J. Basic Microbiol. 43 (2003) 5, 430 – 436 DOI: 10.1002/jobm.

200310277

(Culture Collection of Yeasts, Institute of Chemistry, Slovak Academy of Sciences, Dúbravská cesta 9,
842 38 Bratislava, Slovakia)

The diversity of yeasts in the agricultural soil

ELENA SLÁVIKOVÁ* and RENATA VADKERTIOVÁ

(Received 07 March 2003/Accepted 25 April 2003)

One hundred and eleven yeast strains were isolated from 60 agricultural soil samples. The samples
were taken from four various fields located in the southwest of Slovakia. Cryptococcus laurentii, Can-
dida maltosa, Metschnikowia pulcherrima, and Sporobolomyces salmonicolor were the predominant
species in the samples collected from all four types of fields. These species represented 78.4 – 86.6%
of the total yeast counts.
The results obtained enabled comparisons to be made between forest and agricultural soil yeast
population. We have found out that the yeast population in tilled soils was significantly reduced. The
number of yeasts in the tilled soils ranged from 40 to 6.8 × 103 CFU/g soil and the average number
reached approximately 1.12 × 103. This number is more than ten times lower in comparison with the
forest soils.

Microorganisms form the basis of the ecological balance of the biosphere. The composition
of the microbial communities influences the transformation of plant residues into soil or-
ganic matter and plant available nutrients, and also stabilises soil aggregates, reduces ero-
sion and maintains the water-holding capacity (BEARE et al. 1993, KENNEDY and GEWIN
1997). Yeasts are important organisms in many ecosystems and form a significant contribu-
tion to biodiversity (FLEET 1998).
The soil is the ultimate repository for storage and an even development of certain species
of yeasts. The yeast density may range from none or a few to several thousand cells per
gram of soil. Yeast flora found in soil more or less reflects the yeast flora associated with
plants, animal, and fungal life above, but many yeast species are typical inhabitants of soil.
The presence of yeast organisms in soil depends also on many factors, e.g. type of soil,
rainfall, and climate. It is evident that the ability of yeasts to survive in this habitat plays a
fundamental role (PHAFF and STARMER 1987).
The occurrence of yeasts in soil has been studied in various parts of the world (JENSEN
1963, VISHNIAC 1996, DMITRIEV et al. 1997). Yeasts were found in tropical (MOK et al.
1984) as well as antarctic soils (BAUBLIS et al. 1991). Soil biodiversity is generally high in
forests, which may represent “biodiversity hot spots” in agricultural landscapes. Forest soils
tend to be species-rich and represent stable and often old environments (HÄGVAR 1998).
Only little information on yeasts associated with soil on the territory of Slovakia and
neighbouring countries is available. This work is a part of a broader survey of the occur-
rence of yeasts in various types of soils in the southwest of Slovakia. In our previous inves-
tigations we studied the occurrence of yeasts and yeast-like species in forest and grass-
grown soil (SLÁVIKOVÁ and VADKERTIOVÁ 2000, 2003). The aim of this work was to study
the occurrence of yeast populations in soil samples taken from four various fields where
different crops were growing, and to compare them with those found in no-tilled soils.

* Corresponding author: Dr. E. SLÁVIKOVÁ; e-mail: chemslav@savba.sk

© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 0233-111X/03/0509-0430

The diversity of yeasts in the agricultural soil 431

Materials and methods

The soil samples were taken from four various fields located in the southwest of Slovakia: maize field,
sugar beet field, potato-field, and grein-field. The collection of soil was carried out in August when
fertilizing and pesticides applications were completed and intensive sunshine and dryness occurred.
Fifteen samples were collected from every field. In total, this resulted in 60 samples from which yeasts
and yeast-like organisms were isolated. The samples were taken from a depth of 5 cm and put into the
sterile bottles, transported to the laboratory, and processed within 2 hours after collection.
Five grammes of the soil sample were suspended in 50 ml of sterile tap water and shaken on a
rotary shaker for 1 h. The suspension was diluted to a 1 % solution. An inoculum of 0.5 ml was put on
each 10 cm plate with agar and spread with a glass spreader. For cultivation, malt-extract agar and
glucose agar (2% glucose, 0.3% yeast extract, and 0.1% (NH4)2SO4, 0.02% K2HPO4, 0.01% KH2PO4,
0.02% MgSO4 ⋅ 7 H2O, 0.02% NaCl, 0.01% K2SO4, 2% agar, water) were used. Both media contained
80 µg · ml–1 streptomycin. The plates were incubated at 25 °C (2 – 3 d) and 7 °C (14 d). Sodium pro-
pionate (0.25%) reduces the growth of hyphal fungi and was added to the part of medium used for
cultivation at 25 °C. Colonies of different appearance were counted in both medium variants, and their
representatives were purified according to SLÁVIKOVÁ et al. (1992). Yeast counts were calculated as
the number of colony-forming units (CFU) per dry gramme of soil sample. Dry weights for soil
samples were determined gravimetrically by subsampling c. 1 g of soil and drying in a drying oven at
105 °C to constant weight.
The morphological and physiological characteristics of isolates were examined by the methods
described by VAN DER WALT and YARROW (1984). Strains were identified to species according to
KURTZMAN and FELL (1998).

Results and discussion

One hundred and eleven yeast strains belonging to 8 genera and 11 species were isolated
from 60 soil samples. Table 1 provides a list of isolated species. Cryptococcus laurentii,
Candida maltosa, Metschnikowia pulcherrima, and Sporobolomyces salmonicolor were the
most frequently isolated species from the samples taken in all four types of fields. These
species represented 78.4–86.6% of the total yeast counts found in the soil samples.
The species Cryptococcus laurentii was very common in soil samples taken from all four
fields, and it formed 16.4–35.6% of the total yeast counts found out in the soil samples of
the individual fields (Table 1). This species was very frequently found also in the forest and
grass-grown soils, where it formed 13.0–18.8% and 18.9–45.1% of the total yeast popu-
lation, respectively (SLÁVIKOVÁ and VADKERTIOVÁ 2000, 2003). C. laurentii is often
associated with plants and soils. An investigation of various soils indicate that this species
was the most frequently encountered in tundra (BABJEVA and AZIEVA 1980), Antarctic
(VISHNIAC 1996), and prairie (SPENCER and SPENCER 1997) soils. On the other hand,
C. laurentii formed just 4.1 and 6.0% of the total yeast population occurring in eutrophised
Danube and Morava river water, respectively (SLÁVIKOVÁ and VADKERTIOVÁ 1997,
1997a). It assimilates a very wide range of sugars and related compounds and has also been
shown to use flavonoids (SPENCER et al. 1971). Species of Cryptococcus belong to the
group of capsulated yeasts, which survive better in habitats poor in nutrients and during
periods of desiccation. Capsules may serve to protect cells from physical and biological
stresses encountered in their natural habitat and may influence the ability of the cells to
survive low moisture conditions (SPENCER and SPENCER 1997, GOLUBEV 1991). These
insoluble extracellular polysaccharides can also act as soil binding agents (CAESAR-TON
THAT et al. 2001).
Candida maltosa made up 5.1–45.8% of the yeast counts found out in soil samples taken
from the four tilled fields (Table 1). It also belonged to the predominant species occurred in
the river waters (SLÁVIKOVÁ and VADKERTIOVÁ 1997, 1997a), but in the forest and
grass-grown soils was found very rarely. CERNIGLIA et al. (1992) demonstrated the ability of
432 E. SLÁVIKOVÁ and R. VADKERTIOVÁ

Table 1
The occurrence of species isolated from soil samples collected from the studied fields
Species Occurrence of individual species related to the total yeast
counts (%) in each field
maize sugar beet grein- potato-
field field field field
Candida maltosa 16.7 17.1 5.1 45.8
Candida valida 0.3
Cryptococcus albidus 1.1
Cryptococcus laurentii 35.6 34.2 27.5 16.4
Cystofilobasidium capitatum 1.5 10.3 3.3
Debaryomyces occidentalis var. 6.6
occidentalis
Metschnikowia pulcherrima 9.1 32.8 37.8 11.9
Sporobolomyces salmonicolor 17.0 2.7 15.8 7.7
Trichosporon cutaneum 2.7 2.6
Trichosporon pullulans 19.4 14.9
Williopsis saturnus var. saturnus 3.5

C. maltosa to oxidize polycyclic aromatic hydrocarbons, which can have an adverse effect
on the environment.
Metschnikowia pulcherrima is the third species, which was found very frequently in the
soil samples taken from all four fields. It formed 9.1–37.8% of the total yeast counts found
out in soil samples of the individual fields. This species did not occurred in the forest and
grass-grown soils and it was rarely isolated from various types of water. It was reported that
M. pulcherrima belonged to the most widespread species isolated from various parts of
plants collected in North-East Mecklenburg in Germany (KOCKOVÁ-KRATOCHVÍLOVÁ et al.
1972).
The last species, which was often presented in the soil samples taken from all four fields,
is Sporobolomyces salmonicolor. The proportion of this carotenoids producing species was
2.7–17.0% (Table 1). It was also found in forest and grass-grown soils, but in smaller
proportion. SPENCER et al. (1971) isolated Sp. salmonicolor from soils of the Matador site in
southern Saskatchewan. This species forms ballistokonidia and is often associated with the
phyllosphere of plants. During periods of bright sunlight, carotenoids protect the
photosynthetic apparatus of plants against photodestruction (GOODWIN 1981). Similarly,
they may also protect the vital structures and processes of red yeasts in upper layers of soil.
Other carotenoids producing species Cystofilobasidium capitatum was also regularly
isolated organisms from the soil samples taken from maize field, sugar beet field, and po-
tato-field, where formed 1.5–10.3% of yeast counts. It was not found only in the samples
taken from the grein-field. Cyst. capitatum was the dominant species in the forest and grass-
grown soils (SLÁVIKOVÁ and VADKERTIOVÁ 2000, 2003).
The genus Trichosporon was represented by the two species. Tr. cutaneum was present in
the samples taken from the sugar beet field and from the grein-field; it formed only 2.6 and
2.7% of the total yeast population, respectively. On the other hand, Tr. pullulans occurred in
higher proportions in the soil samples taken from the maize field and the potato-field and it
constituted 19.4 and 14.9% of the total yeast population, respectively (Table 1). Both these
species were found also in the forest and grass-grown soils (SLÁVIKOVÁ and VADKERTIOVÁ
2000, 2003). The frequent occurence of the above yeast species in soil was also observed by
other authors. JENSEN (1963) documented Trichosporon sp. in his studies on the microflora
of Danish beech forest soils. SPENCER and SPENCER (1997) isolated numerous cultures of
the species Tr. cutaneum from Californian and Florisa soils. We have found out that the
The diversity of yeasts in the agricultural soil 433

strain Tr. pullulans, which we have isolated from soil can act as a biodegrader of lignin, the
most resistant component of lignocellulosics in nature, and so this organisms could improve
the transformation of plant residues into soil organic matter (SLÁVIKOVÁ et al. 2002).
Four species (Candida valida, Williopsis saturnus var. saturnus, Cryptococcus albidus,
and Debaryomyces occidentalis var. occidentalis) occurred only in the samples taken from
one of four fields and their contribution to the yeast counts found in the agricultural soils
was lower.
As Table 2 shows, the yeast community isolated from tilled soils consisted of species
with the ability to assimilate the hemicellulose-derived sugars xylose and cellobiose
together with the nonreducing disaccharide trehalose. These sugars are widespread in nature
and all strains assimilated them. Approximately one half of yeasts isolated from three fields
and 74% of the yeasts isolated from maize field were capable to utilize also arabinose and
soluble starch. The smaller part of the yeast community (31–57%) could grown also on
lactose and inozitol. Only 13–38% of yeasts utilize nitrate as the sole nitrogen source. The
fermentation of glucose was positive for 26–58% of the yeast community, contrary to the
yeasts isolated from forest soils, where the fermentation of glucose was positive only for
0.6–5.6% of isolates (SLÁVIKOVÁ and VADKERTIOVÁ 2000). It apears from Table 2 that the
presence of basidiomycetous and ascomycetous yeasts was approximately equal in the soil
samples taken from three fields, but in the samples taken from maize field predominated the
basidiomycetous yeasts. The marked prevalence of basidiomycetous yeasts (94–99%) was
found in samples taken from forest soils (SLÁVIKOVÁ and VADKERTIOVÁ 2000).
We have also found quantitative differences between yeast counts occurring in the agri-
cultural and forest soils. The number of yeasts in the agricultural soils ranged from 40 to
6.8 × 103 CFU/g soil and the average number reached approximately 1.12 × 103. This
number is more than ten time lower in comparison with the forest soils, which contained in
average 1.4 × 104 CFU/g soil. In August the average numbers of yeasts counted in no-tilled
soils were about seven to eight time higher than those in agricultural soils.
As it can be seen from Fig. 1, the quantitative representation of the dominant yeast
species found in soil samples of different four fields is distinct in proportion to the total
yeast counts found in all samples. The highest number of yeasts was determined in samples
collected in maize field. They formed up to 56% of the total yeast counts. The number of
yeasts found in soil samples taken from grein-field and potato-field was lower and
represented 22 and 15% of the total yeast counts, respectively. The lowest number of yeasts
was found in samples taken from sugar beet field, only 7% of the total yeast counts. The

Table 2
Survey of some features of yeast population
Feature % of the occurrence of individual feature
maize field sugar beet field grein-field potato-field
Presence of urease 74 50 47 42
Fermentation of saccharides 26 50 53 58
Assimilation of nitrate 38 13 20 26
Assimilation of D-xylose 100 100 100 100
Assimilation of L-arabinose 74 50 54 42
Assimilation of cellobiose 100 100 100 100
Assimilation of trehalose 100 100 100 100
Assimilation of lactose 57 47 38 35
Assimilation of soluble starch 74 47 38 35
Assimilation of inositol 57 47 31 35
434 E. SLÁVIKOVÁ and R. VADKERTIOVÁ

25
[%]

20
Metschnikowia pulcherrima
Candida maltosa
15 Cryptococcus laurentii
Sporobolomyces salmonicolor

10

0
1 field
maize 2 beet field
sugar potato-field grain-field
Fig. 1
Occurrence of the most frequently isolated species related to the total yeast population found in all
samples taken from four fields

differences among the yeast counts found in soil samples taken from four fields could be
caused by several factors. One of them is application of different pesticides and fertilizers
during the cultivation. The species Cryptococcus laurentii and Sporobolomyces salmoni-
color were very frequently isolated not only from soil samples but also from the surface of
the maize leaves (unpublished results), from where they could be flushed by a rain into the
soil. Also the size of plants can influence the number of yeast organisms in the soil, because
the higher plants better protect the yeasts in the upper layer of the soil against an inhibitory
effect of the sunshine and they also keep the soil humidity for a longer time.
We can concluded that the qualitative representation of the dominant yeast species found
in soil samples of different fields was very similar, but the total yeast counts were distinct.
In comparison with the forest and grass-grown soils, the yeast population in agricultural soil
was significantly reduced. The main reason is probably tillage and fungicide application,
which may negatively affect the presence of yeast organisms. This conclusion is supported
also by other authors, who found out, that the microbial biomass is lower in tilled soils than
soils under no till, reduced tillage or perennial crops (DORAN 1980, SMITH and PAUL 1990).
Fertilizer input to soils cause an accumulation of potentially toxic elements such as heavy
metals, which can also reduce yeast diversity, because the microorganisms are far
more sensitive to heavy metal stress than soil animals or plants growing on the same soils
(PRAHALAND and SEENAYYA 1988).

Acknowledgements

This work was supported by grant no 2/1054/21 from the Vega grant agency for biological and
ecological sciences.
The diversity of yeasts in the agricultural soil 435

References

BABYEVA, I. P. and AZIEVA, E. E., 1980. The taxonomic composition and ecological characters of
yeasts occurring in tundra soils in west Taimir. Mikologija i fitopatologia, 14, 99 – 103.
BAUBLIS, J. A., WHARTON, Jr. R. A. and VOLZ, P. A., 1991. Diversity of microfungi in an Antarctic
dry valley. J. Basic Microbiol., 31, 3 – 12.
BEARE, M. H., POHLAD, B. R., WRIGHT, D. H. and COLEMAN, D. C., 1993. Residue placement and
fungicide effects on fungal communities in conventional and no-tillage systems. Soil Sci. Soc. Am.
J., 57, 392 – 399.
CAESAR-TON THAT, T. C. and COCHRAN, V. L., 2000. Soil aggregate stabilization by a saprophytic
lignin decomposer basidiomycete fungus. I. Microbiological aspects. Biol. Fertil. Soils, 32,
374 – 380.
CERNIGLIA, C. E., SUTHERLAND, J. B. and CROW S. A., 1992. Fungal metabolism of aromatic
hydrocarbons. In: Microbial Degradation of Natural Products (G. WINKELMANN, Editor), pp.
193 – 217. VCH Press, Weinheim.
DMITRIEV, V. V., GILICHINSKII, D. A., FAIZUTDINOVA, R. N., SHERSHUNOV, I. N., GOLUBEV, V. I. and
DUDA, V. I., 1997. Detection of viable yeast in 3-million-year-old permafrost soils of Siberia. Mi-
crobiology, 66, 546 – 550.
DORAN, J. W., 1980. Soil microbial and biochemical changes associated with reduced tillage. Soil Sci.
Soc. Am. J., 44, 765 – 771.
FLEET, G. H., 1998. Yeasts in natural habitats. Food Technol. Biotechnol., 36, 285 – 289.
GOLUBEV, W. I., 1991. Capsules. In: The Yeasts (A. H. ROSE and J. S. HARRISON, Editors), Volume 4,
pp. 175 – 198. Academic Press London.
GOODWIN, T. W., 1981. The Biochemistry of the Carotenoids. Vol. 1. 360 p. Chapman and Hall Lon-
don.
HÄGVAR, S., 1998. The relevance of the Rio-Convention on biodiversity to conserving the biodiversity
of soils. Appl. Soil Ecol., 9, 1 – 7.
JENSEN, V., 1963. Studies on the microflora of Danish beech forest soils. IV. Yeasts and yeast-like
fungi. Zentralbl. Bakteriol., Parasitenkunde, Infektionskrankheiten u. Hygiene, II. Abt., 117, 41 – 65.
KENNEDY, A. C. and GEWIN, V. L., 1997. Soil microbial diversity: present and future considerations.
Soil Science, 162, 607 – 617.
KOCKOVÁ-KRATOCHVÍLOVÁ, A., WEGENER, K. A. and ONDRUŠOVÁ, D., 1972. Ein Beitrag zur Ökologie
der Hefen aus Nordost-Mecklenburg. Mycopathologia et Mycologia Applicata, 48, 191 – 212.
KURTZMAN, C. P. and FELL, J. W., 1998. The Yeasts, a Taxonomic Study. Fourth Revised and
Enlarged Edition. 1055 p. Elsevier, Amsterdam.
MOK, W. I., LUIZÁO, R. C. C., DO SOCORRO BARRETO DA SILVA M., TEIXEIRA, M. F. S. and MUNITZ,
E. G., 1984. Ecology of pathogenic yeasts in Amazonian soil. Appl. Environ. Microbiol., 47,
390 – 394.
PHAFF, H. J. and STARMER, W. T., 1987. Yeasts associated with plants, insects and soil. – In:
The Yeasts (A. H. ROSE and J. S. HARRISON, Editors), Volume 1, pp. 123 – 180. Academic Press
London.
PRAHALAND, A. K. and SEENAYYA, G., 1988. Bioavailability of zinc and cadmium and their effect on
microbial growth and metal uptake. Bull. Environ. Contam. Toxicol., 41, 921 – 927.
SLÁVIKOVÁ, E. and VADKERTIOVÁ, R., 1997. Yeasts and yeast-like organisms occurring in the river
Morava. Food Technol. Biotechnol., 35, 293 – 297.
SLÁVIKOVÁ, E. and VADKERTIOVÁ, R., 1997 a. Seasonal occurrence of yeasts and yeast-like organisms
in the river Danube. Antonie van Leeuwenhoek, 72, 77 – 80.
SLÁVIKOVÁ, E. and VADKERTIOVÁ, R., 2000. The occurrence of yeasts in the forest soils. J. Basic
Microbiol., 40, 207 – 212.
SLÁVIKOVÁ, E. and VADKERTIOVÁ, R., 2003. The occurrence of yeasts in the grass-grown soils. Czech
Mycology, 54, 182 – 187.
SLÁVIKOVÁ, E., VADKERTIOVÁ, R. and KOCKOVÁ-KRATOCHVÍLOVÁ, A., 1992. Yeasts isolated from
artificial lake waters. Can. J. Microbiol., 38, 1206 – 1209.
SMITH, J. L. and PAUL, E. A., 1990. The significance of soil biomass estimates. In: Soil Biochemistry
(J. M. BOLLAG and G. STOTZKY, Editors), Volume. 6, pp. 357 – 396. Marcel Dekker, New York.
SPENCER, J. F. T., BABIUK, L. and MORRALL, R. A. A., 1971. Yeasts of the Matador soils of Sas-
katchewan and their ability to use flavonoids. Can. J. Microbiol., 17, 1248 – 1250.
436 E. SLÁVIKOVÁ and R. VADKERTIOVÁ

SPENCER, J. F. T. and SPENCER, D. M., 1997. Ecology: where yeasts live. In: Yeasts in Natural and
Artificial Habitats (J. F. T. SPENCER and D. M. SPENCER, Editors), pp. 33 – 58. Springer-Verlag Ber-
lin Heidelberg.
VAN DER WALT, J. P. and YARROW D., 1984. Methods for the isolation, maintenance, classification
and identification of yeasts. In: The Yeasts a Taxonomic Study ( N. J. W. KREGER-VAN RIJ, Editor),
pp. 130 – 145. Elsevier Amsterdam.
VISHNIAC, H. S., 1996. Biodiversity of yeasts and filamentous microfungi in terrestrial Antarctic
ecosystems. Biodiversity Conservation, 5, 1365 – 1378.

Mailing address: Dr. ELENA SLÁVIKOVÁ, Culture Collection of Yeasts, Institute of Chemistry, Slovak
Academy of Sciences, Dúbravská cesta 9, 842 38 Bratislava, Slovakia
e-mail: chemslav@savba.sk