Terrestrial growing on Dir Chinar Coat 2,700–3,000 m 34.7992118 S.Nasar shah and
(C. Hope) C. Chr forest floor. N72.5513358 E Fazal Ullah

Athyrium wallichianum SNS-1340 Terrestrial growing on Bishi gram, Swat 3,600–4000 m 35.1266398 N S.Nasar shah and
Ching high mountains 72.7459718 E Fazal Ullah
slopes.

Deparia allantodioides SNS-1420 Along streams and Kwanjay Banda, 23,00 m 34.803278 N S.Nasar shah and
(Bedd.) M. Kato forest paths Shangla 71.6062228 E Wajid Zaman

Deparia japonica SNS-7330 Terrestrial on bank Kwanjay Banda, 23,00 m 35.9050178 S.Nasar shah and
(Thunb.) M. Kato along the tract. Shangla N72.6461528 E Wajid Zaman

Deparia macdonellii SNS-1523 Growing along forest Kikore, Shangla 16, 00–2,000 m 34.8181218 S.Nasar shah and
(Bedd.) M.Kato. paths. N71.6957938 E Fazal Ullah

Deparia petersenii SNS-5765 Terrestrial on slope of Kwanjay Banda, 23,00 m 35.0094988 N S.Nasar shah
(Kunze) M. Kato also grow along the Shangla 72.7511588 E
streams.

Diplazium esculentum SNS-1623 Terrestrial along riv- Kwanjay Khwar, 27, 00 m 34.7992098 N S.Nasar shah
(Retz.) Sw. ers and streams. Shangla 72.5713758 E

Diplazium polypo- SNS-1726 Growing along sides Dolo Banda, Shangla 2,700–3,000 m 34.8181218 N S.Nasar shah
dioides Blume of streams and riv- 71.6957938 E
ers.

area. Stomatal density is calculated as described by Ghosh and Davis anticlinal wall occur in A. atkinsonii (Figure 6) A. attenuatum (Figure 6)
(1973). Stomatal density 5 No of stomata per unit area. Stomatal den- A. mackinnoni (Figure 6), A. wallichianum (Figure 7), D. esculentum
sity calculates as the number of stomata per unit leaf area based on the (Figure 7), D. polypodioides (Figure 7). Irregular lobed wall occur in D.
observation of five samples and the area for counting the stomata of petersenii (Figure 7) and D. allantodioides (Figure 6) whereas D. japonica
each field was (0.0940 mm2). A complete statistical analysis of quanti- have broadly lobed anticlinal wall. Under SEM observation the upper
tative characters are presented in Supporting Information Appendix S1. and lower epidermis surface of A. atkinsonii (Figure 7), A.wallichianum

Altogether, 19 quantitative and qualitative characters were investigated (Figure 8), D. petersenii (Figure 8), and D. esculentum (Figure 8) are

and 10 quantitative characters were evaluated for grouping taxa using striate. While the remaining species have smooth epidermis surface. A
number of lobes per cell vary on abaxial and adaxial surfaces between
the clustering analysis method UPGMA (Figure 9).
the species. On adaxial surface highest number of lobes per cell is 9–
13 are reported in D. allantodioides while lowest lobes per cell is 4–7
3 | RESULTS occur in A. atkinsonii. On abaxial surface the highest number of lobes
per cell is 7–11 occurred in D. japonica whereas the lowest is occurred
The qualitative and quantitative characters of leaf micromorphology of in 4–7 in A. attenuatum (Table 2).
Athyriaceae are summarized in Table 2. Selected light and SEM micro- A significant variation is observed in epidermal cell number and
graphs are presented in Figures 628. A taxonomic key based on foliar size on both abaxial and adaxial surface of the investigated species
micromorphological characters was prepared for 10 investigated spe- (Figure 3). The highest number of epidermal cell on adaxial surface is
cies. Morphology of Plant species of family Athyriaceae from research 31 observed in A. attenuatum while lowest cell number counted on this
area (Figure 2). surface is 21 in D. macdonellii . On abaxial surface highest number of
cell were observed is 28 in A. mackinnoni whereas lowest number
counted on this surface is 20 in D. esculentum. Epidermal cell size (L 3
3.1 | Epidermal cell micromorphology
W) varies from minimum 763 25.7 mm (D. polypodioides) to maximum
The epidermal cells shapes of all species as seen under LM were usually 123 3 24.8 mm (Athyrium attenuatum) on adaxial surface. On abaxial
irregular (Figures 628). The anticlinal walls are lobed and irregular, surface minimum cell size observed is 58 326.6 mm (D. polypodioides)
broadly lobed, irregular lobed and sinuous in all the investigated species maximum cell size is 113 3 26.4 mm (D. macdonellii). Epidermal cell size
(Figures 628). The pattern of anticlinal varies between species sinuate on upper surface is slightly longer than those on the lower surface.
T AB LE 2 Qualitative and quantitative characters of foliar epidermal anatomy of 10 species of Athyriaceae
SHAH

Mean
Mean stomatal Trichomes
ET AL.

Shape of Epidermal Epidermal cell Shape of stomatal pore Stomatal size (mm)
epidermal Lobes Wall Epidermis cell mean size (mm) Type of Shape of stomatal Shape of size (mm) size (mm) density Stomatal L 3 W
S. No Plant name cell/AD/AB per cell pattern surface number L3W stomata stomata pore guard cell L 3 W L3W mm2 index and shape

1. Athyrium Irregular 4–6 Sinuous Striate 22 72 323.2 Absent Absent Absent Absent Absent Absent Absent Absent Absent
atkinsonii
Irregular 4–7 Sinuous Striate 20 763 32.6 Polocytic Elliptic Narrow Narrow 353 22.2 333 3 200.8 20.3 Absent
elliptic kidney
shape

2. Athyrium Irregular 5–7 Sinuous Smooth 31 123 3 24.8 Absent Absent Absent Absent Absent Absent Absent Absent Absent
attenuatum
Irregular 4–7 Sinuous Smooth 30 1103 22.3 Polocytic Elliptic Broad Narrow 373 17.1 34 33.2 352.4 22.4 Absent
elliptic kidney
shape

3. Athyrium Irregular 6–9 Sinuous Smooth 25 118 333.6 Absent Absent Absent Absent Absent Absent Absent Absent 803 6
mackinnoni Unicellular
(Nonglandular)
Irregular 5–10 Sinuous Smooth 28 92 326.5 Polocytic Elliptic Narrow Narrow 50 3 30.2 44 32.4 204.2 17.4 Absent
elliptic kidney
shape

4. Athyrium Irregular 6–12 Sinuous Striate 24 803 21.9 Absent Absent Absent Absent Absent Absent Absent Absent
wallichianum
Irregular 8–10 Sinuous Striate 20 813 33.5 Polocytic Wide Narrow Narrow 37 314.1 263 3 120.6 20.4 Absent
Elliptic elliptic kidney
shape

5. Deparia Irregular 9–13 Lobed and Smooth 24 82 339.2 Absent Absent Absent Absent Absent Absent Absent Absent
allantodioides irregular
Irregular 6–13 Lobed and Smooth 20 60 330 Polocytic Elliptic Narrow Narrow 20 318.8 26 3 2.3 144.4 16.6 Absent
irregular elliptic kidney
shape

6. Deparia Irregular 7–10 Broadly Smooth 26 933 23.2 Absent Absent Absent Absent Absent Absent Absent Absent Absent
japonica lobed
Irregular 7–11 Broadly Smooth 28 803 22.4 Anomocytic Elliptic Narrow Narrow 30 310 18 31.8 72.4 12.3 Absent
lobed elliptic kidney
shape

7. Deparia Irregular 6–9 Sinuous Smooth 21 913 34.8 Absent Absent Absent Absent Absent Absent Absent Absent
macdonellii
Irregular 6–10 Sinuous Smooth 22 113 326.4 Polocytic Elliptic Narrow Narrow 34 327.3 243 2.9 135.7 15.4 Absent
elliptic kidney
shape
(Continues)
|
5
 6

T AB LE 2 (Continued)
|

Mean
Mean stomatal Trichomes
Shape of Epidermal Epidermal cell Shape of stomatal pore Stomatal size (mm)
epidermal Lobes Wall Epidermis cell mean size (mm) Type of Shape of stomatal Shape of size (mm) size (mm) density Stomatal L 3 W
S. No Plant name cell/AD/AB per cell pattern surface number L3W stomata stomata pore guard cell L 3 W L3W mm2 index and shape

8. Deparia Irregular 7–10 Irregular Striate 27 893 36.6 Absent Absent Absent Absent Absent Absent Absent Absent
petersenii lobed
Irregular 7–12 Irregular Striate 31 91 332.2 Anomocytic Elliptic Narrow Narrow 353 21.2 31.33 55.2 11.1 Absent
lobed elliptic kidney
shape

9. Diplazium Irregular 6–9 Sinuous Striate 23 105 326.5 Absent Absent Absent Absent Absent Absent Absent Absent
esculentum
Irregular 6–9 Sinuous Striate 20 76 332.4 Anomocytic Elongate Broad Kidney 333 17.3 24.3 2.8 252.9 18.2 Absent
elliptic elliptic shape

10 Diplazium Irregular 5–7 Sinuous Striate 20 763 25.7 Absent Absent Absent Absent Absent Absent Absent Absent
polypodioides
Irregular 6–7 Sinuous Striate 18 58 326.6 Anomocytic Elongate Broad Kidney 263 21.2 323 3 144.2 8.3 Absent
elliptic elliptic shape

Note: AD, Adaxial; AB, Abaxial; L, Length; W, Width.

FIGURE 3
wileyonlinelibrary.com]

3.2 | Stomatal morphology

(j) Diplazium polypodioides [Color figure can be viewed at

Variation of epidermal cell size in 10 species of

atkinsonii (b) Athyrium attenuatum (c) Athyrium mackinnoni (d)
SHAH

F I G U R E 2 Morphology of Plant species Athyriaceae (a) Athyrium

Athyrium wallichianum (e) Deparia allantodioides (f) Deparia japonica

Athyriaceae [Color figure can be viewed at wileyonlinelibrary.com]

(g) Deparia macdonellii (h) Deparia petersenii (i) Diplazium esculentum

Under SEM observation, the shape of stomata is elliptic and wide ellip-
of stomata were observed in studied species, polocytic and anomocytic.
only abaxial surface (Hypostomatic) in all studied species. The two types
ET AL.

tic and in some species elongate elliptic shape is observed. Shape of

Stomatal features are presented in (Table 2). Stomata are present on
SHAH ET AL. | 7

wallichianum had similar identity. In fourth cluster D. esculentum and D.

polypodioides and in fifth D. macdonellii and D. allantodioides found to
be closely related. The sixth cluster consisted of D. japonica and D.
petersenii were found to have similar morphological characters with
definite distance.
Identification keys based on leaf micromorphological characters
for Pakistani Athyriaceae species

1. 1Anticlinal wall lobed and irregular. . . . . . . . . . . . . . . . . . . . . . . . . . . 2

_Anticlinal wall sinuous. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .8
2. 1Lobes per cell 9–13 on adaxial, polocytic stomata on abaxial,
FIGURE 4 Variation in stomatal size of 10 species of Athyriaceae stomatal index onabaxial 144.4. . . . . . . . . . . . . . . . . . . . . . . .Deparia
[Color figure can be viewed at wileyonlinelibrary.com]
allantodioides
stomatal pore in all studied species observed is broad and narrow ellip- - Lobes per cell 5–7 on adaxial, stomata absent on adaxial, sto-
tic. Shape of guard cell is narrow kidney shape in most of the species matal index 352.4. . .3
while in some species the kidney shape guard is also seen. 3. 1 Epidermal cell surface smooth, stomatal size 37 3
A remarkable variation was observed in stomatal size, stomatal 17.1. . . . . .Athyrium attenuatum
density, stomatal pore size and stomatal index (Figure 4 and 5). The - Epidermal cell surface striate, stomatal size 33 3 3. . .
largest stomata were present in A. mackinnoni (50 3 30.2 mm) and . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4
smallest stomata in D. allantodioides (20 318.8 mm). The longest stoma-
4. 1 Lobes per cell on adaxial 4–6, trichomes absent, stomatal den-
tal pores were present in A. mackinnoni (44 32.4 mm) and smallest in D.
sity 200.8. . . . . . . . . . . .
japonica (18 31.8 mm). The stomatal index varies from lowest in D. pol-
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Athyrium
ypodioides (8.3) to highest in A. attenuatum (22.4). The highest stomatal
atkinsonii
density was observed in A. attenuatum (352.4 mm2) whereas the low-
- Lobes per cell on adaxial 6–9, trichomes unicellular on adaxial,
est density were present in D. petersenii (55.2 mm2).
stomatal density 204.2. . . . . . . . . . . . . . . . . . . . . . . . . . .5
5. 1 Shape of stomata elliptic, narrow elliptic stomatal pore.. . .Athy-
3.3 | Trichomes characteristics
rium mackinnoni
Unicellular non glandular trichomes are observed on adaxial surface in - Shape of stomata wide elliptic, stomatal pore narrow elliptic
only one species, A. mackinnoni whereas trichomes are absent in the . . . . . . . . . . . . . . . . . . . . ..6
remaining species. The light and SEM micrograph of trichomes are illus- 6. 1 Epidermal cell wall striate, size of stomata 37 3
trated in Figures 6 and 8. The mean length of trichomes is 80 mm and 14.1. . . . . .Athyrium wallichianum
width is 6 mm. The nonglandular unicellular trichomes have elongated - Epidermal cell sinuous, size of stomata 24 3 27.3. . . . . .
shape tapering at the apex. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ..7
7. 1 Lobes per cell 6–10 on abaxial, stomatal pore narrow ellip-
3.4 | Cluster analysis tic. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Deparia macdonellii
The UPGMA phenogram of the quantitative data provide 6 main clus- - Lobes per cell on abaxial 6–9, stomatal pore broad
ters (Figure 9). In first main cluster A. attenuatum and A. mackinnoni elliptic. . . . . . . . . . . . . . . . . . . . . . . . . . .8
have similar identity. In second and third cluster A. atkinsonii and A. 8. 1 Stomatal pore size 24 3 2.8, stomata density
252.9. . . . . . . . . . . .Diplazium esculentum
- Stomatal pore size 32 3 3, stomata density
144.2. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9

9. 1 Lobes per cell 5–7, shape of epidermal cell sinuous, stomata

size 26 3 21. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Diplazium polypodioides
- Lobes per cell 7–11, shape of epidermal cell broadly lobed, sto-
mata size 30 3 10. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
. . . . . . . . . . . ..10
10. 1 Epidermal cell size 93 3 23.2, stomatal density
72.4. . . . . . . . . . . . . . ..Deparia japonica
FIGURE 5 Variation in stomatal index of 10 species of -Epidermal cell size 89 3 36.6, stomatal density
Athyriaceae [Color figure can be viewed at wileyonlinelibrary.com] 55.2. . . . . . . . . . . . . . .Deparia petersenii
8 | SHAH ET AL.

including Athyriaceae (Bondada, Tu, & Ma, 2006; Chuang & Liu, 2003;
Lee & Oh, 1988; Mali & Yi, 1997; Nayar, 1962; Zhang et al., 2011). A
number of micromorphological characters here can be used in this
study to identify genera and species of Athyriaceae.
Almost all the quantitative and qualitative leaf epidermal charac-
ters were quite variable and inconsistent in studied species. The epider-
mal cell shapes in all studied species are irregular. According to (Van
Cotthem, 1970) irregular epidermal cell have been observed in many
species of Athyriaceae. The irregular epidermal shape of Athyrioid ferns
have been reported in many studies (Wang et al., 2009; Yan-Ping &
Jian-Meng, 2012; Zhang et al., 2011). The present studies reveal some
novel epidermal features in Athyriaceae. For instance, the number of
lobes per cell on both adaxial and abaxial surfaces were counted and
found may be useful for taxonomic purposes up to certain limit in the
family. The highest number of lobes per cell on adaxial surface were
observed is 9–13 in D. allantodioides while lowest lobes per cell is 4–7
occur in A. atkinsonii. The counting of number of lobes of epidermal cell
have been reported is taxonomically useful in certain plant groups
(Rahman, Ahmad, Zafar, Mumtaz, & Shah, 2017).
The anticlinal walls are sinuous in the species of Athyrium and
Diplazium; A. atkinsonii, A. attenuatum, A. mackinnoni, A. wallichianum, D.

FIGURE 6 Light micrograph of shape of cell, pattern of anticlinal

wall, trichomes and stomata type of Athyriaceae 6. Athyrium
atkinsonii 7. Athyrium attenuatum 8–9-10 Athyrium mackinnoni 11–
12 Deparia allantodioides 13–14 Deparia japonica [Color figure can
be viewed at wileyonlinelibrary.com]

4 | DISCUSSION

The plant species studied of Athyriaceae are growing wild in different

regions of Malakand division. The species were selected within differ-
ent eco climatic conditions (Table 1). This article is the first report on
the microscopic investigation of foliar micromorphology of Pakistani
ferns family Athyriaceae. Initially, light microscopic investigations were
conducted to check variability of micromorphological characters on
both adaxial and abaxial surfaces. Further investigation was performed
through scanning electron microcopy for an accurate interpretation of
micromorphological characters and possible relationship of species and
genera within Athyriaceae. The data set obtained here in this study
through microcopy confirmed the importance of foliar epidermal anat-
omy is a source of taxonomic data in Athyriaceae and proved very
rewarding and resulted in exploration of valuable intergeneric and
intraspecific variations in the family. The generic circumscription and
species delimitation in Athyriaceae is still controversial (Liu et al., 2011;
FIGURE 7 Light micrograph of shape of cell, pattern of anticlinal
Wang et al., 2002; Wei et al., 2013). Generally among leaf anatomical
wall and stomata type of Athyriaceae 15–16 Athyrium wallichianum
features in ferns the leaf epidermis provides significant characters for 17 Deparia macdonellii 18–19 Deparia pettersenii 20–21 Diplazium
systematic and taxonomy (Wylie, 1949, Nayar, 1962; Van Cotthem, esculentum 22–23 Diplazium polypodiodes [Color figure can be
1970), and it has been used in identification of many species of ferns viewed at wileyonlinelibrary.com]
SHAH ET AL. | 9

are considered for taxonomic delimitation and each have a particular

terminology. For better understating of ferns stomata type the ontoge-
netic studies are critically important and are necessary to find out dif-
ferent type of mature stomata in ferns Van Cotthem (1970), Thurston
(1969), Sen and De (1992). The stomata are hypostomatic (Stomata are
distributed only on abaxial surface) in all investigated species here.
Hypostomatic leaves are commonly observed in many fern families
(Kramer, 1990; Kramer, Chambers, & Hennipman, 1990; Van Cotthem,
1970). Two types of stomatal complex are studied here in this study.
The stomatal complex of Athyrium species is polocytic which is one of
most widely studied genus of Athyriaceae and polycytic type of stoma-
tal complex is common in this genus (Malik & Bhardwaja, 1993; Mehra
& Soni, 1983; Van Cotthem, 1970). Polocytic stomata were also
observed in two species of Deparia. Anomocytic stomata complex have
been reported as a common type at family level (Mali & Yi, 1997).
Anomocytic stomata were observed in species of D. japonica, D. peter-
senii, D. esculentum, and D. polypodioides. The different stomata types
in the investigated Athyriaceae species can be used up to some extent
to delimit genera.
The SEM observation of stomata shows three types of stomatal
shapes in studied species which may be useful up to certain limit in the
differentiation of species. The shape of stomata is elliptic in A. atkinso-
nii, A. attenuatum, A. mackinnoni, D. allantodioides, D. japonica, D. mac-
donellii, D. petersenii. The wide elliptic stomata were observed in only
one species A. wallichianum. The two species of Diplazium genus have
elongate elliptic stomata. The other stomatal micromorphological char-
FIGURE 8 SEM micrograph of 10 species of Athyriaceae shape of acters of taxonomic importance studied here include shape of stomatal
cell, pattern of anticlinal wall, trichomes and stomata type 24.
pore and guard cell shape. Narrow elliptic stomatal pore were observed
Athyrium atkinsonii 25. Athyrium attenuatum 26–27 Athyrium
in A.atkinsonii, A. mackinnoni A.wallichianum, D. allantodioides, D. japon-
mackinnoni 28. Athyrium wallichianum 29. Deparia allantodioides 30.
Deparia japonica. 31 Deparia macdonellii 32–33 Deparia petersenii ica, D. macdonellii, and D. petersenii. The two species of Diplazium, D.
34 Diplazium esculentum 35 Diplazium polypodiodes esculentum, D. polypodioides have broad elliptic stomatal pore. Narrow
kidney shape guard cells are observed in genera of Athyrium and
esculentum, and D. polypodioides. The anticlinal walls of D. allantodioides
and Deparia petersenii is irregular lobed, however the wall is character-
ized as broadly lobed in D.japonica. The current pattern of anticlinal
wall observed in all investigated species largely agreed with those of
Chinese Athyriaceae (Mali & Yi, 1997). The epidermal anticlinal walls
patterns are suitable micromorphological character for delimitation of
number of species of Athyriaceae. Epidermal and anticlinal wall pattern
provided many important taxonomic delimitation in ferns (Van Cot-
them, 1970). The shape and pattern of epidermal anticlinal walls is con-
sidered as environmental adaptation, as mesophytic species have
usually sinuous walls and xerophytes have straight walls (Gifford,
1989). The epidermis surface seen under SEM is smooth in most spe-
cies but striate epidermis surface were also observed in many species.
Epidermal cell size varies in the investigated species and remarkable
variation is observed in cell size which may have less taxonomic value
and it can be affected by environmental influences. The frequency of
epidermal cell in pert unit mm2 were counted and no of epidermal cells
on both upper and lower surface varies in Pakistani Athyriaceae.
The distribution of stomata and stomata type are considered of
taxonomic value especially at higher taxonomic level (Metcalfe & Chalk, FIGURE 9 UPGMA phenogram for 10 species of Athyriaceae
1950). In ferns different stomatal classification were proposed which based on quantitative characters
10 | SHAH ET AL.

Deparia. Kidney shape guard cells were observed in only two species of descriptions of other Pakistani Athyriaceae species are recommended
Diplazium, D.esculentum, and D. polypodioides. to use these features in a broader taxonomic and evolutionary context.
The stomatal frequency is often determined but it considered hav-
ing less taxonomic value as it influenced by prevailing external environ- ACKNOWLE DGME NTS
mental condition (Metcalfe & Chalk, 1950). Variation in stomatal
We are grateful to staff of Central Resource Labatoray, University of
density and stomatal index have great implication in paleoclimatic
Peshawar for providing the facility of scanning electron microcopy.
reconstruction (Poole, Weyers, Lawson, & Raven, 1996). The density of
stomata varies between the species and found useful taxonomic micro-
ORC ID
morphological characters in studied species. The stomatal index also
Syed Nasar Shah http://orcid.org/0000-0002-9002-6110
distinguished species to some extent in different genera studied here.
Mushtaq Ahmad http://orcid.org/0000-0003-2971-2848
The stomatal index varies from lowest in D. polypodioides (8.3) to high-
Muhammad Zafar http://orcid.org/0000-0003-2002-3907
est in A. attenuatum (22.4). The stomatal size (guard cell size) have
Fazal Ullah http://orcid.org/0000-0001-5856-3189
widely been used for taxonomic segregation in certain plant groups
Wajid Zaman http://orcid.org/0000-0001-6864-2366
(Adedeji & Jewoola, 2008; Prabhakar, 2004; Rudall & Mathew, 1990).
The stomatal size varies between the species and appears to reflect
RE FE RE NC ES
taxonomic segregation in Athyriaceae studied here (Table 2). The other
Adedeji, O., & Jewoola, O. (2008). Importance of leaf epidermal charac-
stomatal epidermal feature of taxonomic importance in this study
ters in the Asteraceae family. Notulae Botanicae Horti Agrobotanici
which was chosen is stomatal pore measurement. The stomatal aper-
Cluj-Napoca, 36(2), 7–16.
ture size have been studied by many scientists using scanning electron
Arana, M., Mynssen, C., & Ponce, M. (2017). Synopsis of Diplazium (Poly-
microcopy and light microcopy techniques (Gardingen, Jeffree, & podiales: Athyriaceae) from Argentina. Phytotaxa, 291(1), 53–65.
Grace, 1989)

Barthlott, W., Wiersch, S., Colić, Z., & Koch, K. (2009). Classification of
The taxonomic importance of trichomes in ferns are well docu- trichome types within species of the water fern Salvinia, and ontog-
mented in botanical literature (Bondada et al., 2006; Chuang & Liu, eny of the egg-beater trichomes. Botany, 87(9), 830–836.

2003; Saunders & Fowler, 1992, 1993; Wagner, 1977). The glandular Bir, S. (1969). The stelar anatomy of Diplazium latifolium Moore. American
Fern Journal, 59(1), 23–26.
trichomes have been reported in some species of Athyriaceae (Fengqin,
Bondada, B., Tu, C., & Ma, L. (2006). Surface structure and anatomical
Qingmei, Shuyu, & Jianxiu, 2006). Multicellular trichomes are common
aspects of Chinese brake fern (Pteris vittata; Pteridaceae). Brittonia,
in genus Diplazium (Arana et al., 2017). However, trichomes are absent 58(3), 217–228.
in most of the species studied here have little diagnostic features in Carlquist, S., & Schneider, E. L. (1997). SEM studies on vessels in ferns.
Pakistani Athyriaceae. Unicellular nonglandular trichomes are observed 4. Astrolepis. American Fern Journal, 87(2), 43–50.
in only one species Athyrium mackinnoni. Cluster analysis (Figure 9) per- Carlquist, S., & Schneider, E. L. (2000). SEM studies on vessels in ferns.
formed on 10 quantitative characters were also useful in demonstrating 16. Pacific tree ferns (Blechnaceae, Cyatheaceae, Dicksoniaceae).
Pacific Science, 54(1), 75–86.
distinct group of taxa. The cluster analysis have been used in many
Carlquist, S., & Schneider, E. L. (2001). Vessels in ferns: structural, eco-
plant groups and has been useful in validating the leaf micro morpho-
logical, and evolutionary significance. American Journal of Botany, 88
logical features as a method of separating the species and genera (1), 1–13.
(Manly & Alberto, 2016) Ching, R.-C. (1964). On some confused genera of the family Athyriaceae.
The present study signifies the foliar epidermal anatomical charac- Acta Phytotaxonomica Sinica, 9(2), 41–84.
ters have importance in the taxonomy of Athyriaceae at generic and Christenhusz, M. J., Zhang, X.-C., & Schneider, H. (2011). A linear
species level. In addition, it also signifies the use of qualitative and sequence of extant families and genera of lycophytes and ferns. Phy-
totaxa, 19(1), 7–54.
quantitative micromorphological characters by using LM and SEM by
Christensen, C. (1906). Index Filicum. Hagerup, Copenhagen. 1938. Filici-
taxonomist to generate taxonomic keys which can be used as confirma-
nae. Manual of Pteridology. Martinus Nijhoff, The Ha.
tory keys in identification of selected taxa and surmount the similarity
Christensen, C. (1917). Index filicum: supplement preliminaire pour les
problem between the genera and species of Athyriaceae.
annees 1913, 1914, 1915, 1916.
Chuang, Y.-Y., & Liu, H.-Y. (2003). Leaf epidermal morphology and its sys-
tematic implications in Taiwan Pteridaceae. Taiwania, 48(1), 60–71.
5 | CONCLUSIONS Chunxiang, L., Shugang, L., Xiaoyan, S., & Qun, Y. (2011). Phylogenetic
positions of the enigmatic asiatic fern genera Diplaziopsis and Rhachi-
This is the first microscopic investigations that describe leaf micromor- dosorus from analyses of four plastid genes. American Fern Journal,
101(3), 142–155.
phology of 10 species of Pakistani Athyriaceae belonging to 3 genera
Copeland, E. B. (1947). Genera Filicum-the genera of ferns. Genera
(Athyrium, Deparia and Diplazium). Leaf micromorphological characters
Filicum-the genera of ferns.
through light microscopy and scanning electron microscopy is
Fengqin, Z., Qingmei, G., Shuyu, T., & Jianxiu, L. (2006). Morphology and
described here is a good source of taxonomic information that can help anatomy of two Athyriaceae genera growing in Shandong. Acta Bota-
the species and genera delimitation. Yet, micromorphological nica Boreali-Occidentalia Sinica, 26(8), 1569–1574.
SHAH ET AL. | 11

Fraser-Jenkins, C. (2006). Woodsiaceae. A revised handbook to the flora Manly, B. F., & Alberto, J. A. N. (2016). Multivariate statistical methods: A
of Ceylon 15(2): 532–576. primer. Chapman and Hall/CRC. 253 p.
Gardingen, P., Jeffree, C., & Grace, J. (1989). Variation in stomatal aper- Mehra, P., & Soni, S. (1983). Stomatal patterns in pteridophytes-an evo-
ture in leaves of Avena fatua L. observed by low-temperature scan- lutionary approach. Proceedings of the Indian National Science Acad-
ning electron microscopy. Plant, Cell and Environment, 12(9), 887– emy, 2, 155–203.
898. Metcalfe, C. R., & Chalk, L. (1950). Anatomy of the dicotyledons. Oxford:
Ghosh, M., & Davis, T. A. (1973). Stomata and trichomes in leaves of Clarendon Press.
young plants. Phytomorphology, 23, 216–229. Nakaike, T., & Malik, S. (1992). A list of pteridophytes collected from
Gifford, E. M. (1989). Morphology and evolution of vascular plants (3rd Pakistan in 1990. Cryptogamic Flora of Pakistan, 1, 261.
ed.). New York: W.H. Freeman and Co. Nakaike, T., & Malik, S. (1993). Cryptogamic flora of Pakistan: National
Hasebe, M., Wolf, P. G., Pryer, K. M., Ueda, K., Ito, M., Sano, R., . . . Mur- Science Museum (Vol. 2, pp. 262).
akami, N. (1995). Fern phylogeny based on rbcL nucleotide sequen- Nayar, B. (1956). Correlation between gametophytic and sporophytic tri-
ces. American Fern Journal, 85(4), 134–181. chomes in ferns. Science & Culture, 21(2), 455–457.
Hayata, B. (1927). On the systematic importance of the stelar system in Nayar, B. (1962). Studies in Pteridaceae. V. Contributions to the mor-
the Filicales, 1. Bot. Mag. Tokyo, 41(492), 697–718. phology of some species of the maidenhair ferns. Journal of the Lin-
Hernandez-Hernandez, V., Terrazas, T., Mehltreter, K., & Angeles, G. nean Society of London, Botany, 58(372), 185–199.
(2012). Studies of petiolar anatomy in ferns: structural diversity and Nester, J. E. (1985). Scanning electron microscopy of antheridia and
systematic significance of the circumendodermal band. Botanical Jour- archegonia of Anemia mexicana Klotzsch. American Journal of Botany,
nal of the Linnean Society, 169(4), 596–610. 72(5), 777–780.
Karafit, S. J., Rothwell, G. W., Stockey, R. A., & Nishida, H. (2006). Evi- Ogura, Y. (1921). On the gaps in the stele of some Polypodiaceae. Shoku-
dence for sympodial vascular architecture in a filicalean fern rhizome: butsugaku Zasshi, 35(415), 113–125.
Dickwhitea allenbyensis gen. et sp. nov.(Athyriaceae). International
Pant, D., & Khare, P. (1971). Epidermal structure of Psilotales and stoma-
Journal of Plant Sciences, 167(3), 721–727.
tal ontogeny of Tmesipteris tannensis Bernh. Annals of Botany, 35(1),
Kato, M. (1972). The vascular structure and its taxonomic significance in 151–157.
the Athyriaceae. Acta Phytotaxonomica et Geobotanica, 25(2), 79–91.
Poole, I., Weyers, J., Lawson, T., & Raven, J. (1996). Variations in stoma-
Kato, M. (1977). Classification of Athyrium and allied genera of Japan. tal density and index: implications for palaeoclimatic reconstructions.
The Botanical Magazine Tokyo, 90(1), 23–40. Plant, Cell and Environment, 19(6), 705–712.
Kato, M. (1979). Taxonomic study of the genus Cornopteris: Athyriaceae. Prabhakar, M. (2004). Structure, delimitation, nomenclature and classification
Acta Phytotaxonomica Et Geobotanica, 30(4–6), 101–118. of stomata. Acta Botanica Sinica-English Edition, 46(2), 242–252.
Kato, M., & Darnaedi, D. (1988). Taxonomic and phytogeographic rela- Rahman, F., Ahmad, M., Zafar, M., Mumtaz, A., & Shah, S. (2017). Taxo-
tionships of Diplazium flavoviride, D. pycnocarpon, and Diplaziopsis. nomic implications of foliar epidermis in Impatiens (Balsaminaceae):
American Fern Journal, 78(3), 77–85. Investigating 12 Pakistani taxa as an example. Plant Biosystems, 151
Kramer, K. (1990). Schizaeaceae. In K. Kubitzki (Ed.), The families and (4), 642–648.
genera of vascular plants, vol. 1. Pteridophytes and gymnosperms. K. Rothfels, C. J., Sundue, M. A., Kuo, L.-Y., Larsson, A., Kato, M., Schuett-
Kramer, P. S. Green (Eds.). (pp. 258–263). Narosa: Springer. pelz, E., & Pryer, K. M. (2012). A revised family-level classification for
Kramer, K., Chambers, T., & Hennipman, E. (1990). Blechnaceae. In Pteri- eupolypod II ferns (Polypodiidae: Polypodiales). Taxon, 61(3), 515–
dophytes and gymnosperms (pp. 60–68). Springer. 533.
Krings, M., Kellogg, D. W., Kerp, H., & Taylor, T. N. (2003). Trichomes of Roux, J. P. (2009). Synopsis of the Lycopodiophyta and Pteridophyta of
the seed fern Blanzyopteris praedentata: implications for plant–insect Africa, Madagascar and neighbouring islands. South African National
interactions in the Late Carboniferous. Botanical Journal of the Lin- Biodiversity Institute. 296 pp.
nean Society, 141(2), 133–149. Rudall, P., & Mathew, B. (1990). Leaf anatomy in Crocus (Iridaceae). Kew
Kuo, L. Y., Ebihara, A., Kato, M., Rouhan, G., Ranker, T. A., Wang, C. N., Bulletin, 45(3), 535–544.
& Chiou, W. L. (2018). Morphological characterization of infra- Salisbury, E. (1928). On the causes and ecological significance of stoma-
generic lineages in Deparia (Athyriaceae: Polypodiales). Cladistics, 34 tal frequency, with special reference to the woodland flora. Philosoph-
(1), 78–92. ical Transactions of the Royal Society of London. Series B, Containing
Large, M. F. (1989). A spore atlas of New Zealand ferns and fern allies: Papers of a Biological Character, 216(431–439), 1–65.
ResearchSpace@ Auckland. Salisbury, E. (1932). The interrelations of soil, climate and organism, and
Lee, C. S., & Oh, Y. C. (1988). A taxonomical study of Korean Pterida- the use of stomatal frequency as an integrating index of the water
ceae on the morphology of leaf epidermis. Korean Journal of Plant relations of the plant. Beih. bot. Zbl, 49(3), 408–420.
Taxonomy, 18(4), 275. Sano, R., Takamiya, M., Kurita, S., Ito, M., & Hasebe, M. (2000). Diplazium
Liu, Y.-C., Chiou, W.-L., & Kato, M. (2011). Molecular phylogeny and tax- subsinuatum and D. tomitaroanum should be moved to Deparia
onomy of the fern genus Anisocampium (Athyriaceae). Taxon, 60(3), according to molecular, morphological, and cytological characters.
824–830. Journal of Plant Research, 113(2), 157–163.
Mali, W., & Yi, R. (1997). Comparative morphological studies on leaf epi- Saunders, R., & Fowler, K. (1992). A morphological taxonomic revision of
derm of athyriaceae. Acta Botanica Boreali-Occidentalia Sinica, 17(5), Azolla Lam. section Rhizosperma (Mey.) Mett.(Azollaceae). Botanical
37–43. Journal of the Linnean Society, 109(3), 329–357.
Malik, S., & Bhardwaja, T. (1993). Structure and histochemistry of sto- Saunders, R. M., & Fowler, K. (1993). The supraspecific taxonomy and
mata and epidermal cells in the three species of genus Athyrium. Phy- evolution of the fern genus Azolla (Azollaceae). Plant Systematics and
tomorphology, 42(2), 35–42. Evolution, 184(3–4), 175–193.
 12 | SHAH ET AL.

Schneider, E., & Carlquist, S. (1998). SEM studies on vessels in ferns. 9. Wang, M., Chen, Z., Zhang, X., Lu, S., & Zhao, G. (2002). Phylogeny of
Dicranopteris (Gleicheniaceae) and vessel patterns in leptosporangiate the Athyriaceae: evidence from chloroplast trnL-F region sequences.
ferns. American Journal of Botany, 85(7), 1028. Acta Phytotaxonomica Sinica, 41(5), 416–426.
Schneider, H., Smith, A. R., Hovenkamp, P., Prado, J., Rouhan, G., Salino, Wang, M., Xie, Y., & Zhao, G. (2004). A revised subdivision of the Athyr-
A., . . . Sessa11, E. B. (2016). A community-derived classification for iaceae. Acta Phytotaxonomica Sinica, 42(6), 524–527.
extant lycophytes and ferns. Journal of Systematics and Evolution, 54 Wang, R., Deng, X., Li, J., Deng, J., & Lu, S.-G. (2009). Leaf Micromor-
(6), 563–603. phology of 12 Species of Polypodiaceae from Guangxi of China and
Sen, U., & De, B. (1992). Structure and ontogeny of stomata in ferns. Its Taxonomic Significance. Journal of Guangxi Normal University (Nat-
Blumea-Biodiversity, Evolution and Biogeography of Plants, 37(1), 239– ural Science Edition), 4, 030.
261. Wang, R., & Lu, S. (2010). Leaf epidermis micromorphology of subfam.
Sen, U., & Hennipman, E. (1981). Structure and ontogeny of stomata in Polypodioideae Nayar (Polypodiaceae) and its taxonomic significance.
Polypodiaceae. Blumea, 27(1), 175–201. Journal of Wuhan Botanical Research, 30(4), 410–416.
Shao, W., Lu, S.-G., & Shang, Q.-C. (2011). Comparative morphology of Wei, R., Ebihara, A., Zhu, Y.-M., Zhao, C.-F., Hennequin, S., & Zhang, X.-
leaf epidermis in the fern genus Phymatopteris (Polypodiaceae). Plant C. (2018). A total-evidence phylogeny of the lady fern genus Athy-
Diversity and Resources, 33(2), 174–182. rium Roth (Athyriaceae) with a new infrageneric classification. Molec-
Smith, A. R., Pryer, K. M., Schuettpelz, E., Korall, P., Schneider, H., & ular Phylogenetics and Evolution, 119(3), 25–36.
Wolf, P. G. (2006). A classification for extant ferns. Taxon, 55(3), Wei, R., Schneider, H., & Zhang, X.-C. (2013). Toward a new circumscrip-
705–731. tion of the twinsorus-fern genus Diplazium (Athyriaceae): A molecular
Smith, A. R., Pryer, K. M., Schuettpelz, E., Korall, P., Schneider, H., & phylogeny with morphological implications and infrageneric taxon-
Wolf, P. G. (2008). Fern classification. Biology and evolution of ferns omy. Taxon, 62(3), 441–457.
and lycophytes (pp. 417–467). Cambridge: Cambridge University Wei, R., & Zhang, X.-C. (2016). Athyrium sessilipinnum: A new lady fern
Press. (Athyriaceae) from southern China. Brittonia, 68(4), 440–447.
Stewart, R. R. (1967). Check list of the plants of Swat State, Northwest Wylie, R. B. (1949). Variations in leaf structure among Adiantum pedatum
Pakistan. plants growing in a rock cavern. American Journal of Botany, 36(3),
Stewart, R. R., Ali, S., & Nasir, E. (1972). An annotated catalogue of the 282–287.
vascular plants of West Pakistan and Kashmir: printed at Fakhri Print Xu, H., & Wang, M.-L. (2009). Study on tracheary elements of three gen-
Press. era in Athyriaceae. Guihaia, 3, 006.
Suseela, M., & Devi, S. (1998). Scanning electron microscopic studies on Yan-Ping, X. C.-D. C., & Jian-Meng, F. (2012). Micromorphological fea-
the associations of vesicular arbuscular mycorrhizae in some Indian tures of epidermis of 20 species ferns from Yunnan. Bulletin of Botan-
ferns. Archives of Phytopathology & Plant Protection, 31(5), 423–428. ical Research, 16(1), 4–20.
Thurston, E. L. (1969). Taxonomic significance of stomatal patterns in the Zhang, H., Ye, J., Liu, Z., & Jing, Y.-H. (2011). The microscopic observa-
ferns. American Fern Journal, 59(2), 68–79. tion on leaf epidermis cells of several ferns in Wu’an National Forest
Tryon, A. F., & Lugardon, B. (2012). Spores of the Pteridophyta: surface, Park. Northern Horticulture, 18(1), 54–67.
wall structure, and diversity based on electron microscope studies. Ziegler, H. (1987). The evolution of stomata. Stomatal Function, 29–57.
Springer Science & Business Media. 648 pp.
Tryon, R., & Tryon, A. F. (1982). Additional taxonomic and nomenclatural
SUP POR TI NG INFOR MATION
notes on ferns. Rhodora, 84(837), 125–130.
Tryon, R. M., & Tryon, A. F. (2012). Ferns and allied plants: with special Additional Supporting Information may be found online in the sup-
reference to tropical America. Springer Science & Business Media. porting information tab for this article.
850 pp.
Van Cotthem, W. (1970). Comparative morphological study of the sto-
mata in the Filicopsida. Bulletin Du Jardin Botanique National De Belgi-
How to cite this article: Shah SN, Ahmad M, Zafar M, et al.
que/Bulletin Van De Nationale Plantentuin Van Belgie, 40(2), 81–88.
Foliar epidermal micromorphology and its taxonomic implica-
Viane, R., & Cotthem, W. (1977). Spore morphology and stomatal characters
of some Kenyan Asplenium species. Plant Biology, 90(1), 219–239. tions in some selected species of Athyriaceae. Microsc Res Tech.

Wagner, W. H. (1977). Systematic implications of the Psilotaceae. Britto- 2018;00:1–12. https://doi.org/10.1002/jemt.23055

nia, 29(1), 54–63.

View publication stats