Extracellular enzymatic activities of basidiomycetous yeasts isolated from glacial and subglacial waters of northwest Patagonia (Argentina)

As part of a project aimed at the selection of cold-adapted yeasts expressing biotechnologically interesting features, the extracellular enzymatic activity (EEA) of basidiomycetous yeasts isolated from glacial and subglacial waters of northwest Patagonia (Argentina) was investigated. Ninety-one basidiomycetous yeasts (belonging to the genera Cryptococcus, Leucosporidiella, Dioszegia, Mrakia, Rhodotorula, Rhodosporidium, Sporobolomyces, Sporidiobolus, Cystofilobasidium, and Udeniomyces) were screened for extracellular amylolytic, proteolytic, lipolytic, esterasic, pectinolytic, chitinolytic, and cellulolytic activities. Over 15% of the strains exhibited three or more different EEAs at 4 degrees C and more than 63% had at least two EEAs at the same temperature. No chitinolytic or cellulolytic activities were detected at 4 and 20 degrees C. Cell-free supernatants exhibited significantly higher (P < 0.01) protease and lipase activities at < or = 10 degrees C, or even at 4 degrees C. In light of these findings, cold environments of Patagonia (Argentina) may be considered a potential source of cold-adapted yeasts producing industrially relevant cold-active enzymes.     

Biodiversity of cold-adapted yeasts from glacial meltwater rivers in Patagonia, Argentina

The occurrence of culturable yeasts in glacial meltwater from the Frías, Casta?o Overo and Río Manso glaciers, located on Mount Tronador in the Nahuel Huapi National Park (Northwestern Patagonia, Argentina) is presented. Subsurface water samples were filtered for colony counting and yeast isolation. The total yeast count ranged between 6 and 360 CFU L(-1). Physiologic and molecular methods were employed to identify 86 yeast isolates. In agreement with yeast diversity data from studies for Antarctic and Alpine glaciers, the genera Cryptococcus, Leucosporidiella, Dioszegia, Rhodotorula, Rhodosporidium, Mrakia, Sporobolomyces, Udeniomyces and Candida were found. Cryptococcus and Leucosporidiella accounted for 50% and 20% of the total number of strains, respectively. Among 21 identified yeast species, Cryptococcus sp. 1 and Leucosporidiella fragaria were the most frequent. The typically psychrophilic Mrakia yeast strain and three new yeast species, yet to be described, were also isolated. All yeast strains were able to grow at 5, 10, and 15 degrees C. Among yeast strains expressing extracellular enzymatic activity, higher proteolytic and lipolytic activities were obtained at 4 degrees C than at 20 degrees C.     

Alkalitolerant yeasts from natural biotopes

Using a solid nutrient medium containing alkaline buffer (pH 10) and an antibiotic, alkalitolerant yeasts were isolated from samples of soda-rich saline soils (solonchaks) of Armenia (Arazdayan) and the Transbaikal Region (the Kungur Steppe). The species diversity of the yeast populations of the tested soda-rich soils was relatively insignificant. They only contained alkalitolerant representatives of asporogenic capsulated yeasts belonging to the species Cryptococcus laurentii, C. albidus, Rhodotorula glutinis, R. mucilaginosa, and Sporobolomyces roseus. C. laurentii representatives clearly dominated the isolates obtained, their number exceeding that of the other species by 2-3 orders of magnitude. All of the isolates grew on acidic wort agar, suggesting that they did not include obligate alkaliphiles.     

红富士苹果园酵母多样性研究

从北京顺义和山东泰安红富士苹果园采集果实、叶片、树皮和土壤等不同基物,分离酵母菌,利用26S rDNA的D1/D2区域序列分析并结合形态学特征和SSCP分析对这些菌株进行了分类学研究,探讨了苹果园酵母的物种多样性及其分布。北京苹果园共分离酵母菌129株,鉴定为13属21种,优势属为Pichia（4个种）,Cryptococcus（3个种）,Pseudozyma（3个种）,子囊菌占较大优势,分布于8属12种,占总种数的57.1%。山东苹果园共分离酵母291株,鉴定为13属26种,优势属为假丝酵母Candida（6个种）,毕赤酵母Pichia（4个种）和隐球酵母Cryptococcus（3个种）,并且子囊菌占较大优势,分布于7属17种,占总种数的65.4%。     

Isolation of proteolytic psychrotrophic yeasts from fresh raw seafoods

A total of 103 cultures of yeasts were isolated from seven kinds of fresh raw seafoods. The isolates comprised six genera, Candida, Cryptococcus, Debaryomyces, Rhodotorula, Sterigmatomyces and Trichosporon , and included 21 different species. All the isolates were psychrotrophic yeasts. Proteolytic activities of 50 psychrotrophic strains were studied by use of skim milk within the temperature range of 0–42°C. All the strains showed various degrees of proteolysis. In particular, Candida lipolytica. Trichosporon pullulans and Candida scottii were active species at low temperatures. Sensory spoilage due to the proteolytic yeasts were observed in mackerel homogenates stored at 10°C. C. lipolytica -inoculated homogenates caused spoilage with ammoniacal odours after 1 week of storage. Values of total volatile basic nitrogen at 10°C were highest with C. lipolytica among 35 strains tested, followed by Tr. pullans. Trichosporon cutaneum, C. scottii, Rhodotorula glutinis and Cryptococcus luteolus. Proteolytic psychrotrophic yeasts were widely distributed in raw seafoods.     

Isolation of oligotrophic yeasts from supraglacial environments of different altitude on the Gulkana Glacier (Alaska)

Psychrophilic yeasts have been isolated from supra- and subglacial ice at many sites worldwide. To understand the ecology of psychrophilic yeasts on glaciers, we focused on their adaptation to wide range of nutrient concentrations and their distribution with altitude on the Gulkana Glacier in Alaska. We found various culturable psychrophilic yeasts on the ice surfaces of the glacier, and 11 species were isolated with incubation at 4?°C in four different dilutions of agar medium. Some of our isolated species (Rhodotorula psychrophenolica, Rhodotorula aff. psychrophenolica, Rhodotorula glacialis, and Basidiomycota sp. 1) can grow on the low dissolved organic matter (DOC) concentrations medium (7.6?mg?L(-1) ) which is close to the typical level of supraglacial melt water, suggesting that these species can inhabit in any supraglacial meltwater. Otherwise, most of other species were isolated only from higher DOC concentration medium (183?mg?L(-1) -18.3?g?L(-1) ), suggesting that these are inhabitant around the cryoconite, because DOC concentrations in melted surface-ice contained cryoconite is much higher than in melted water. Similarity of altitudinal distribution between culturable yeast and algal biomass suggests that the ecological role played by the cold-adapted yeasts is as organic matter decomposers and nutrient cyclers in glacier ecosystem.     

Alkalitolerant Yeasts from Natural Biotopes

Using a solid nutrient medium containing alkaline buffer (pH 10) and an antibiotic, alkalitolerant yeasts were isolated from samples of soda-rich saline soils (solonchaks) of Armenia (Arazdayan) and the Transbaikal region (the Kungur Steppe). The species diversity of the yeast populations of the tested soda-rich soils was relatively insignificant. They only contained alkalitolerant representatives of asporogenic capsulated yeasts belonging to the species Cryptococcus laurentii, C. albidus, Rhodotorula glutinis, R. mucilaginosa,and Sporobolomyces roseus. C. laurentii representatives clearly dominated the isolates obtained, their number exceeding that of the other species by two to three orders of magnitude. All of the isolates grew on acidic wort agar, suggesting that they did not include obligate alkaliphiles.     

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.     

Biotransformation of polycyclic aromatic hydrocarbons by yeasts isolated from coastal sediments.

Yeast abundance in the sediments of 13 coastal sites in Massachusetts was quantified, and the potential of yeast isolates to biotransform polycyclic aromatic hydrocarbons (PAHs) was determined. Plate counts of yeasts varied between 10(2) to 10(7) CFU g (dry weight) of sediment-1. The most abundant genera isolated and identified included Candida, Cryptococcus, Rhodotorula, Torulopsis, and Trichosporon. More than 50% of the isolates from heavily contaminated sites transformed phenanthrene, as determined by spray-plate screening. The plate counts of phenanthrene-transforming yeasts correlated significantly to the sediment concentrations of phenanthrene. Transformation of [9-14C]phenanthrene and [12-14C]benz[a]anthracene by individual isolates varied greatly, ranging from 0.15 to 8.15 mumol of PAH g-1 in 120-h incubations. Of the isolated yeasts, Trichosporon penicillatum exhibited the greatest capacity for phenanthrene transformation. The ability to transform PAHs appears to be widespread among yeasts in coastal sediments.     

Relative Incidence of Ascomycetous Yeasts in Arctic Coastal Environments

Previous studies of fungi in polar environments have revealed a prevalence of basidiomycetous yeasts in soil and in subglacial environments of polythermal glaciers. Ascomycetous yeasts have rarely been reported from extremely cold natural environments, even though they are known contaminants of frozen foods. Using media with low water activity, we have isolated various yeast species from the subglacial ice of four glaciers from the coastal Arctic environment of Kongsfjorden, Spitzbergen, including Debaryomyces hansenii and Pichia guillermondii, with counts reaching 10⁴ CFU L⁻¹. Together with the basidiomycetes Cryptococcus liquefaciens and Rhodotorula mucilaginosa, these yeasts represent the stable core of the subglacial yeast communities. Other glacial ascomycetous species isolated included Candida parapsilosis and a putative new species that resembles Candida pseudorugosa. The archiascomycete Protomyces inouyei has seldom been detected anywhere in the world but was here recovered from ice in a glacier cave. The glacier meltwater contained only D. hansenii, whereas the seawater contained D. hansenii, Debaryomyces maramus, Pichia guilliermondii, what appears to represent a novel species resembling Candida galli and Metschnikowia bicuspidata. Only P. guilliermondii was isolated from sea ice, while snow/ice in the fjord tidal zone included C. parapsilosis, D. hansenii, P. guilliermondii and Metschnikowia zobellii. All of these isolated strains were characterized as psychrotolerant and xero/halotolerant, with the exception of P. inouyei.     

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.     

云南5个地区戟叶酸模花中酵母菌和类酵母的多样性

[目的]研究云南5个地区(晋宁、祥云、程海、泸沽湖、洱海)的戟叶酸模(Rumex hastatus)花中的酵母菌和类酵母.[方法]采用涂布平板法对5个地区的戟叶酸模花中酵母菌和类酵母进行分离,通过26S rDNA Dl/D2区域序列分析并结合形态观察对分离获得的酵母菌和类酵母进行鉴定;采用胞外酶定性筛选培养基进行产酶筛选;用苏丹黑B染色法筛选产油脂菌株.[结果]从戟叶酸模花中分离得到82株酵母菌和99株类酵母;82株酵母菌鉴定为6个属16个种和1个潜在新种,99株类酵母鉴定为短梗霉属(Aureobasidium)的普鲁兰类酵母(A.pullulans)及3个变种;戟叶酸模花中的优势属是类酵母短梗霉属,其次为红酵母属(Rhodotorula)和隐球酵母属(Cryptococcus);筛选到134株具有产胞外酶活性和83株产油脂的酵母菌和类酵母.[结论]研究结果显示5个地区的戟叶酸模花中酵母菌和类酵母种类多样性较为丰富,并具有产淀粉酶、蛋白酶、纤维素酶、脂肪酶和油脂的特点,有潜在的应用前景.     

Identification and Molecular Analysis of Pathogenic Yeasts in Droppings of Domestic Pigeons in Beijing, China

Feral pigeons are known as reservoirs of pathogenic yeasts that cause opportunistic infections in human. In the outskirts of Beijing, China, pigeons are more frequently raised at homes than are encountered in public areas. Many studies have focused on the presence of pathogenic yeasts in the excreta (fresh or withered) of a variety kinds of birds, pigeon crop and cloacae. One hundred and forty-three samples of fresh droppings were collected from three suburban pigeon-raising homes in an area of northern Beijing, China. The internal transcribed sequences (ITS) of all strains (except for 8 strains of Rhodotorula sp. ) were sequenced and compared with those of the databases of the National Center for Biotechnology Information website ( http://www.ncbi.nlm.nih.gov ) using the Basic Local Alignment Search Tool (BLAST). Yeasts representing 8 genera, Cryptococcus, Filobasidium, Rhodotorula, Candida, Debaryomyces, Saccaromyces, Trichosporon and Sporidiobolus, were identified from 120 isolates. Cryptococcus was the most prolific genera represented by eight species. The populations of yeast species isolated from fresh pigeon droppings were different among homes. Although it is well established that Cryptococcus neoformans exists mainly in old pigeon guano, several C. neoformans strains were still isolated from fresh pigeon excreta, providing a clue that live cryptococcal cells could move through the gastrointestinal tract of the pigeons. Eight genera identified from fresh droppings of domestic pigeons further confirm that pigeons serve as reservoirs, carriers and even spreaders of Cryptococcus species and other medically significant yeasts. The proportion of pathogenic yeasts in all isolates is more than 90 %.     

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.     

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 high Arctic glaciers: the discovery of a new habitat for eukaryotic microorganisms

Recently a new habitat for microbial life has been discovered at the base of polythermal glaciers. In ice from these subglacial environments so far only non-photosynthetic bacterial communities were discovered, but no eukaryotic microorganisms. We found high numbers of yeast cells, amounting to a maximum of 4,000 CFU ml⁻¹ of melt ice, in four different high Arctic glaciers. Twenty-two distinct species were isolated, including two new yeast species. Basidiomycetes predominated, among which Cryptococcus liquefaciens was the dominant species (ca. 90% of total). Other frequently occurring species were Cryptococcus albidus, Cryptococcus magnus, Cryptococcus saitoi and Rhodotorula mucilaginosa. The dominant yeast species were psychrotolerant, halotolerant, freeze-thaw resistant, unable to form mycelium, relatively small-sized and able to utilize a wide range of carbon and nitrogen sources. This is the first report on the presence of yeast populations in subglacial ice.     

Yeast diversity in a mesotrophic lake on the karstic plateau of Lagoa Santa,MG-Brazil

The yeast diversity in a paleo-karstic lake of the Lagoa Santa plateau was studied during March 1986–March 1987. Water samples were collected monthly at five stations and the yeasts were isolated at 25 °C on Sabourad dextrose agar supplemented with 0.5% of yeast extract and 10 mg% of chloramphenicol. August and February showed the highest numbers of isolates, coinciding, respectively with the beginning and the ending of stratification. Of 56 isolated species, Aureobasidium pullulans, Candida famata, Cryptococcus albidus, Cryptococcus laurentii, Rhodotorula glutinis, Rhodotorula rubra and Trichosporon cutaneum occurred at the highest frequencies. A. pullulans showed greatest prevalence during the dry period (winter), whereas C. famata, Cr. albidus, Cr. laurentii and Rh. rubra were prevalent during the rainy season (summer). Tr. cutaneum was distributed between July and November. Most isolates yeasts are associated with plants and soils, and are important in decomposition of aquatic macrophytes. Furthermore, a possible role of these species as indicators of pollution is discussed.     

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.     

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.