APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Mar. 1999, p. 879–885 Vol. 65, No.

3
0099-2240/99/$04.0010
Copyright © 1999, American Society for Microbiology. All Rights Reserved.

Microbial Degradation of Paintings

ORIO CIFERRI*
Dipartimento di Genetica e Microbiologia and Centro Interdipartimentale di Studi e di Ricerche per la Conservazione
dei Beni Culturali, Università degli Studi di Pavia, 27100 Pavia, Italy

“. . .l’alliance possible et désiderable de la Science et de l’Art. . .,” should settle once and forever the arguments about the possi-
Louis Pasteur, when he was nominated to the first chair in ble role of microorganisms in the degradation of our cultural
physical chemistry at the Ecole des Beaux Arts in Paris. heritage. The conditions that led to the microbial bloom on the
Lascaux Cave paintings probably represent an extreme case,
INTRODUCTION but it may be argued convincingly that even less harsh envi-
ronmental stresses than those that occurred in the less than 20
In 1940 four young men discovered the Lascaux Cave in the years since the opening of the Lascaux Cave may cause irre-
Dordogne region of France. The cave contained an impressive versible aesthetic and structural damage to almost any type of
display of prehistoric art: the main cavern and several galleries art work.
connected to it were decorated with engraved, drawn, and This minireview focuses on the colonization of art works by
painted figures of animals. The approximately 600 paintings, microorganisms and its effects. Its scope will be limited to
done with mineral pigments mixed with animal fat in various paintings, both on canvas and panel, as well as on walls. Thus,
shades of yellow, red, brown, and black, were dated to the late other art works, such as those in stone, wood, paper, and
Aurignacian period (15,000 to 13,000 B.C.). With few excep- masonry, as well as those in more esoteric materials, such as
tions, the paintings, some as long as 5 m, represented different leather, parchment, glass, and metal, will not be considered.
animals (some imaginary), and their quality was such that the For a more comprehensive treatment of the role of microor-
cave was designated by some the Sistine Chapel of the Paleo- ganisms in the degradation of our cultural heritage, the reader
lithic. In 1948 Lascaux Cave was opened to visitors, but in 1963 should refer to the reviews already published (2, 6, 12, 13, 20,
it was closed indefinitely to the public. Closing was imposed 29, 30, 36, 46, 55, 56). The treatment of the subject will not be
after the discovery of a green patina (from which comes the exhaustive but will focus on aspects that, in the writer’s opin-
term maladie verte, or green disease) covering the painted ion, appear to be most interesting. At the end, a few ideas on
portions (34). Quite unexpectedly, although other algae to- how, again in the writer’s opinion, the research in this field
gether with cyanobacteria, bacteria, and fungi were isolated in might proceed will be expressed.
different parts of the cave, the green patina was composed
exclusively of the unicellular alga Bracteacoccus minor (order
Chlorococcales). The influx of workers and visitors brought THE SUBSTRATE
into the cave considerable amounts of soil and of the organic
Paintings, whether easel or mural, contain a wide range of
compounds present in people’s breath and sweat and increased
organic and inorganic constituents and provide different eco-
the concentration of carbon dioxide to almost pathological
logical niches that may be exploited by a large variety of mi-
levels. The lighting system, installed in the cave and operating
crobial species. Many of the components of paintings are bio-
almost continuously, created the conditions for a massive
degradable, and so are the additives (glues, emulsifiers,
growth of photosynthetic organisms. Extensive analysis of the
thickeners, etc.) that facilitate drawing or application of paint
composition of, and the variations in, the microbial population
layers or enhance the aesthetic quality of the finished product.
of the painted areas as well as of the unpainted rocks and the
In easel paintings, the support material (the cellulose of
surrounding environment led to the conclusion that the pop-
paper, canvas, and wood and the proteins of parchment, silk,
ulation of Bracteacoccus minor, responsible for the maladie
and wool) may be easily degraded by microorganisms, as may
verte, also increased when the cave was closed to the public and
the materials (animal or plant glues) used to “size” the support
kept in continuous darkness for long periods. Indeed, after 3
and to prepare a ground layer. Paintings on paper or silk are
months of total darkness and closure to the public, algal pro-
laid, in general, directly on the support, since a ground or
liferation on painted areas was found to have increased by 1
underlay is lacking, but the pigments are kept in emulsion with
order of magnitude (35). Thus, it was concluded that the alga
organic binders. Thus, besides the organic nature of the sup-
could grow even under heterotrophic conditions by utilizing
port, easel paintings contain organic molecules that many mi-
the organic molecules brought in the cave by visitors or result-
croorganisms may utilize for growth, such as sugars, gums, and
ing from the degradation of biological residues. It was postu-
other polysaccharides, proteins, linseed and other oils, waxes,
lated that, before discovery and opening of the cave, the com-
etc., but also less chemically defined mixtures of biomolecules
munity of heterotrophic microorganisms, bacteria and fungi,
such as egg yolk, bile, and even urine. (A list, certainly not
present in the cave had mineralized all organic molecules
exhaustive, of the organic components that may be present on
present, so that heterotrophic growth of the alga was pre-
paintings can be found in references 15 and 55).
vented, as was autotrophic growth as a result of the absence of
Mural paintings rely on techniques and materials differing
light.
from those utilized in easel paintings. Essentially, pigments are
The Lascaux Cave is perhaps the most emblematic example
suspended in water or oil, often in the presence of binders such
of the damage that microorganisms may cause to art work and
as casein and milk, and applied on the damp lime plaster. The
calcium carbonate formed on contact with air consolidates the
* Mailing address: Dip. Genetica e Microbiologia, Via Ferrata 1, pigments. Thus, by and large, frescoes contain mainly inor-
27100 Pavia, Italy. Phone: 39 382 505576 or 39 382 505577. Fax: 39 382 ganic components and the microbial flora that colonize these
528496. E-mail: [email protected]. substrates may, at least in the first steps, differ from that

879
880 MINIREVIEW APPL. ENVIRON. MICROBIOL.

present on easel paintings. For both types of paintings the in this investigation, determination of acetylene reduction in
spectrum of compounds that may be present is further in- situ demonstrated that nitrogen fixation occurred, albeit at a
creased by those that are added at later times during retouch- reduced rate, in the microbial biofilm covering the frescoes. In
ing, restoration, or relining or when a fresco is detached and addition, cyanobacteria and algae can provide an important
transferred to a canvas or a board. In one case at least, exten- source of organic material on which heterotrophic bacteria and
sive fungal colonization was reported even with frescoes that, fungi may thrive, thus causing further aesthetic and structural
after cleaning and consolidation, were removed from walls and damage to the paintings. Finally, cyanobacteria and algae may
transferred to a fiberglass support (42). Finally, dirt, soot, and colonize the mortar, bricks, or stone supporting frescoes. In-
other environmental contaminants, accumulating on the deed, these organisms have been reported to contribute to the
painted surface, may represent another not insignificant source weathering process of masonry (31).
of nutrients.
Given the wide range of organic and inorganic molecules THE FLORA
that are present in both types of paintings, many different types
of microorganisms may grow on such substrates provided that With a few exceptions, characterization of the microbial
favorable environmental conditions (humidity, temperature, flora present on frescoes or easel paintings has been limited to
light, and, to a lesser extent, pH) are met. It sounds almost selected groups of microorganisms rather than to all types of
tautological to state that, besides the chemical composition of microorganisms that might be present on a given substrate.
an art work, the environment conditions the development of a Thus, in general, surveys have often been limited to fungi (1, 8,
microbial flora, as it is quite obvious that a specific microbial 10, 14, 17, 22–26, 37, 47, 50, 57), bacteria (7, 18, 32, 33, 44, 45),
flora will develop, for instance, on a fresco on the facade of a or cyanobacteria and eukaryotic algae (9, 16, 21, 40, 58). In a
church where it receives a considerable amount of light and a few cases, more comprehensive analyses aiming to determine
different flora will develop on a similar fresco inside the same all, or the majority of, the biota present on a painting have
building in which light is very reduced. Likewise, if tempera- been reported (27, 41). This comprehensive data may provide
ture, moisture, and light are not controlled, the microbial com- the foundation for ascertaining the existence of associations or
munities of two paintings produced with exactly the same ma- successions among the components of a microbial flora. Re-
terials will differ considerably if one painting is kept in the cently, a method of identifying microorganisms by sequencing
northern latitudes and the other is kept in the tropics. It may a portion of the DNA coding for the 16S rRNA has been used
be added that high levels of humidity, temperature, and light, with cultures of bacteria isolated from frescoes (7, 44) and even
as may be found, for instance, in warmer climates, may shorten with DNA samples extracted directly from a fresco (44, 45).
the, one could say, life span of a painting by exacerbating the This technique, extensively employed in macromolecular ecol-
damages caused by air pollution, biological attack, and natural ogy to identify, without culturing, members of microbial com-
aging. munities, will certainly lengthen the list of microorganisms
Growth of microorganisms on paintings may cause aesthetic present on any given substrate by permitting, for instance, the
and structural damage. As aesthetic damage one must consider identification of species that are present at very low cell con-
pigment discoloration, stains, and formation of a biofilm on the centrations and of those that cannot be cultured in the labo-
painted surface, whereas as structural damage one must con- ratory. However, it will not determine if the DNA derives from
sider cracking and disintegration of paint layers, formation of living or dead microorganisms and, more importantly, it will
paint blisters, and degradation of support polymers or of glues not allow us to distinguish between microorganisms responsi-
and binders resulting in detachment of the paint layer from the ble for the observed damage (one could call them the para-
support. Of course, the two types of damage are strongly sites) and those that do not contribute to it (the saprophytes).
linked, and in the long run, structural damage profoundly af- Similar limitations will greatly reduce the usefulness of other
fects the aesthetic quality of a painting. Conversely, aesthetic molecular biological techniques, such as fluorescence in situ
damage may precede serious injuries to the materials. For hybridization, that permit identification of microorganisms
instance, in fungal colonization of mural paintings, Saiz-Jime- without their isolation and culture.
nez and Samson (47) have shown that, at the beginning, growth Perusal of the lists of taxa isolated shows that the most
of fungi on a mural’s surface caused only aesthetic damage common soil inhabitants, both fungi (species of Penicillium,
since there was little or no alteration of the painted surface. Aspergillus, Cladosporium, Chaetomium, and Alternaria) and
Later on, fungal growth in depth occurred. Hyphae penetrated bacteria (species of Pseudomonas, Arthrobacter, and Streptomy-
the painted layer, degrading some of its components (especial- ces), are present in many of the samples analyzed. However,
ly glues and binders), which resulted in a decrease in the wide quantitative variations are evident. For instance, from a
cohesion of the painted layers, thus giving rise to exfoliations, fresco in St. Damian’s Monastery in Assisi, Italy, more than 33
cracking, and loss of the paint. To these damages one should different species of fungi belonging to at least 17 genera were
add those inflicted by metabolites, often acidic in nature, and isolated (approximately 25% of all isolates were not identified)
by extracellular enzymes excreted by microorganisms. These (22). On the other hand, from a mural in Canterbury Cathedral
compounds may modify the colors as well as the stability of the only one fungal species, Beauveria alba (Engyodontium album),
painted layer and of the substrate. was repeatedly isolated (26) and, similarly, on damaged fres-
Similarly, cyanobacteria and algae growing on paintings ex- coes in an Italian church only one species of Cladosporium was
posed to light, such as frescoes on the facades of buildings, may found (37).
cause considerable damage. Besides the aesthetic damage Gettens and coworkers were among the first to point out, in
caused by a green, black, brown, or yellow algal patina covering 1941, that paintings could be “defaced or destroyed by the
the painted portions, these organisms may cause weathering of growth of those small, parasitical plants commonly called
the surface layers, accelerating detachment of portions of the ‘mold’ or ‘mildew’” (15). Further, in laboratory experiments,
painted layer as well as the underlying plaster (40). The pres- they demonstrated that treatment with fungicides could arrest
ence in a number of Italian frescoes of species of nitrogen- or prevent microbe-induced damage to paintings. About 20
fixing Nostoc indicates that cyanobacteria may colonize fres- years later, Tonolo and Giacobini (59) confirmed that micro-
coes in which combined nitrogen may be absent (58). Indeed, organisms could damage works of art by providing examples of
VOL. 65, 1999 MINIREVIEW 881

frescoes disfigured by growth of eukaryotic algae (members of the isolates from Ghirlandaio’s fresco but was not isolated
Chlorophyceae), bacteria (Sarcina lutea or Streptomyces spp.), from Botticelli’s fresco. Such differences in two frescoes
or fungi (species of Penicillium, Aspergillus, Cephalosporium, painted at the same time (1480) in the same building, presum-
and some Dematiaceae). The authors reported that these or- ably with similar or identical materials, and restored and
ganisms could cause changes to the paintings’ surfaces through cleaned at the same time appear rather striking. In laboratory
staining, discoloration, or formation of patinas and efflores- experiments, 19 species of the fungi isolated from the two
cence. In addition, they showed that many such organisms, frescoes were tested for the capacity to grow on the materials
especially the fungi, could grow between the paint layers and used for restoration (calcium caseinate, animal glue, and ma-
the ground, causing a swelling of the paint film that could lead sonite, used as a support panel). Although qualitative differ-
to detachment of portions of the painted layer and disaggre- ences were observed, essentially all the fungal species isolated
gation of the underlying ground. This in turn could promote from both frescoes grew quite well on calcium caseinate, to a
separation of the painted surface from the ground or of the lesser extent on masonite, and to an even lesser extent on
ground from the masonry on which the fresco was laid. animal glue. The only exception was provided by the two spe-
After these pioneering papers, Gargani (14) and Tiano and cies of Cladosporium, which, although being among the most
Gargani (57) published a detailed investigation of the micro- frequent isolates from the two frescoes, did not grow well on
bial floras of art works, mostly frescoes. Their work was greatly any of these materials. In the opinion of the investigators, this
stimulated by the finding that, after the flooding of Florence in genus is one of the most commonly isolated from frescoes
November 1966, a great number of paintings, both mural and because it is resistant to variations in external factors (temper-
easel, were severely damaged and that the damage could be at ature, humidity, etc.). However, as the two tested species of
least in part associated with the growth of microorganisms. Cladosporium did not grow on casein, masonite, or animal
Using a technique of dermatologic mycology, they determined glue, the investigators assumed that this genus did not contrib-
that direct microscopic examination of the microbial structures ute significantly to the degradation of paintings. This assump-
adhering to transparent cellulose tape pressed on the painted tion is in contrast to the opinion of other scientists who con-
surface revealed the presence of fungal elements, such as hy- sider Cladosporium one of the major biological agents, if not
phae, typical of most filamentous fungi. However, species iden- the most significant agent, responsible for fresco degradation
tification and determination of the microbial load were possi- (2, 19, 37, 47). In conclusion, the differences observed in the
ble only when cultures on different media were made with fungal colonizations of the two frescoes are not easily ex-
small fragments of the painted surface or cotton swabs brushed plained. Assuming that no great differences exist in the mate-
on such a surface. The analyses were essentially limited to the rials used when the two frescoes were painted (but this cannot
fungal population and demonstrated that clear differences ex- be proved), the only possible explanation is that the locations
isted between the numbers of species isolated from art works of the two frescoes in the church are such that they affect
and those isolated from the environment in which the art work differentially the fungal colonizations of the two murals. One
was located. For instance, from the surface of a fresco by Beato can argue that the positions of the frescoes relative to openings
Angelico in St. Mark’s Convent in Florence, Italy, 17 different (windows or doors), sources of moisture and heat, and other
species of hyphomycetes encompassing 10 genera were iso- factors may be responsible for the differences in the fungal
lated whereas from the environment 9 species (six genera) colonizations. In an extensive investigation of the fungal colo-
were isolated. These data could be taken as an indication of the nizations of frescoes in eight different Moldavian monasteries,
presence of a fungal flora specifically developing on the paint- Ionita (25) isolated 26 different species of fungi from stains
ing and differing, at least in part, from that present in the appearing on the frescoes, from areas of efflorescence, and
environment. However, sampling at different intervals from zones in which the painted layer was fissured and portions
(months) revealed significant differences in the compositions were breaking away from the support. No apparent recogniz-
of the flora of the painted surface whereas there was little able pattern in the fungal distribution could be observed. For
variation from sampling to sampling in the flora of the envi- instance, from three areas with stains of the same color,
ronment. Similar wide variations in the species isolated from present on the same portion of a fresco, different fungi were
different periods were reported in the analyses of the micro- isolated. In addition, the same fungal species was isolated from
bial, essentially fungal, flora present on wall paintings in the spots of different colors as well as from fissured fragments of
Buddhist shrines of Ajanta in India (1). Of 40 different species the frescoes that were apparently not stained. Further, As-
of fungi isolated from the wall paintings on three different pergillus niger, one of the most ubiquitous fungal contaminants,
visits, only 11 species were always present and more than 50% was isolated in only one case.
were isolated only once. Such variations in the fungal floras present in samples taken
Two 15th-century murals in the Ognissanti church in Flo- at different times, or in frescoes of the same age and in the
rence, restored in 1969 after the flood of 1966, were cleaned same location, were often observed and do not allow us to
and treated with nystatin in the late 1970s but, in 1985, showed establish conclusively that the fungi present on a painted sur-
the appearance of greenish-brown–to–black spots on the face, even when they are absent from the environment, are
painted surface (49). Isolation of fungal species from such responsible for the damage observed on the paintings. Further,
areas demonstrated the presence of 15 different species from no attempt has been made to identify the species responsible
the samples taken from Botticelli’s fresco and a similar number for the damage, both aesthetic and structural, and the species
(13) from that by Ghirlandaio. However, striking differences in that are just saprophytes living on the painted surface may be
the types of species were evident: the most abundant fungi on growing at the expense of other microorganisms colonizing the
the fresco by Botticelli were two species of Penicillium and frescoes. However, the idea that fungi may be the primary
Cladosporium cladosporiodes, whereas the most common fungi microbiological agents responsible for degradation of art
in the fresco by Ghirlandaio were Aspergillus versicolor and works is so entrenched that often antibacterial agents are
Cladosporium sphaerospermum. Even more striking was the added routinely to the media used for the isolation of the
finding that the two penicillia most abundant on the Botticelli microbial contaminants presumed responsible for the degra-
fresco were undetected on the fresco by Ghirlandaio, as was dation of art works (22, 26).
the case with Aspergillus versicolor, which accounted for 74% of With frescoes located underground, such as those in crypts,
882 MINIREVIEW APPL. ENVIRON. MICROBIOL.

tombs, and grottoes, it has been reported that the predominant are limited to one group of microorganisms (fungi, bacteria, or
species and, possibly, the first colonizers are members of the algae) and rarely include all the microorganisms present.
order Actinomycetales, most of which are in the genus Strepto- Nevertheless, in a few cases attempts have been made to
myces and a few of which are in the genus Nocardia (18). Over present a more comprehensive analysis of the different micro-
200 strains of actinomycetes were isolated from 13 frescoes in bial groups present, to unravel the chemical modifications
different Italian hypogean sites. In some of these, cell concen- brought about by the microbial colonization, and to determine
trations reached up to 1 million cells/gram of sample. Of the the succession of the microbial colonizers. For instance, Saiz-
200 isolates, 46 were identified as members of 19 different Jimenez and Samson (47) have analyzed the microbial flora of
species of Streptomyces and 5 were identified as members of a large fresco painted in the late 1920s in an old Spanish
the genus Nocardia. According to the researchers, colonization monastery. Two types of aesthetic damage were observed,
by actinomycetes begins as soon as the sites are opened and the white efflorescence and green-to-black stains. From both types
frescoes are excavated, becoming quite evident only 2 months of alterations Cladosporium sphaerospermum was the fungus
after excavation and exposure to air. In a short period, other most frequently isolated (approximately 75 to 88% of all iso-
microorganisms (bacteria, fungi, and algae) become associated lates from the two types of lesions), followed by Engyodontium
with the predominant population of actinomycetes. When the album (slightly more than 10% of all isolates from both efflo-
hypogean rooms of the Domus Aurea in Rome were opened to rescent and stained areas). However, the fungi were consid-
visitors in 1951, very rapidly green crusts appeared on the ered secondary colonizers of the fresco. The first microorgan-
frescoes in lighted areas; their development was so rapid that, isms colonizing the fresco were supposed to be sulfur-cycling
in 1981, illumination had to be discontinued (21). A study of bacteria (48), well known to play an important role in stone
the microbial community composing such crusts showed a pre- and masonry deterioration (12). The decay of the fresco was
dominance of cyanobacteria (two species of Lyngbya, accom- thought to have begun around the 1970s, coincident with the
panied by unidentified bacteria) and chlorophytes (species of establishment in the vicinity of the monastery of a series of
Chlorella, Pseudococcomyxa, and Pseudopleurococcus) (4). The industrial plants that emitted into the atmosphere considerable
composition of the algal population associated with the dam- amounts of pollutants, especially sulfur dioxide. The sulfuric
age was studied for 4 years. During this period, the two species acid produced from sulfur dioxide dissolved the calcium car-
of Lyngbya were by far the predominant ones. The chloro- bonate of the fresco, leading, eventually, to the production of
phytes Pseudococcomyxa simples and Pseudopleurococcus print- a precipitate of dihydrous calcium sulfate (gypsum). Gypsum
zii were always present but at much lower cellular concentra- deposition resulted in the formation of white crystal aggregates
tions. These findings were confirmed in laboratory experiments responsible for the efflorescence observed on the fresco. Mi-
in which samples of the microbial mats from the frescoes were crobiological analyses showed that the efflorescence contained
grown under fluorescent or incandescent light at two different up to 65,000 sulfur-oxidizing and 200 sulfur-reducing bacteria
light intensities (5). In these experiments too the two Lyngbya per gram. In the investigator’s view, the sulfur-utilizing bacte-
species appeared to be the predominant ones. According to the ria were the first colonizers of the fresco. Death and lysis of
investigators, the presence of thick sheaths of these cyanobac- these bacteria provided the organic substrates necessary for
teria not only favored their adhesion to the painted surface but the growth of heterotrophic bacteria and fungi. Growth of the
provided also the substrates for the establishment of a popu- latter was considered responsible for the colored stains present
lation of heterotrophic bacteria (3). on the fresco’s surface as well as the mechanical damage ob-
In conclusion, although an impressive number of publica- served, such as the detachment of portions of the painted layer.
tions report that from damaged paintings it is possible to iso- Thus, environmental pollutants, especially sulfur dioxide and
late a wide range of microorganisms, with very few exceptions related compounds, caused direct damage to the fresco but
no attempts have been made to distinguish between microor- also provided the substrates that promoted growth of aerobic
ganisms responsible for the deterioration and those that play and anaerobic sulfate-cycling bacteria. These, in turn, supplied
no role, direct or indirect, in the process leading to a painting’s the organic nutrients that allowed the establishment of a com-
defacement. munity of scavenger bacteria and fungi that further contributed
to the degradation of the fresco.
A somewhat similar sequence of events was postulated by
MECHANISM OF AGGRESSION AND MICROBIAL Karpovich-Tate and Rebrikova to occur on frescoes and ma-
SUCCESSION sonry in a Russian cathedral (27). According to these research-
ers, even in the presence of organic substrates such as compo-
Presenting a unified scheme for determining the mechanism nents of fresco and plaster, the first colonizers were the
of microbial damage of painted surfaces is rather difficult. Such autotrophic, nitrifying bacteria found on many different types
difficulty resides in the fact that the chemical compositions of of stone and masonry and considered responsible for the bio-
paintings vary considerably, and at times, they are even impos- logically induced corrosion of stone and other building mate-
sible to ascertain. Although historical records and, more sig- rials (11). These bacteria oxidized to nitrate the ammonia
nificantly, chemical analyses may indicate with sufficient accu- present in the atmosphere and thus promoted growth of het-
racy the pigments that have been used in older art works, it is erotrophic microorganisms (bacteria as well as fungi that were
less easy to determine which components were used for sizing present in concentrations up to 106 cells/g of material) that also
the ground, emulsifying the pigments, protecting the finished utilized the cellular components of the first colonizers. Accord-
painted surfaces, etc. As already mentioned, another difficulty ing to the researchers, support for this conclusion was given by
lies in the fact that most of the published reports are essentially the finding that most of the heterotrophs that were present on
catalogues of the microorganisms isolated from painted sur- frescoes were capable of hydrolyzing bacterial and yeast cell
faces, especially from the areas in which visual inspection has walls. A somewhat similar analysis of the bacteria present on
revealed aesthetic damage due to changes in the colors of frescoes in northern Moldavia monasteries gave different re-
paints and appearance of stains, variations in the structure of sults. Over 90 bacterial strains were isolated, all of which were
the painted layer, etc. Further, as already noticed, quite often heterotrophs and most of which were in the genera Bacillus,
the lists of microorganisms isolated from a damaged painting Arthrobacter, Micrococcus, Sarcina, and Pseudomonas (32). The
VOL. 65, 1999 MINIREVIEW 883

presence of bacteria was constantly demonstrated in the sam- the only fungal species present on the panels. Such results
ples collected from portions of the fresco disfigured by a whit- confirmed those of an earlier report on the succession of fungi
ish, powdery layer, whereas the absence of bacteria was dem- on this type of paint, namely, that initially species of Aspergil-
onstrated in samples from apparently undamaged portions of lus, followed by species of Alternaria, and, eventually, Aureo-
the fresco. In a courageous attempt to verify Koch’s postulates, basidium pullulans were found. The last represented 80% of
control experiments demonstrated that when pure cultures of the climax community, the remaining 20% being represented
many of these bacteria were transferred to sterile cotton wool by Alternaria spp. (60). The possibility that Aureobasidium pul-
wads and these were applied to and kept on undamaged por- lulans grew at the expense of the polysaccharides of the
tions of the same fresco for 3 to 4 weeks, almost half of the 40 Pseudomonas capsules and the other bacterial species coloniz-
isolates tested produced stains similar to those observed in the ing the panels was investigated (39). Although dead bacterial
damaged portions. From the artificially produced areas of cells adhering to the paint layer did stimulate growth of the
staining the researchers reisolated the bacterial species used fungus, further experiments provided evidence that bacterial
for inoculation. Bacteria, especially of the genus Arthrobacter, colonization of the painted surface had chemically modified
were reported to be among the first colonizers of murals in a some of the components of the paint, rendering them utilizable
medieval church in Rostov, Russia (41), and to be responsible by the fungus (38). Indeed, a previous report showed that
for oxidation of the lead present in pigments, resulting in the Aureobasidium pullulans was unable to utilize hydroxyethylcel-
production of brown-black spots of lead oxides. Indeed, when lulose, a component of the paint, for growth but that it utilized
samples taken from the damaged areas of the murals or bac- this compound pretreated with cells of Pseudomonas or even
teria isolated from such samples were incubated in mineral with a cellulase produced by the bacterium (51).
media in the presence of lead-containing pigments such as Somewhat different conclusions were reached in our inves-
white lead, lead ocher, or red lead, good microbial growth tigations with samples of painted canvases (mock paintings)
together with the formation of a brown precipitate composed prepared with traditional materials by following the standard
of lead dioxide was observed. The fact that no brown precip- recipes used for paintings (52). Essentially, mock paintings
itate was formed in uninoculated media or in those inoculated consisted of a linen canvas, sized with animal glue in water and
with an unidentified fungus isolated from the same area of the with a ground of chalk and animal glue. A paint film of lead
fresco gave strong, presumptive evidence that the bacteria white in linseed oil was laid on the smoothed-out ground. The
present in the damaged fresco were responsible for the oxida- main soil microorganisms, fungi and bacteria, growing on the
tion of divalent lead to tetravalent lead oxide and hence to the mock painting were identified. Bacillus pumilus was the bacte-
appearance of dark-brown–to–black spots on areas in which rial species present at the highest cell concentration, by far,
lead-containing pigments were used. In addition, other labo- and Aspergillus niger and Penicillium chrysogenum were the
ratory experiments indicated that black spots of lead sulfide fungal species present at the highest cell concentrations. Re-
could be produced on the frescoes from the reaction between construction experiments showed that pure cultures of the
the lead oxide of pigments and the hydrogen sulfide produced main bacterial species, including Bacillus pumilus, essentially
by other bacterial species present in the samples. did not grow when they were incubated with mock paintings. In
In conclusion, the few reports in which the mechanism of this type of experiment only the viable counts of the fungi
microbial colonization of frescoes has been investigated indi- Aspergillus niger and Penicillium chrysogenum increased in the
cate that bacteria may be the first colonizers. However, the first period of incubation. However, the presence of Aspergillus
majority of reports are limited to analyses of the fungal flora niger stimulated growth and survival on mock paintings of
isolated from the substrates and make no attempt to establish Bacillus pumilus and the stimulatory effect of the fungus was
whether these microorganisms are the first to colonize the abolished by the addition of cycloheximide, an inhibitor of
substrates. protein synthesis and growth in eukaryotes but not in pro-
With easel paintings, experiments performed on wood pan- karyotes. These findings, indicating that growing fungal cells
els coated with a white acrylic latex and exposed to soil in an are necessary to promote growth and survival of Bacillus pumi-
environmental cabinet or in the field led to the isolation of lus, could be explained by the fact that Aspergillus niger was
members of 7 bacterial genera and 15 fungal genera, with no found to possess cellulolytic and proteolytic activities, activities
great difference between the numbers of genera isolated in the that were not identified in Bacillus pumilus and the other
laboratory (20 isolated) and in the field samples (23 isolated) bacteria, all of which were gram positive, isolated from mock
(38). The time course (over 2 weeks) of the colonization by the paintings exposed to soil. Thus, it was postulated that the
different genera showed that some organisms, termed transient fungus stimulated growth and survival of the bacteria by sup-
species (Acremonium, Penicillium, and Helmintosporium spp.) plying the latter with the products of the hydrolysis of macro-
were present only during certain periods but that not only were molecules, such as cellulose and proteins, present on the paint-
other organisms, termed permanent species (Alternaria and ings. This conclusion was strengthened by the finding that the
Pseudomonas spp.) present in all samples but also their num- most abundant bacterial species, mostly gram-negative organ-
bers often increased throughout the period of exposure. Mem- isms, isolated from a severely degraded 16th century fresco
bers of the genera Alcaligenes, Bacillus, Flavobacterium, and that had been transferred in the 19th century to a canvas
Pseudomonas represented the most frequent bacterial species support hydrolyzed cellulose and casein, grew to a certain
present at all times. Whereas the population of most bacterial extent, and survived for a longer period of time on mock
species remained constant or increased only slightly during the paintings than did the bacteria isolated from soil (53). Unlike
duration of the experiments, that of Pseudomonas increased Bacillus pumilus, growth and survival on mock paintings of the
linearly with the time of incubation (during 12 weeks of incu- bacteria isolated from the fresco were not stimulated by the
bation, the number of colonies of Pseudomonas spp. per square presence of Aspergillus niger. The differences between our data
centimeter increased by more than 1 order of magnitude). and those of O’Neil’s and Schmitt’s could be easily explained
With fungi, only colony numbers of Aureobasidium (Pullularia) by the differences in the materials used (acrylic paint on wood
pullulans, considered by some the main biological agent of in one case, oil paint on cloth in the other) and indicate that
paint deterioration (28, 43), increased steadily with the time of the succession of the different microbial taxa colonizing works
incubation so that, after 12 weeks, this species was essentially of art depends also on the chemical nature of the substrate.
884 MINIREVIEW APPL. ENVIRON. MICROBIOL.

Indeed, work under way in my laboratory has demonstrated ing of the substrates (supports, pigments, binders, glues, etc.)
the existence of differences in the microbial colonizations of that make up an art work, how the substrate is modified by the
mock paintings when different pigment binders (oil or distem- microbial colonization, and how these modifications lead to
per) were used or when the same type of painting was relined the establishment of different microbial communities. Simi-
with different glues (unpublished data). larly, one should try to evaluate in the laboratory how the
That the chemical nature of the substrate conditions the microbial population varies when the environmental condi-
capacity of microorganisms to colonize different art works was tions change (a painting on the exterior of a building will
further demonstrated by the finding that silk (composed of the undergo colonization by microorganisms different from those
proteins fibroin and sericin but often of fibroin only) is easily colonizing a similar painting located inside the same building).
colonized and degraded by bacteria (especially species of Finally, one must evaluate how aging, which may be simulated
Pseudomonas and Arthrobacter) but that it is hardly attacked by in the laboratory, may bring about variations in the chemical
fungi (54). However, if the textile was artificially aged in the structures of many components of works of art (from the sup-
laboratory by exposure to the light of a xenon lamp or to heat, port polymers to the different binders and glues) and how these
treatments that result in a chemical modification of the pro- chemical variations may influence the colonization by different
tein, then it became susceptible also to fungal attack (unpub- microbial taxa. From such research it will be possible to learn
lished data). how to monitor and evaluate the onset and the rate of micro-
Thus, one should take into consideration how the microbial bial colonization and the changes in the microbial population
flora colonizing an art work varies according to the chemical as a function of the substrate composition and environmental
composition of such a work. Further, the biochemical reactions conditions and, eventually, how to proceed for disinfestation.
catalyzed by the different microbial species may vary with the Finally, these data will be useful in indicating the most suit-
different makeup of the substrate and also when external fac- able materials to be used, including those for restoration and
tors, including age, alter the chemical structures of some of the relining. We expect the life spans of works of art to be on the
components of the substrate. The number of variables to be order of centuries if not millennia. It is inconceivable that we
taken into consideration becomes almost unlimited, presenting will find compounds that will ensure protection from microbial
a difficult but not unsurmountable challenge, since reliable attack for periods of such lengths. If, for any reason, control of
information can be gathered in laboratory experiments per- humidity, temperature, and light, as occurs in museums, is not
formed with standardized models. When the microbiologist is possible, then protection of objects of artistic or historical
confronted with a request to investigate the presence (and interest rests only on the intrinsic components of such objects
role) of microorganisms on a defaced art work, he or she is that can render them refractory to microbial colonization.
called to do so with a substrate on which, quite often, microbial
colonization has taken place for years. The microbial flora that ACKNOWLEDGMENTS
he or she will find is probably the result of successive coloni- The research performed in my laboratory was supported by grants
zations by different groups of microorganisms. Such variations from Progetto Finalizzato Beni Culturali of the Italian Research
are the result of modifications of the chemical composition of Council (C.N.R.).
the substrate, to which the microorganisms themselves may I am grateful to many colleagues for supplying reprints of papers and
have contributed in part. Thus, the investigator will have only to Maria Gravagna for constant and generous help in the bibliographic
a snapshot of the state of the artifact at that precise moment search for and in the preparation of the manuscript.
and not a time-elapsed picture of the development of the
REFERENCES
microbial communities that may have existed during the life
1. Agrawal, O. P., S. Dhawan, K. L. Garg, F. Shaheen, N. Pathak, and A. Misra.
span of the art work. In addition, he or she will be called to give 1988. Study of biodeterioration of the Ajanta wall paintings. Int. Biodeterior.
an answer in a short period and to provide in great haste the 24:121–129.
information necessary for corrective interventions. A microbi- 2. Agrawal, O. P., S. Dhawan, and K. L. Garg. 1989. Microbial deterioration of
ologist should be asked to characterize the microbial flora paintings—a review, p. 1–51. Intach Conservation Centre, Lucknow, India.
3. Albertano, P., and M. Grilli Caiola. 1989. A hypogean algal association.
present on a work of art when it appears to be still in its Braun-Blanquetia 3:287–292.
pristine condition and well before any alteration becomes ev- 4. Albertano, P., L. Luongo, and M. Grilli Caiola. 1991. Observations on cell
ident. By determining which microorganisms are present at structure of micro-organisms of an epilithic photoprophic community com-
time zero, he or she will be able to make a reasonable assump- peting for light. Nova Hedwigia 53:369–381.
5. Albertano, P., L. Luongo, and M. Grilli Caiola. 1991. Influence of different
tion about how the microbial colonization will develop. In this lights on mixed cultures of microalgae from ancient frescoes. Int. Biodete-
way, the microbiologist will be able to suggest the nature and rior. 27:27–38.
the mode of treatment that will stop microbial colonization 6. Allsopp, D., and K. J. Seal. 1986. Biodeterioration of refined and processed
before damage becomes visible and irreversible. materials, p. 51–53. In Introduction to biodeterioration. Edward Arnold,
London, United Kingdom.
It seems to me that the time is now ripe to acknowledge that 7. Altenburger, P., P. Kampfer, A. Makristathis, W. Lubitz, and H.-J. Busse.
studies of the microbial colonization of art works should go 1996. Classification of bacteria isolated from a medieval wall painting. J. Bio-
beyond the descriptive stage, that is, cataloguing which organ- technol. 47:39–52.
isms are found on which substrate. It is undeniable that this 8. Arai, H. 1984. Microbiological studies on the conservation of mural paintings
in tumuli, p. 117–124. In Y. Emoto and S. Miura (ed.), International Sym-
type of information is important to establish which organisms, posium on the Conservation and Restoration of Cultural Property. Tokyo
or which types of organisms (bacteria, algae, fungi, etc.), col- National Research Institute of Cultural Property, Tokyo, Japan.
onize a given art work, since this information is necessary for 9. Ariño, X., M. Hernandez-Marine, and C. Saiz-Jimenez. 1996. Ctenocladus
any disinfestation treatment. However, I think that we are now circinnatus (Chlorophyta) in stuccos from archaelogical sites of southern
Spain. Phycologia 35:183–189.
in the position to begin to study and understand the mecha- 10. Bianchi, A., M. A. Favali, N. Barbieri, and M. Bassi. 1980. The use of
nisms underlying the microbiological attack. In other words, fungicides on mold-covered frescoes in S. Eusebio in Pavia. Int. Biodeterior.
we should try to set up standardized laboratory models using Bull. 16:45–51.
the most common types of support as well as the most com- 11. Bock, E., W. Sand, M. Meincke, B. Wolters, B. Ahlers, C. Meyer, and F.
Sameluck. 1988. Biologically induced corrosion of natural stones—strong
monly employed ingredients. These models will allow us to contamination of monuments with nitrifying organisms, p. 436–440. In D. R.
establish, under controlled conditions, which species colonize a Houghton, R. N. Smith, and H. O. W. Eggins (ed.), Biodeterioration, vol. 7.
given substrate, how the microbial flora will change on chang- Elsevier Applied Science, New York, N.Y.
VOL. 65, 1999 MINIREVIEW 885

12. Bock, E., and W. Sand. 1993. The microbiology of masonry biodeterioration. cellulosic textiles: a review. Int. Biodeterior. 28:209–226.
J. Appl. Bacteriol. 74:503–514. 37. Nugari, M. P., M. Realini, and A. Roccardi. 1993. Contamination of mural
13. Bravery, A. F. 1988. Biodeterioration of paint—a state-of-the-art comment, paintings by indoor airborne fungal spores. Aerobiologia 9:131–139.
p. 466–485. In D. R. Houghton, R. N. Smith and H. O. W. Eggins (ed.), 38. O’Neill, T. B. 1986. Succession and interrelationships of microorganisms on
Biodeterioration, vol. 7. Elsevier Applied Science, New York, N.Y. painted surfaces. J. Coatings Technol. 58:51–56.
14. Gargani, G. 1968. Fungus contamination of Florence art—masterpieces be- 39. O’Neill, T. B. 1988. Succession and interrelationships of microorganisms on
fore and after the 1966 disaster, p. 252–257. In A. H. Walters and J. J. painted surfaces. Int. Biodeterior. 24:373–379.
Elphick (ed.), Biodeterioration of materials. Elsevier P.C., Amsterdam, The 40. Ortega-Calvo, J. J., M. Hernandez-Marine, and C. Saiz-Jimenez. 1993. Cya-
Netherlands. nobacteria and algae on historic buildings and monuments, p. 173–203. In
15. Gettens, R. J., M. Pease, and G. I. Stout. 1941. The problem of mold growth K. L. Garg, N. Garg, and K. G. Mukerji (ed.), Recent advances in biodete-
in paintings. Techn. Stud. Fine Arts 9:127–143. rioration and biodegradation, vol. I. Naya Prokash, Calcutta, India.
16. Giacobini, C., C. Andreoli, G. Casadoro, B. Fumanti, P. Lanzara, and N. 41. Petushkova, J. P., and N. N. Lyalikova. 1986. Microbiological degradation of
Rascio. 1979. Una caratteristica alterazione delle murature e degli intonaci, lead-containing pigments in mural paintings. Stud. Conserv. 31:65–69.
p. 289–299. In Atti del 3° Congresso Internazionale sul Deterioramento e la 42. Realini, M., N. Barbieri, and G. Sala. 1988. Fungal growth on frescoes in the
Conservazione della Pietra, Venice, Italy. University of Padua, Padua, Italy basilica of S. Vincenzo in Galliano, p. 340–343. In C. A. C. Sequeira and
17. Giacobini, C., and M. Firpi. 1981. Problemi di microbiologia nei dipinti su A. K. Tiller (ed.), Microbial corrosion, vol. 1. Elsevier Applied Science,
tela, p. 203–211. In Atti del Convenzione sul Restauro delle Opere d’Arte. London, United Kingdom.
Opificio delle Pietre Dure e Laboratorio di Restauro di Firenze, Polistampa, 43. Reynolds, E. S. 1950. Pullularia as a cause of deterioration of paint and
Florence, Italy. plastic surfaces in South Florida. Mycologia 42:432–448.
18. Giacobini, C., M. A. De Cicco, I. Tiglie, and G. Accardo. 1988. Actinomycetes 44. Rolleke, S., G. Muyzer, C. Wawer, G. Wanner, and W. Lubitz. 1996. Iden-
and biodeterioration in the field of fine art, p. 418–423. In D. R. Houghton, tification of bacteria in a biodegraded wall painting by denaturing gradient
R. N. Smith, and H. O. W. Eggins (ed.), Biodeterioration, vol. 7. Elsevier gel electrophoresis of PCR-amplified gene fragments coding for 16S rRNA.
Applied Science, New York, N.Y. Appl. Environ. Microbiol. 62:2059–2065.
19. Giacobini, C., M. Pedica, and M. Spinucci. 1991. 31. Problems and future 45. Rolleke, S., A. Witte, G. Wanner, and W. Lubitz. 1998. Medieval wall paint-
projects on the study of biodeterioration: mural and canvas paintings, p. ings—a habitat for archaea: identification of archaea by denaturing gradient
275–286. In Proceedings of the 1st International Conference on the Biode- gel electrophoresis (DGGE) of PCR-amplified gene fragments coding for
terioration of Cultural Property. Macmillan India Limited, New Delhi, India. 16S rRNA in a medieval wall painting. Int. Biodeterior. and Biodegrad.
20. Griffin, P. S., N. Indictor, and R. J. Koestler. 1991. The biodeterioration of 41:85–92.
stone: a review of deterioration mechanisms, conservation case histories, and 46. Ross, R. T. 1963. Microbiology of paint films. Adv. Appl. Microbiol. 5:217–
treatment. Int. Biodeterior. 28:187–207. 234.
21. Grilli Caiola, M., C. Forni, and P. Albertano. 1987. Characterization of the 47. Saiz-Jimenez, C., and R. A. Samson. 1981. Biodegradacion de obras de arte.
algal flora growing on ancient Roman frescoes. Phycologia 26:387–390. Hongos implicados en la degradacion de los frescos del monasterio de la
22. Guglielminetti, M., C. De Giuli Morghen, A. Radaelli, F. Bistoni, G. Car-
Rabida (Huelva). Bot. Macaronesica 8–9:255–264.
ruba, G. Spera, and G. Caretta. 1994. Mycological and ultrastructural studies
48. Saiz-Jimenez, C., and R. A. Samson. 1981. Microorganisms and environmen-
to evaluate biodeterioration of mural paintings. Detection of fungi and mites
tal pollution as deteriorating agents of the frescoes of the monastery “Santa
in frescos of the monastery of St. Damian in Assisi. Int. Biodeterior. Biode-
Maria de la Rabida”, Huelva, Spain. Presented at the 6th Triennial Meeting
grad. 34:269–283.
of the International Council of Museums Committee for Conservation, Ot-
23. Inoue, M., and M. Koyano. 1991. Fungal contamination of oil paintings in
tawa, Canada.
Japan. Int. Biodeterior. 28:23–35.
49. Sampò, S., and A. M. Luppi Mosca. 1989. A study of the fungi occurring on
24. Ionita, I. 1971. Contributions to the study of the biodeterioration of the
15th century frescoes in Florence, Italy. Int. Biodeterior. 25:343–353.
works of art and of historic monuments. II. Species of fungi isolated from oil
50. Savulescu, A., and I. Ionita. 1971. Contributions to the study of the biode-
and tempera paintings. Rev. Roum. Biol. Ser. Bot. 16:377–381.
terioration of the works of art and historic monuments. I. Species of fungi
25. Ionita, I. 1973. Contributions to the study of the biodeterioration of the
works of art and historical monuments. IV. Fungi involved in the deterio- isolated from frescoes. Rev. Roum. Biol. Ser. Bot. 16:201–206.
ration of mural paintings from the monasteries of Moldavia. Rev. Roum. 51. Schmitt, J. A. 1974. The microecology of mold growth. J. Paint Technol.
Biol. Ser. Bot. 18:179–189. 46:59–64.
26. Jeffries, P. 1986. Growth of Beauvaria alba on mural paintings in Canterbury 52. Seves, A. M., S. Sora, and O. Ciferri. 1996. The microbial colonization of oil
Cathedral. Int. Biodeterior. 22:11–13. paintings. A laboratory investigation. Int. Biodeterior. Biodegrad. 37:215–
27. Karpovich-Tate, N., and N. L. Rebrikova. 1990. Microbial communities on 224.
damaged frescoes and building materials in the Cathedral of the Nativity of 53. Seves, A. M., S. Sora, and O. Ciferri. 1996. La flora batterica, p. 229–233. In
the Virgin in the Pafnutii-Borovskii Monastery, Russia. Int. Biodeterior. “L’affresco di Sant’ Agata al Monte di Pavia: ricerche ed analisi per il
27:281–296. restauro.” Memorie dell’Istituto Lombardo—Accademia di Scienze e Let-
28. Klens, P. F., and J. R. Lang. 1956. Microbiological factors in paint preser- tere, vol. XL, no. 4. Instituto Lombardo di Scienze e Lettere, Milan, Italy.
vation. J. Oil Colour Chemists’ Assoc. 38:887–899. 54. Seves, A. M., M. Romanò, T. Maifreni, S. Sora, and O. Ciferri. 1999. The
29. Koestler, R. J., T. Warscheid, and F. Nieto. 1997. Biodeterioration: risk microbial degradation of silk: a laboratory investigation. Int. Biodeterior.
factors and their management, p. 25–36. In N. S. Baer and R. Snethlage Biodegrad. 42:203–211.
(ed.), Saving our architectural heritage: the conservation of historic stone 55. Strzelczyk, A. B. 1981. Paintings and sculptures, p. 203–234. In A. H. Rose
structures. J. Wiley and Sons, London, United Kingdom. (ed.), Microbial deterioration. Academic Press, London, United Kingdom.
30. Kowalik, R. 1980. Microbiodeterioration of library materials. Part 2. Micro- 56. Tiano, P. 1993. Biodeterioration of stone monuments: a critical review, p.
biodecomposition of basic organic library materials. Restaurator 4:135–219. 301–321. In K. L. Garg, N. Garg, and K. G. Mukerji (ed.), Recent advances
31. Krumbein, W. E., and C. Lange. 1978. Decay of plaster paintings and wall in biodeterioration and biodegradation, vol. I. Naya Prokash, Calcutta, India.
material of the interior of buildings via microbial activity, p. 687–697. In 57. Tiano, P., and G. Gargani. 1981. Controlli microbiologici su alcuni affreschi
Environmental biogeochemistry and geomicrobiology. Proceedings of the fiorentini, p. 341–358. In Atti del Convegno sul Restauro delle Opere d’Arte.
3rd International Symposium on Environmental Biogeochemistry. Ann Ar- Opificio delle pietre dure e laboratori di restauro di Firenze. Polistampa,
bor Science Publishers, Inc., Ann Arbor, Mich. Florence, Italy.
32. Lazar, I. 1971. Investigations on the presence and role of bacteria in dete- 58. Tomaselli, L., M. C. Margheri, and G. Florenzano. 1979. Indagine speri-
riorated zones of Cozia Monastery painting. Rev. Roum. Biol. Ser. Bot. mentale sul ruolo dei cianobatteri e delle microalghe nel deterioramento di
16:437–444. moumenti ed affreschi, p. 313–325. In Atti del 3° Congresso Internazionale
33. Lazar, I., and L. Dumitru. 1973. Bacteria and their role in the deterioration sul Deterioramento e la Conservazione della Pietra, Venice, Italy. University
of frescoes of the complex of monasteries from northern Moldavia. Rev. of Padua, Padua, Italy.
Roum. Biol. Ser. Bot. 18:191–197. 59. Tonolo, A., and C. Giacobini. 1961. Microbiological changes on frescoes, p.
34. Lefèvre, M. 1974. La “maladie verte” de Lascaux. Stud. Conserv. 19:126–156. 62–64. In G. Thompson (ed.), Recent advances in conservation. Butter-
35. Lefèvre, M., G. Laporte, and J. Bauer. 1964. Sur les microorganismes enva- worths, London, United Kingdom.
hissant les peintures rupestres de la grotte préhistorique de Lascaux. C. R. 60. Winters, H., I. R. Isquith, and M. Goll. 1975. A study of the ecological
Acad. Sci. 258:5116–5118. succession in biodeterioration of a vinyl acrylic paint film. Dev. Ind. Micro-
36. Montegut, D., N. Indictor, and R. J. Koestler. 1991. Fungal deterioration of biol. 17:167–171.