Aureobasidium
EJ van Nieuwenhuijzen, CBS-KNAW Fungal Biodiversity Centre, Utrecht, The Netherlands
Ó 2014 Elsevier Ltd. All rights reserved.
This article is a revision of the previous edition article by T. Roukas, volume 1, pp 109–112, Ó 1999, Elsevier Ltd.
Characteristics of the Genus
Habitat
Aureobasidium is a worldwide-distributed fungus mainly
present on diverse organic and inorganic outdoor materials,
such as phylloplanes, soil, wood, marble, and water. Examples
of isolation from extreme environments include the outer
space, salterns, and a damaged nuclear reactor. Indoors, it can
be found in house dust and in wet environments like bathroom
walls. In food factories, it sometimes can be isolated from
painted surfaces. Aureobasidium has been isolated from a wide
range of foods but only rarely has been designated as a cause of
spoilage. For wine grapes, it is known that Aureobasidium is
dominant on healthy grape berries, but is overgrown by others,
when the grape skin clearly is damaged. It is commonly isolated
from the surface of fresh fruits and vegetables – for example,
apple, pear, blueberries, peaches, strawberries, cabbage, lettuce,
and broccoli. Old records describe its presence in shrimp, grain,
flour, oats, and nuts. Many years ago, it was found in some
frozen foods and was involved in the black spot spoilage of
long-term-stored beef. Occasionally, it is found in raw milk,
cheese, and smoked beef.
Taxonomy
The genus Aureobasidium belongs to Ascomycota, order
Dothideales, family Dothideaceae. Officially described species
are Aureobasidium iranium and Aureobasidium pullulans, in which
the last species is divided into the subspecies pullulans, mela-
nogenum, subglaciale, and namibae. Their taxonomy is based on
morphology and phylogenetic studies and represents only
a small geographic area compared with the global presence of
this fungus. For example, analysis of isolates from Thailand and
Iceland resulted in 14 different phylogenetic clades.
Cultural Characteristics
Colonies on malt extract agar (MEA) at 25
C expand rapidly
attaining at least 20 mm diameter after 7 days and appear
smooth. The shine, slimy, or matt appearance depends on the
(sub)species. The same applies to the border texture varying
from smooth to raveling. Aerial mycelium sometimes formed
scanty, thinly floccose.
Isolates are typically off-white to pale pink or black on solid
media, whereas some tropical isolates have been described as
‘color variants’ with pigments of pink, brown, yellow, or
purple. The color of a single isolate can vary due to the type of
solid media. Although colonies on MEA and potato dextrose
agar can show dark- or bright-colored pigmentation, colonies
on Dichloran Glycerol (DG18) are mostly white.
On MEA, most strains start as pinkish white colony. They
turn dark brown or black after some time varying from a day to
a few weeks. Colonies can turn dark along the whole colony,
Encyclopedia of Food Microbiology, Volume 1 http://dx.doi.org/10.1016/B978-0-12-384730-0.00017-3
only in the center or partially in sectors (see Figure 1). Dark-
ening of cultures is due to the formation of chlamydospores,
which contain the pigment melanin.
Micromorphology
Aureobasidium can grow as budding yeast or as mycelia in dark
or hyaline appearance, depending on environmental condi-
tions and (sub)species. On MEA the following microscopic
characteristics can be seen in the officially described species:
smooth hyaline thin-walled hyphae, maximum 13 mm wide.
The occurrence of hyaline hyphae converting to dark-brown
hyphae depends on the (sub)species varying from sometimes
locally in older cultures to rather soon in all cultures. These
thick-walled hyphae may act as a chlamydospore chain or fall
apart into separate dark cells commonly called chlamydospores
or chlamydoconidia. Conidiogenous cells are undifferentiated,
intercalary, or terminal on hyaline hyphae. Conidia produced
synchronously in dense groups from small denticles, and in
most strains are formed percurrently from single butts on short
lateral branches. Hyaline conidia are one-celled, smooth,
ellipsoidal, and variable in shape and size. Budding of hyaline
and dark-brown conidia frequently is noticed. Chlamydoco-
nidia are mostly 1-2-celled, being bigger than hyaline conidia.
Endoconidia occasionally have been seen.
Of the tropical isolates of A. pullulans, young colonies on
MEA showed polymorphic forms with blastospores, swollen
cells, and pseudohyphae, with older colonies showing hyphae
and chlamydospores.
Physiology
Aureobasidium chlamydospores can survive low temperatures
and reduced water activity present in frozen foods. For micro-
bial growth, the temperature has to rise above 4
C. Optimum
growth is 20–25
C and a maximum of 37
C is reported. The
fungus cannot grow in highly salted food but might survive the
salinity because it is reported as halotolerant. It is an aerobic
fungus, needing oxygen for growth. Chlamydospores were
found in outer space, which demonstrates that they can survive
without oxygen. High doses of gamma irradiation are needed
to sterilize Aureobasidium in food products. Assumptions are
made that the melanin present in the chlamydospores are
responsible for the high tolerance against the gamma irradia-
tion and especially ultraviolet (UV) irradiation.
Industrial Application of Aureobasidium
Aureobasidium pullulans is an industrially important microor-
ganism especially because of its capability to produce pullulan
(poly-a-1,6-maltotriose). Pullulan is a commercially exploited
biodegradable extracellular polysaccharide used in coatings
and wrappings potential and as a food ingredient. Also the
105
106 Aureobasidium

Figure 1 Aureobasidium pullulans. (a)–(b), (d)–(e): Colonies grown at 25
C for 7 days on MEA and DG18. (c)–(f): Colonies grown at 25
C for 14 days on
MEA. (g): Conidiogenous cells producing blastoconidia synchronously. (h): Hyphae. (i)–(j): Conidia. Scale bar ¼ 10 mm.
 Aureobasidium 107

Table 1 Food additives and medical supplements produced by Aureobasidium

Product Strains Application

Pullulan Most strains isolated Coating and wrapping agent and as a food ingredient
b-Glucan A. Pullulans NP1221 Supplement to enhance immune system and lower blood pressure
Erythritol Aureobasidium sp. SN-124A An artificial sweetener used as food ingredient
Gluconic acid A. pullulans AHU 9190, DSM 7085 Flavoring and leavening agent, reducer of fat absorption
L-Malic acid A. pullulans FERM-P2760 Acidulant
Poly(b-L-malic acid) Most strains isolated Drug carrier
Fructoligosaccharides Strains producing fructosyl transferase Nondigestible sweeteners; they have applications in health foods

Table 2 Enzymes produced by Aureobasidium

Temperature
Enzyme optimum (
C) pH optimum Food industry application

Alkaline protease 45, 48–52 9.0 Cleaves peptide bonds of proteins; detergent, food processing
Acidophilic endo-1,4-b-xylanase 50 2 Hydrolyses xylan, clarification of fruit pulp and juices,
production of wine
b-Xylosidase 80 3.5 Hydrolyses xylan, clarification of fruit pulp and juices,
production of wine
Glucoamylase 50–60 4–4.5, 5.75 Starch saccharification, detergent, bread and baking processing,
production of high-fructose syrup
a-Amylase 55 5.0 Starch saccharification, detergent, bread and baking processing,
production of high-fructose syrup
Lipase 35 8.5 Breaks down milk fat, flavoring cheeses, food processing
Endo-b-1,4-mannanase Not published Reduces viscosity of coffee extracts
a-Galactosidase Not published Reduces viscosity of coffee extracts
b-Galactosidase 45 6.8 Hydrolyses lactose in whey or milk
b-Mannosidase Not published Reducing viscosity of coffee extracts
Pectinase (unspecified) 12 3.5 Wine production
Protopectinase Not published 4.5 Maceration of fruit pulps and for clarification of juices and wines
Polygalacturonase 50–60 4.6–5.5 Maceration of fruit pulps and for clarification of juices and wines
Endopolygalacturonase 37 3.8 Maceration of fruit pulps and for clarification of juices and wines
Pectin lyase 40 5–7.5 Maceration of fruit pulps and for clarification of juices and wines
Pectin methylesterase Not published Maceration of fruit pulps and for clarification of juices and wines
b-fructofuranosidase I 50–55 5.5 Production of prebiotics, sweetener
Fructosyl transferase 65 4.4 Production of prebiotics, sweetener
b-Glucosidase 75 4 Removal of bitterness from citrus fruit juices,
wine production, diary processing
Endoglucanase 60 4.5 Food fermentation
Exoglucanase Not published 5.5 Preparation of dehydrated vegetables and food products
Xylitol dehydrogenase 25 10–10.5 Oxidizes xylitol to D-xylulose
Laccase 25–35 4.5–6.4 Bioremediation, beverage (wine, fruit juice and beer) processing,
ascorbic acid determination, sugar beet pectin gelation, baking
L-Fucose dehydrogenase 30 9.5 Converts L-fucose to L-fuconic acid
Phosphatase Not published
Polyamine oxidase Not published Biological inactivation of polyamines component
of clinical diagnostic assay kits
L-Rhamnose dehydrogenase Not published 9.0 Hydrolyses L-rhamnose
Sucrase 35 4.5 Hydrolyses sucrose

polysaccharide b-glucan produced by Aureobasidium is transferases. Consequently, it has become an important

commercially available. It is identified as an effective substance
to improve animal health condition. Other metabolites
produced by A. pullulans that are used as medical supplement
or additives in food are presented in Table 1.
Different strains of A. pullulans isolated from different
environment can produce many important enzymes, such as
amylase, protease, lipase, cellulose, xylanase, mannose, and
organism in applied microbiology. In Table 2, the different
enzymes are listed together with the corresponding optimum
reaction conditions and a description of the possible applica-
tion in the food industry. Of the multiple studied Aureobasi-
dium enzymes, a general description has been made:
Proteases in general have been shown to have many
applications; however, studies on the production and
108 Aureobasidium
characterization of proteases derived from Aureobasidium are
rather new. Marine-derived strains as well as a terrestrial-
derived strain, secrete extracellular proteases.
Aureobasidium pullulans has shown to be a xylan-degrading
fungus. The enzymatic degradation of xylan to xylose requires
the catalysis of both endoxylanase and b-xylosidase. Xylanases
have applications in paper, fermentation, and food industries,
as well as in waste treatment.
Amylases have applications in bread and baking industry,
starch liquefaction and saccharification, textile desizing, paper
industry, detergent industry, and food and pharmaceutical
industries. Amylases hydrolyze starch molecule into glucose,
maltose, and dextrin. They can be classified into a-amylase, b-
amylase, and glucoamylase. Aureobasidium pullulans is known to
produce a-amylase a glucoamylase.
A few studies exist on the extracellular lipases produced by
Aureobasidium isolates from marine environment. Lipases
catalyze a wide range of reactions, including hydrolysis of
lipids, interesterification, alcoholysis, acidolysis, esterification,
and aminolysis. They are the enzyme of choice for potential
applications in the food, detergent, pharmaceutical, leather,
textile, cosmetic, and paper industries.
In the 1990s, different strains were found to produce
mannanases. Mannanases are useful in many fields, including
the coffee industry. Mannan is distributed widely in nature as
part of the hemicelluloses (polyoses) fraction in plant cell
walls. One strain A. pullulans was mentioned to produce all
enzymes required for complete degradation of galactomannan
and galactoglucomannan: endo-b-1,4-mannanase is secreted
into the culture fluid, b-mannosidase is strictly intracellular,
and a-galactosidase and b-glucosidase are found both extra-
cellular and intracellular.
Aureobasidium also produces pectinases. These enzymes are
widely used for fruit pulping and for the clarification of juices
and wines. Protopectinases, polygalacturonases, pectin lyase,
and pectin esterases are among the studied enzymes produced
by Aureobasidium.
The enzymes b-fructofuranosidases and fructosyltransferase
produced by A. pullulans have been used to produce fructo-
oligosaccharides (FOS). These FOS are a class of prebiotics,
used as food material. Its taste is close to that of conventional
sweeteners such as sucrose.
Cellulases are enzymes that degrade crystalline cellulose to
glucose. Cellulases have diverse applications also in the food
industry. Three types of cellulases, endoglucanases, cellobio-
hydrolases, and b-glucosidases, are considered to be needed to
degrade cellulose to glucose. It has been observed that most of
the cultures of A. pullulans usually have failed to show any
cellulolytic activity. Mostly strains that produce b-glucosidase
were found. Some isolates of A. pullulans of tropical
origin produced CMCase (endoglucanase) and a-cellulase
(exoglucanase).
Laccase production from A. pullulans was studied in the
1970s but up-to-date reports are also available. Laccases are
well known as a component of fungal enzyme systems of lignin
degradation. Potentially, they can be used in several areas, such
as textile, paper, and food industries.
Aureobasidium is mentioned repeatedly as producer of single
cell proteins that are used as protein supplement in human
foods or animal feeds. Although dried cells of microorganism
generally have an application potential due to high nutritive
values, there is doubt on the replacement of conventional
protein sources because of the slower digestibility and uncer-
tainty on possible allergic reactions.
Aureobasidium as a natural living system can be used in
various applications. It is commercially developed as a micro-
bial pest control agent protecting the blossom of pome fruit
against the plant pathogen Erwinia amylovora. The mode of
action against E. amylovora is explained by an increased resis-
tance of host plants toward the fire-blight pathogen by
competition for nutrients and space. Another high-potential
application is the use of Aureobasidium as a black biofilm pro-
tecting wood against wood rot or UV degradation. Wood that is
treated with linseed oil can naturally form a completely
covered black film on the wood surface during outside exhi-
bition of the treated wood. Studies revealed that chlamydo-
spores are responsible for the black color of the film.
Aureobasidium is mentioned in reports to be an indicator of
environmental perturbations generated by chemicals or other
biological organisms on leaf surfaces.
Method of Detection
Aureobasidium can be isolated from food by homogenization of
solid food in peptone water or swapping the surface of the food
with sterile cotton wool and shaking it in water. After plating
and several days of incubating on MEA, pure subcultures can be
made. Identification of the pure cultures can be done by the
classical method of morphology analysis or by molecular
methods.
The morphology of Aureobasidium can be used as a diag-
nostic feature. The synchronous conidia production from
young expanding hyphae distinguishes Aureobasidium from
other related genera. This conidiation is also known in
sporodochial Kabatiella species; its micromorphology on
plate is very similar to that of A. pullulans. The additional
percurrent condition of Aureobasidium is identical to that in
the anamorph genus Hormonema, which makes identification
sometimes difficult. Although no commercial products
presently are available for specific detection of Aureobasidium,
molecular methods like polymerase chain reaction and DNA
sequence analysis of the internal transcribed spacer (ITS) loci
are well suited for a more secure identification. Phylogenetic
analysis of the ITS region is useful to distinguish Aureobasi-
dium isolates on species level.
Importance of Aureobasidium as Spoiler
for Food Industry
In the 1980s A. pullulans was determined to be one of the
causative molds for the black spot spoilage of frozen meat
transported long distances by sea. Colonies produced by
A. pullulans mainly were located in the subsurface of the meat
tissue, with the hyphae spreading along the intercellular
junctions, possibly in response to the arid conditions at the
frozen meat surface. During the past decades, temperature
control was better controlled during shipping and distribution
and black spot spoilage seems no longer to be an issue.

The occurrence of Aureobasidium on other frozen products no
longer is reported.
Assumptions are made that A. pullulans can cause rotting of
healthy fruits and vegetables because of its ability to produce
pectinolytic enzymes. Pectin is a part of the cell wall of fruits
and might be degraded by Aureobasidium. The production of
pectinolytic enzymes is no guarantee for spoilage because
pectin will be broken down during natural ripening of the
fruits. Furthermore, A. pullulans is isolated from a wide range of
fruits and vegetables, but only rarely is designated as a cause of
spoilage.
To control fungi on vegetables and fruits, modified atmo-
sphere can have positive effects, such as increasing the lag phase
of fungal growth, repressing mycelial growth, and decreasing
spore development. Increasing the carbon dioxide content or
decreasing the oxygen content of the atmosphere has been
reported to be fungistatic on A. pullulans.
Health Impact of Aureobasidium in Food
Human mycotoxins are not known to be produced by
Aureobasidium. A few clinical records exist on the presence of A.
pullulans in immunocompromised or traumatically injured
patients causing divergent mycosis, such as phaeohyphomy-
cosis, keratomycosis, or pulmonary mycoses.
Aureobasidium 109
Symptoms vary with the portal of entry and condition of the
host. Health test that were done with A. pullulans strains
selected as preharvest fungicide showed no toxicity, infection,
or pathogenicity to rats. No health hazards have been reported
associated with the presence of Aureobasidium on food.
See also: Spoilage of Meat; Mycotoxins: Classification.
Further Reading
Deshpande, M.S., Rale, V.B., Lynch, J.M., 1992. Aureobasidium pullulans in applied
microbiology: a status report. Enzyme and Microbial Technology 14, 514–527.
Gaur, R., Singh, R., Gupta, M., Gaur, M.K., 2010. Aureobasidium pullulans, an
economically important polymorphic yeast with special reference to pullulan.
African Journal of Biotechnology 9, 7989–7997.
Gill, C.O., Lowry, P.D., Di Menna, M.E., 1981. A note on the identities of organisms
causing black spot spoilage of meat. Journal of Applied Microbiology 51,
183–187.
Manitchotpisit, P., Skory, D., Peterson, S.W., et al., 2011. Poly(b-L-malic acid)
production by diverse phylogenetic clades of Aureobasidium pullulans. Journal of
Industrial Microbiology and Biotechnology 39, 125–132.
Pitt, J.I., Hocking, A.D., 2009. Fungi and Food Spoilage, third ed. Springer, Dordrecht,
Heidelberg, London, New York.
Samson, R.A., Houbraken, J., Thrane, U., Frisvad, J.C., Andersen, B., 2010. Food and
Indoor Fungi, first ed. CBS-KNAW Fungal Biodiversity Centre, Utrecht.
Zalar, P., Gostincar, C., De Hoog, C.S., 2008. Redefinition of Aureobasidium pullulans
and its varieties. Studies in Mycology 61, 21–38.