Variable adhesion and diurnal population patterns of epiphytic yeasts on creeping bentgrass

Irrigation and an in vitro agitation assay were used to determine the percentage of the epiphytic yeast community (Cryptococcus, Pseudozyma, Rhodotorula, and Sporobolomyces) adhering to the phylloplane of creeping bentgrass (Agrostis palustris (Huds.) Pers.). Colony-forming units (cfu) of total epiphytic yeast populations (adherent and nonadherent cells) and of adherent populations (cells not removed by agitation) were determined by leaf washing and dilution plating. In an in vitro assay, 40.0% and 57.1% of the yeast adhered to the leaves, whereas, in initial field trials the percentage of adherent yeasts ranged from 40.0% to 71.9% of the total population. Adherent yeast cfu on leaves in the morning were significantly lower on bentgrass (8.0 x 103 to 3.1 x 104 cfu.cm(-2)) compared with total yeast cfu (1.4 x 104 to 4.7 x 104 cfu.cm(-2)) on the nonirrigated control. No differences in yeast populations were observed between irrigated and nonirrigated plots 2 h after the 0900 treatments. Yeast populations followed a diurnal pattern, with larger cfu recovered from bentgrass leaves in the morning and significantly lower populations recovered in the afternoon. At 1400 the adherent yeast were 83.1%-100% of the total yeast population recovered from the leaves. The relative adhesiveness of the epiphytic yeast community on bentgrass leaves is dynamic with nonadherent cells making up a larger percentage of the population in the mornings than the afternoons.     

In vitro attachment of phylloplane yeasts to Botrytis cinerea, Rhizoctonia solani, and Sclerotinia homoeocarpa

The ability of yeasts to attach to hyphae or conidia of phytopathogenic fungi has been speculated to contribute to biocontrol activity on plant surfaces. Attachment of phylloplane yeasts to Botrytis cinerea, Rhizoctonia solani, and Sclerotinia homoeocarpa was determined using in vitro attachment assays. Yeasts were incubated for 2 d on potato dextrose agar (PDA) prior to experimentation. A total of 292 yeasts cultured on PDA were screened for their ability to attach to conidia of B. cinerea; 260 isolates (89.1%) attached to conidia forming large aggregates of cells, and 22 isolates (7.5%) weakly attached to conidia with 1 or 2 yeast cells attached to a few conidia. Ten yeasts (3.4%), including 8 isolates of Cryptococcus laurentii, 1 isolate of Cryptococcus flavescens, and an unidentified species of Cryptococcus, failed to attach to conidia. All non-attaching yeasts produced copious extracellular polysaccharide (EPS) on PDA. Seventeen yeast isolates did not attach to hyphal fragments of B. cinerea, R. solani, and S. homoeocarpa after a 1 h incubation, but attachment was observed after 24 h. Culture medium, but not culture age, significantly affected the attachment of yeast cells to conidia of B. cinerea. The 10 yeast isolates that did not attach to conidia when grown on agar did attach to conidia (20%-57% of conidia with attached yeast cells) when cultured in liquid medium. Attachment of the biocontrol yeast Rhodotorula glutinis PM4 to conidia of B. cinerea was significantly greater at 1 x 10(7) yeast cells x mL(-1) than at lower concentrations of yeast cells. The ability of yeast cells to attach to fungal conidia or hyphae appears to be a common phenotype among phylloplane yeasts.     

Seasonal dynamics in a yeast population on the Oxalis acetosella L. leaves

Analysis of an epiphytic yeast population on the leaves of the evergreen common wood sorrel Oxalis acetosella L. throughout a year showed that the density and the species composition of this population underwent regular seasonal changes. There were almost no yeasts on the young spring leaves. However, the yeast population on the mature leaves tended to increase in the autumn, reaching a maximum after the formation of continuous snow cover. Then the yeast population on the leaves tended to decrease, reaching a minimum in the spring. The species diversity of the yeasts was maximum in the autumn. The population of the epiphytic yeast species Cystofilobasidium capitatum, Rhodotorula fujisanensis, Leucosporium scottii, and Cryptococcus flavus peaked in the autumn. On the other hand, the population of the widespread epiphytic species Cryptococcus laurentii on the wood sorrel leaves peaked in January. The relative abundance of the red-pigmented phytobionts Rhodotorula glutinis and Sporobolomyces roseus virtually did not change throughout the year. The relative abundance of the euribiotic species Cryptococcus albidus showed irregular monthly variations. The data obtained show that the epiphytic microbial population of various plants can be comprehensively studied only by analyzing this population throughout the vegetative period of the plants.     

Seasonal Dynamics in a Yeast Population on Leaves of the Common Wood Sorrel Oxalis acetosella L.

Analysis of an epiphytic yeast population on the leaves of the evergreen common wood sorrel Oxalis acetosella L. throughout a year showed that the density and the species composition of this population underwent regular seasonal changes. There were almost no yeasts on the young spring leaves. However, the yeast population on the mature leaves tended to increase in the autumn, reaching a maximum after the formation of continuous snow cover. Then the yeast population on the leaves tended to decrease, reaching a minimum in the spring. The species diversity of the yeasts was maximum in the autumn. The population of the epiphytic yeast species Cystofilobasidium capitatum, Rhodotorula fujisanensis, Leucosporium scottii, and Cryptococcus flavus peaked in the autumn. On the other hand, the population of the widespread epiphytic species Cryptococcus laurentii on the wood sorrel leaves peaked in January. The relative abundance of the red-pigmented phytobionts Rhodotorula glutinis and Sporobolomyces roseus virtually did not change throughout the year. The relative abundance of the euribiotic species Cryptococcus albidus showed irregular monthly variations. The data obtained show that the epiphytic microbial population of various plants can be comprehensively studied only by analyzing this population throughout the vegetative period of the plants.     

Groups and sources of yeasts in house dust

House dust contains bacteria, mycelial fungi, microarthropods, and yeasts. The house dust samples collected in 25 apartments in Moscow and the Moscow region were found to contain yeasts belonging to the genera Candida, Cryptococcus, Debaryomyces, Rhodotorula, Sporobolomyces, and Trichosporon. The most frequently encountered microorganisms were typical epiphytic yeasts, such as Cryptococcus diffluens and Rhodotorula mucilaginosa, which are capable of long-term preservation in an inactive state. The direct source of epiphytic yeasts occurring in the house dust might be the indoor plants, which were contaminated with these yeasts, albeit to a lesser degree than outdoor plants. Along with the typical epiphytic yeasts, the house dust contained the opportunistic yeast pathogens Candida catenulata, C. guillermondii, C. haemulonii, C. rugosa, and C. tropicalis, which are known as the causal agents of candidiasis. We failed to reveal any correlation between the abundance of particular yeast species in the house dust, residential characteristics, and the atopic dermatitis of the inhabitants.     

Psychrophilic yeasts in glacial environments of Alpine glaciers

The presence of psychrophilic yeasts in supra- and subglacial sediments, ice and meltwater collected from two glaciers of the Italian Alps (Forni and Sforzellina-Ortles-Cevedale group) was investigated. After incubation at 4 degrees C, subglacial sediments contained from 1.3 x 10(3) to 9.6 x 10(3) CFU of yeasts g(-1). The number of yeast cells in supraglacial sediments was c. 10-100-fold lower. A significant proportion of isolated yeasts exhibited one or more extracellular enzymatic activities (starch-degrading, lipolytic, esterolytic, proteolytic and pectinolytic activity) at 4 degrees C. Selected isolates were able to grow at 2 degrees C under laboratory-simulated in situ conditions. In all, 106 isolated yeasts were identified by MSP-PCR fingerprinting and 26S rRNA gene sequencing of the D1/D2 region as belonging to 10 species: Aureobasidium pullulans, Cryptococcus gilvescens (over 50% of the total), Cryptococcus terricolus, Mrakia gelida, Naganishia globosa, Rhodotorula glacialis, Rhodotorula psychrophenolica, Rhodotorula bacarum, Rhodotorula creatinivora and Rhodotorula laryngis. Four strains, all belonging to a new yeast species, yet to be described, were also isolated.     

Identity and biodegradative abilities of yeasts isolated from plants growing in an arid climate

Plants harvested in the Canary Islands Lanzarote and Fuerteventura were analyzed for the yeasts inhabiting their surface. Half of the isolates (22 out of 44) were identified as Debaryomyces hansenii. Black ascomycetes, viz. Hortaea werneckii and two Hormonema species were represented by 7 strains. Basidiomycetous yeasts, viz. Cryptococcus sp. (8 strains), Rhodotorula sp. (5 strains), Cerinosterus cyanescens (1 strain) and Pseudozyma sp. (1 strain) constituted a minority of 33%. Thirty strains were screened for their ability to assimilate various plant constituents including lipids of the cuticle and the cell membrane, hemicelluloses, nitrogenous compounds (protein, nucleic acids, amino acids) and benzene compounds. All strains were able to assimilate or to hydrolyze lipids, lecithin included. Many strains of D. hansenii, H. dematioides, H. werneckii, C. cyanescens, Cr. laurentii, Pseudozyma sp. and Rh. glutinis were proteolytic. Hemicelluloses like xylan and pectin were assimilated by black ascomycetous yeasts, Cryptococcus sp., Pseudozyma sp. and Rh. glutinis. Ferulic and hydroxycinnamic acids, gallic and tannic acids were assimilated by some strains of H. dematioides, C. cyanescens, Pseudozyma sp. and Rhodotorula sp.     

Groups and Sources of Yeasts in House Dust

House dust contains bacteria, mycelial fungi, microarthropods, and yeasts. House dust samples collected in 25 apartments in Moscow and the Moscow region were found to contain yeasts belonging to the genera Candida, Cryptococcus, Debaryomyces, Rhodotorula, Sporobolomyces, and Trichosporon. The most frequently encountered microorganisms were typical epiphytic yeasts, such as Cryptococcus diffluens and Rhodotorula mucilaginosa, which are capable of long-term preservation in an inactive state. The direct source of epiphytic yeasts occurring in the house dust might be indoor plants, which were contaminated with these yeasts, albeit to a lesser degree than outdoor plants. Along with the typical epiphytic yeasts, the house dust contained the opportunistic yeast pathogens Candida catenulata, C. guillermondii, C. haemulonii, C. rugosa, and C. tropicalis, which are known as the causal agents of candidiases. We failed to reveal any correlation between the abundance of particular yeast species in the house dust, residential characteristics, and the atopic dermatitis of the inhabitants.     

Yeasts in the flowers of entomophilic plants of the Moscow region

Dynamics of abundance and diversity of epiphytic yeasts in entomophilic flowers of 28 species of meadow, forest, and cultivated plants throughout their blooming period was determined. The number of yeasts in the flowers was shown to increase gradually during the vegetation period, and reached the maximum during summer-autumn. The total abundance and ratio of the yeast species in the flowers depended entirely on the blooming time, rather than on the taxonomic position of the plants. Three stages of development of the entomophilic yeast complexes during the vegetation period may be discerned: predominance of eurybiont nonspecific species (Cryptococcus albidus, Debaryomyces hansenii) in spring, mass development of specific nectar-associated yeasts (Metschnikowia reukaufii) in summer, and their substitution by widespread epiphytic species (Rhodotorula mucilaginosa, Cryptococcus magnus) in autumn.     

Endophytic yeasts in leaf galls

Yeast abundance and species diversity of endophytic complexes in galls (cecidia) formed on the leaves of Salix fragilis, Salix caprea, Quercus robur, Tilia cordata, and Ulmus laevis and the epiphytic yeast communities of undamaged leaves of these plants were studied. Dynamics of yeast abundance in the galls was significantly different from that of the epiphytic yeast communities. Maximum numbers of endophytic yeast cells in the galls (up to 10⁴ CFU/g) were comparable to abundance of epiphytic yeasts. A total of 14 species of endophytic yeasts were isolated from galls of different plants. Ascomycetous yeasts were found to predominate in the insect galls on willows and oak, while basidiomycetous yeasts dominated in mite galls on linden and elm, as well as on plant leaves. These results indicate that gall formation may be considered not only as a bidirectional pathological process of the interaction between plants and invertebrates, but also as a process in which the endophytic microbial population of the galls plays an important role.     

Yeasts associated with fresh and frozen pulps of Brazilian tropical fruits

The occurrence of yeasts on ripe fruits and frozen pulps of pitanga (Eugenia uniflora L), mangaba (Hancornia speciosa Gom.), umbu (Spondias tuberosa Avr. Cam.), and acerola (Malpighia glaba L) was verified. The incidence of proteolytic, pectinolytic, and mycocinogenic yeasts on these communities was also determined. A total of 480 colonies was isolated and grouped in 405 different strains. These corresponded to 42 ascomycetous and 28 basidiomycetous species. Candida sorbosivorans, Pseudozyma antarctica, C. spandovensis-like, C. spandovensis, Kloeckera apis, C. parapsilosis, Rhodotorula graminis, Kluyveromyces marxianus, Cryptococcus laurentii, Metchnikowia sp (isolated only from pitanga ripe fruits), Issatchenkia occidentalis and C. krusei (isolated only from mangaba frozen pulps), were the most frequent species. The yeast communities from pitanga ripe fruits exhibited the highest frequency of species, followed by communities from acerola ripe fruits and mangaba frozen pulps. Yeast communities from frozen pulp and ripe fruits of umbu had the lowest number of species. Except the yeasts from pitanga, yeast communities from frozen pulp exhibited higher number of yeasts than ripe fruit communities. Mycocinogenic yeasts were found in all of the substrates studied except in communities from umbu ripe fruits and pitanga frozen pulps. Most of the yeasts found to produce mycocins were basidiomycetes and included P. antarctica, Cryptococcus albidus, C. bhutanensis-like, R. graminis and R. mucilaginosa-like from pitanga ripe fruits as well as black yeasts from pitanga and acerola ripe fruits. The umbu frozen pulps community had the highest frequency of proteolytic species. Yeasts able to hydrolyse casein at pH 5.0 represented 38.5% of the species isolated. Thirty-seven percent of yeast isolates were able to hydrolyse casein at pH 7.0. Pectinolytic yeasts were found in all of the communities studied, excepted for those of umbu frozen pulps. The highest frequency of pectinolytic activity was found in mangaba frozen pulp communities. Around 30% of all isolates produced pectinases. The ability to split arbutin was observed in all communities ranging from 8% in yeasts from pitanga frozen pulps to 40.6% in acerola ripe fruit communities. Among 432 species tested, 125 were active for beta-glucosidase production, and Kloeckera apis, P. antarctica, C. sorbosivorans, and C. spandovensis-like were the most active species.     

The yeast <Emphasis Type="Italic">Candida railenensis</Emphasis> in the fruits of English oak (<Emphasis Type="Italic">Quercus robur</Emphasis> L.)

The cotyledons of whole intact acorns were shown to contain yeasts; their number increased sharply before acorn germination. The yeasts in the cotyledons are mainly represented by one species, Candida railenensis, with the number in the germinating cotyledons reaching 10⁷ CFU/g. After germination or exocarp destruction, the cotyledons were colonized by the usual epiphytic and litter yeasts Cryptococcus albidus, Rhodotorula glutinis, and Cystofilobasidium capitatum.     

Seasonal dynamics of the structure of epiphytic yeast communities

The seasonal dynamics of the species structure of epiphytic yeasts on the leaves and in the flowers of 25 plant species was studied throughout the period of their vegetation. It was shown that, on average for the vegetation period, the composition of epiphytic yeast communities was nonspecific. The same species of epiphytic yeasts dominated on different plant species, irrespective of their taxonomic identity and ecological peculiarities. However, different species of yeasts exhibited different types of seasonal dynamics of relative abundance. Therefore, a combination of the dynamics of yeast species and the ontogenetic cycles of plants creates a pattern of the dynamics of the epiphytic yeast population, which is unique for each plant species. The species diversity of yeasts on the leaves of a plant is determined by the duration of its ontogenetic cycle: the longer the vegetation of a plant, the higher the diversity of the epiphytic yeasts population. The greatest diversity of epiphytic yeasts was revealed on the leaves of perennial hygrophytes and mesophytes; the minimal diversity, on ephemeroids and annuals with a short ontogenetic cycle.     

Influence of pesticides on yeasts colonizing leaves

The effect of nine different pesticides on the growth of yeasts isolated from the leaves of fruit and forest trees was investigated. Four insecticides (with the active ingredients: thiacloprid, deltamethrin, lambdacyhalothrin, and thiamethoxam) and five fungicides (with the effective substances: bitertanol, kresoxim-methyl, mancozeb, trifloxystrobin, and cupric oxychloride) were tested. The concentrations of chemicals were those recommended by the manufacturers for the spraying of trees. The yeast strains isolated from the leaves of fruit trees were not sensitive to any of the insecticides. The majority of yeast strains isolated from the leaves of forest trees were either not sensitive or only to a small extent. While Rhodotorula mucilaginosa and Pichia anomala were not affected by any insecticide, the strains of Cryptococcus laurentii and Rhodotorula glutinis showed the highest sensitivity. The effects of fungicides on the growth of isolated yeasts were more substantial. The fungicide Dithane DG (mancozeb) completely inhibited the growth of all yeasts. All strains isolated from fruit tree leaves were more resistant to the tested fungicides than those isolated from the leaves of forest trees. The most resistant strains from the leaves of fruit trees belonged to the species Metschnikowia pulcherrima, Pichia anomala, and Saccharomyces cerevisiae, whereas Cryptococcus albidus and C. laurentii, originating from the leaves of forest trees, showed the highest sensitivity to fungicides.     

Assessment of endophytic yeast diversity in rice leaves by a culture-independent approach

Endophytic microorganisms inhabit internal plant tissues in the host plant without causing any symptoms or negative effects. Although the diversity of endophytes has been evaluated by both culture-dependent and culture-independent methods, less information is available on yeast communities. Therefore, in this study a culture-independent method was used to examine endophytic yeasts associated with rice leaves based on the large subunit of ribosomal DNA using a semi-nested PCR technique. Sequence analysis indicated that the colonization frequency and the relative species frequency (RF) of endophytic yeast phylotypes were 0.41 and 0.06, respectively, and the majority of the yeast phylotypes were basidiomycetous yeasts. The phylotypes were designated as five known species (Cryptococcus victoriae, Debaryomyces hansenii, Debaryomyces vindobonensis, Meyerozyma guilliermondii and Pseudozyma antarctica), together with seventeen phylotypes closest to Candida metapsilosis, Cryp. foliicola, Cryp. laurentii, Pseudozyma abaconensis, Pseudozyma aphidis and Trichosporon asahii, among which some could be novel species. The most prevalent phylotypes were those closest to Cryp. foliicola (47.5 % RF) followed by D. hansenii (22.8 % RF) and P. antarctica (16.8 % RF). The presence of the phylotypes related to species known for their potential applications as biocontrol agents and plant growth promoting hormone producers suggests that they may have valuable applications. In addition, our findings revealed the occurrence of novel phylotypes at high frequency, which should encourage extensive studies to discover novel yeast species and to understand their roles in the rice leaves.     

Characterization of yeast groupings in the phyllosphere of Sphagnum mosses

Significant differences were revealed in the taxonomic structure of the epiphytic yeast communities formed on Sphagnum mosses and on the leaves of vascular plants. On mosses, low abundance of red yeasts was found (the most typical epiphytes on vascular plant leaves), along with a relatively high content and diversity of nonpigmented dimorphic basidiomycetes related to the order Leucosporidiales. The species composition of epiphytic yeasts from mosses is different from that of both forest and meadow grasses and of the parts of vascular plants submerged in the turf. The specific composition of the Sphagnum mosses yeast community is probably determined by the biochemical characteristics of this environment, rather than by the hydrothermal regime in the turf.     

Fatty acid makeup of the lipids in soil and epiphytic yeasts]

The fatty acid composition of lipids was compared among yeast cultures belonging to the genera Rhodotorula, Lipomyces, and Cryptococcus. These lipids contain C10--C26 fatty acids, mainly with the even number of carbon atoms. Palmitic acid (C16 : 0) and oleic acid (C18 : 0) predominate. In the majority of the strains, the sum of unsaturated acids exceeds the sum of saturated acids. The content of unsaturated acids in the lipids of the epiphytic yeast Rhodotorula is higher than in the soil yeast Lipomyces. Besides C12--C18 acids, C22--C26 acids were identified by GLC at preset temperatures. Lignoceric acid (C24 : 0) was found for the first time in the cultures of Rhodotorula, Lipomyces, and Cryptococcus, and cerotinic acid (C16 : 0) was also detected in the Rhodotorula yeast. Fatty acids with a long chain are registered in the strains of Rhodotorula more often than in the strains of Lipomyces and Cryptococcus.     

Seasonal occurrence of yeasts and yeast-like organisms in the river Danube

One hundred and seventy yeast strains belonging to 14 genera and 29 species were isolated from 112 water samples of the river Danube in the area of Bratislava. The samples were collected through the year from April to March.Saccharomyces cerevisiae, Candida maltosa, Aureobasidium pullulans, Cystofilobasidium capitatum, Rhodotorula glutinis, Geotrichum candidum, and Candida krusei were the most frequent. The basidiomycetous yeasts and yeast-like organisms with oxidative metabolism were present in approximately equal numbers to those with fermentative metabolism. Saccharomyces cerevisiae was the dominant yeast and was isolated from 50% of all samples examined and represented approximately one quarter of the yeast community.Yeast densities ranged from 100 to 21,100 CFU per litre. The highest population density was observed in October. Cryptococcus albidus, Saccharomyces cerevisiae, Rhodotorula glutinis, and Aureobasidium pullulans formed the main part of the yeast population in this month.     

Growth of epiphytic and soil yeasts on wheat seedlings

Colonization of wheat seedlings by epiphytic (Rhodotorula glutinis) and soil (Lipomyces starkeyi) yeasts was studied by scanning electron microscopy. Epiphytic yeast cells dominated on the plant surface. Soil yeast cells were randomly distributed among both the zones of a seedling and the particles of an inorganic substrate. It has been found that epiphytic yeast strains can readily grow on the surface of a plant.     

Endophytic yeasts in <Emphasis Type="Italic">Malus domestica</Emphasis> and <Emphasis Type="Italic">Pyrus communis</Emphasis> fruits under anthropogenic impact

Yeast abundance and species diversity on the surface and in inner tissues of Malus domestica and Pyrus communis fruits under high anthropogenic impact in the city of Moscow (Russia) were studied. Results demonstrated that abundance of epiphytic yeasts on the fruits increased gradually, reaching the maximum of 3.2 × 10⁴ CFU/g on mature fruits. During summer, abundance of endophytes did not change significantly (variation near 2.5 × 10³ CFU/g) until complete maturation, while in September their numbers increased to 10⁴ CFU/g. Basidiomycetous yeasts (Filobasidium wieringae, F. magnum, Rhodotorula glutinis, and Rhodosporidiobolus colostri) predominated on the fruit surface. Ascomycetous species were the most diverse group inside the fruits, which quantitatively increased through maturation. It was found that the share of opportunistic species Candida parapsilosis in internal tissues was significant during the entire period of fruit formation and development under anthropogenic impact in the city. Specific properties of epiphytic and endophytic yeast communities developing in natural ecological niches under synanthropic conditions and anthropogenic impact are discussed.