Mitochondrial DNA loss caused by ethanol in Saccharomyces flor yeasts.

Saccharomyces flor yeasts proliferate at the surface of sherry wine, which contains over 15% (vol) ethanol. Since ethanol is a powerful inducer of respiration-deficient mutants, this alcohol has been proposed to be the source of the high diversity found in the mitochondrial genomes of flor yeasts and other wine yeasts. Southern blot analysis suggests that mitochondrial DNA (mtDNA) polymorphic changes are due to minor lesions in the mitochondrial genome. As determined in this work by pulsed-field gel electrophoresis, restriction analysis, and Southern blot analysis, ethanol-induced petite mutants completely lack mtDNA (rho zero). Ethanol-induced changes in the mitochondrial genome that could explain the observed mtDNA polymorphism in flor yeasts were not found. The transfer of two different mtDNA variants from flor yeasts to a laboratory strain conferred in both cases an increase in ethanol tolerance in the recipient strain, suggesting that mtDNAs are probably subjected to positive selection pressure concerning their ability to confer ethanol tolerance.     

Effects of small increases in copper levels on culturable basidiomycetous yeasts in low‐nutrient soils

Aims: Investigating the effect of perturbations, with relatively low Cu concentrations, on yeast community composition in low‐nutrient virgin soil. Methods and Results: Culturable soil yeast populations were monitored at an experimental site treated with the fungicide copper oxychloride (10 mg Cu per kg soil). Yeast numbers were unaffected by additional Cu; however, a shift in yeast community composition from Hymenomycetes to Urediniomycetes species occurred. Subsequent growth experiments conducted with a synthetic liquid medium revealed that hymenomycetous and urediniomycetous yeasts were affected differently by 1 and 10 mg l^?1 Cu. Soil microcosm experiments then indicated that additional 10 mg kg^?1 Cu may improve the competitive ability of urediniomycetous yeasts in the presence of hymenomycetous yeasts. Conclusions: The shift from hymenomycetous to urediniomycetous yeasts, as a result of slightly increased soil Cu levels, may be because of hymenomycetous yeasts being more sensitive to elevated Cu levels and urediniomycetous yeasts having an improved competitive ability in the presence of elevated Cu levels. Significance and Impact of the Study: Yeast community composition of pristine low‐nutrient soils may change as a result of perturbations with relatively low concentrations of Cu. Urediniomycetous yeasts should be studied as potential bio‐indicators of Cu perturbations.     

The diversity of Pityrosporum (Malassezia) yeasts in vivo and in vitro

Conclusions In considering the diversity of the lipophilic yeasts it has been shown that in vivo both spherical and oval yeasts may be found in normal conditions on the skin and also associated with hyphae in scales from pityriasis versicolor. There is however generally a different distribution pattern on the body for two forms. This may indicate a different ecology for two distinct varieties or the varying conditions at each site may influence changes in the cell shape of a single species. It is striking that the spherical yeasts (P. orbiculare) have not been found in animals. In vitro, several morphological variants can be maintained, but the change from one form to another which is the subject of a number of reports will be one deciding factor leading to the opinion that P. orbiculare and P. ovale are synonymous. However, physiological differences such as growth rate, their viability in subculture and the analysis of proteins, are all characteristics which alone would be insufficient to support a taxonomic division but when added together confirm the morphological separation of isolates. It remains to be seen if DNA studies, which have so far unified the anthropophilic, lipid dependant Pityrosporum yeasts, will in fact continue to show that they should be confined to a single species.This paper was presented at the X^th congress of the International Society for Human and Animal Mycology at Barcelona, Spain from June 27 to July 1, 1988.     

Fluorescence microscopy procedure for quantitation of yeasts in beverages.

Existing methods for quantitating yeasts in beverages include time-consuming plate counts that detect only living cells and hemacytometer counts that are reliable only at very high concentrations (e.g., 10(6) to 20 X 10(6) cells per ml). The new method described here involves the use of fluorescence microscopy with the fluorescent stain aniline blue to differentiate yeasts (and other fungi) from backgrounds for easy counting and also may be used in conjunction with membrane filtration to concentrate yeasts from liquids before cell enumeration. Recoveries averaged 91.5% for beverages spiked with levels of 500 to 600,000 organisms per ml. The correlation coefficient of count to spike level was 0.996.     

Fluorescence microscopy procedure for quantitation of yeasts in beverages

Existing methods for quantitating yeasts in beverages include time-consuming plate counts that detect only living cells and hemacytometer counts that are reliable only at very high concentrations (e.g., 10(6) to 20 X 10(6) cells per ml). The new method described here involves the use of fluorescence microscopy with the fluorescent stain aniline blue to differentiate yeasts (and other fungi) from backgrounds for easy counting and also may be used in conjunction with membrane filtration to concentrate yeasts from liquids before cell enumeration. Recoveries averaged 91.5% for beverages spiked with levels of 500 to 600,000 organisms per ml. The correlation coefficient of count to spike level was 0.996.     

Establishment of intestinal microbiota with focus on yeasts of unweaned and weaned piglets kept under different farm conditions

This study aimed to characterize the intestinal yeasts in weaning piglets and to establish their possible relationships with main bacterial groups. German Landrace piglets were weaned (WP, n=32) at 28 days of age or kept with the dams until day 39 without creep feed (UP, n=32). The experiment was performed at an experimental and a commercial farm (CF). Faeces were collected from the piglets, sows and pen floors on days 28, 33 and 39 for isolation of DNA and cultivation for enumeration of yeasts, enterobacteria, enterococci and lactobacilli. Fragments of the D1 domain of 26S rRNA gene were amplified and separated by denaturing gradient gel electrophoresis (DGGE). No yeasts could be cultured from water and feed samples. No or only low numbers of yeasts were detected among all UP. In WP at CF, yeasts correlated with lactobacilli (r=0.456; P=0.009) and enterobacteria (r=-0.407; P=0.021). Kazachstania slooffiae dominated among the cultured yeasts. It was the only yeast species detected by PCR-DGGE. Yeasts, especially K. slooffiae, established in the porcine gastrointestinal tract after consumption of grain-based feed and may interrelate with the intestinal microbiota. The study provides data indicating importance of K. slooffiae for the development of balanced porcine gut microbiota.     

Natural transformation of Streptococcus crista

Abstract Most brewing strains of Saccharomyces cerevisiae flocculate following growth in beer wort. However, many do not flocculate in laboratory culture media, because their initial pH and buffering capacity do not correspond to the pH range within which these yeasts flocculate. Many, though not all, NewFlo phenotype brewing yeasts flocculate within a narrow pH range only; this is indicative of the existence of more than one NewFlo flocculation phenotype. Such strains may be flocculated by small alterations of pH to within the flocculation range. Induction of flocculation by pH change may be used to separate cells from media at any stage during fermentation.     

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.     

A novel method for estimating killing ability and digestion of Paracoccidioides brasiliensis by phagocytic cells in vitro

We describe a novel method by which phagocytosis, digestion and killing of Paracoccidioides brasiliensis yeast cells by polymorphonuclear leukocytes or other phagocytic cells may be estimated simultaneously. Suspensions of P. brasiliensis (yeast-like phase) were sonicated, counted and incubated at 37 degrees C with known numbers of phagocytes. Control preparations contained no phagocytic cells. At given intervals samples were incorporated into gelatin nutrient medium and droplets of the mixtures were incubated at room temperature. Live yeast-like P. brasiliensis germinate in vitro and produce filaments. After incubation, droplets may be melted and examined under phase contrast optics, or the cells may be washed and stained by a variation of Papanicolaou's method. Digested P. brasiliensis, intact but non-germinating yeasts and filamented (viable) yeasts may be identified and counted. Killing and digestive abilities of phagocytes may be estimated by the difference between values obtained from phagocyte-containing and control preparations.     

Isolation and Screening of Yeasts That Ferment d-Xylose Directly to Ethanol

Natural habitats of yeasts were examined for the presence of strains able to produce ethanol from d-xylose. Black knots, insect frass, and tree exudates were screened by enrichment in liquid d-xylose-yeast extract medium. These and each d-xylose-assimilating yeast in a collection from cactus fruits and Drosophila spp. were tested for alcohol production from this sugar. Among the 412 isolates examined, 36 produced more than 1 g of ethanol liter from 20 g of d-xylose liter, all under aerated conditions. Closer examination of the strains indicated that their time courses of d-xylose fermentation followed different patterns. Some strains produced more biomass than ethanol, and among these, ethanol may or may not be assimilated rapidly after depletion of d-xylose. Others produced more ethanol than biomass, but all catabolized ethanol after carbohydrate exhaustion. Ethanol production appeared best at low pH values and under mild aeration. Possible correlations between the nutritional profiles of the yeasts and their ability to produce ethanol from d-xylose were explored by multivariate analysis. d-Xylose appeared slightly better utilized by yeasts which rate poorly in terms of fermentation. The fermentation of d-glucose had no bearing on d-xylose fermentation. No specific nutritional trait could discriminate well between better d-xylose fermentors and other yeasts.     

Formation of Dulcitol in Aerobic Dissimilation of d-Galactose by Yeasts

During the study of aerobic dissimilation of galactose by yeasts, polyhydric products were isolated in crystalline form from the fermented broths and identified. Yeast species may be divided into two groups on basis of sugar alcohol production: type I yeasts form the same end products from galactose as from glucose; type II yeasts produce dulcitol from galactose with or without other sugar alcohols but they produce no dulcitol from glucose. Isolation of dulcitol from microorganism has not been previously described.     

Affinity Purifications of Aldose Reductase and Xylitol Dehydrogenase from the Xylose-Fermenting Yeast Pachysolen tannophilus

Although xylose is a major product of hydrolysis of lignocellulosic materials, few yeasts are able to convert it to ethanol. In Pachysolen tannophilus, one of the few xylose-fermenting yeasts found, aldose reductase and xylitol dehydrogenase were found to be key enzymes in the metabolic pathway for xylose fermentation. This paper presents a method for the rapid and simultaneous purification of both aldose reductase and xylitol dehydrogenase from P. tannophilus. Preliminary studies indicate that this method may be easily adapted to purify similar enzymes from other xylose-fermenting yeasts.     

Yeasts Isolated from Neotropical Wood-Boring Beetles in SE Peru

Some temperate wood-boring cerambycid beetles harbor intracellular gut yeasts believed to augment host nutrition, but species belonging to the subfamily Lamiinae are thought to lack endosymbionts. Almost 49 percent of Neotropical cerambycid species are lamiines, therefore, comparatively few rain forest species would be expected to host symbiotic gut yeasts. This study reports the isolation of gut yeasts from closely related Neotropical lamiines. We investigated species that feed on trees in the Brazil nut family (Lecythidaceae), because host plant associations are relatively well known. Our objectives were to determine if gut yeasts were present and, if possible, infer their mode of transmission. We collected and dissected 18 beetle specimens from three tree species, including 17 cerambycids and one curculionid. Every insect specimen yielded a gut yeast. DNA sequence libraries were used for a rapid identification of the yeasts and their larval hosts. The cerambycids included five lamiine species and one cerambycine. Six ascomycete yeasts were isolated from their guts; we found no evidence of strict vertical transmission. Larval gut yeasts were genetically similar to yeasts previously isolated from insects associated with wood or fungi, implying potential habitat specificity. The yeasts have not yet been localized, and potential function is not known, but they may contribute to rapid nutrient cycling or serve as the first line of defense against plant toxins.     

Analysis of homothallic Saccharomyces cerevisiae strain mating during must fermentation

Genetic instability and genome renewal may cause loss of heterozygosity (LOH) in homothallic wine yeasts (Saccharomyces cerevisiae), leading to the elimination of the recessive lethal or deleterious alleles that decrease yeast fitness. LOH was not detected in genetically stable wine yeasts during must fermentation. However, after sporulation, the heterozygosity of the new yeast population decreased during must fermentation. The frequency of mating between just-germinated haploid cells from different tetrads was very low, and the mating of haploid cells from the same ascus was favored because of the physical proximity. Also, mating restriction between haploid cells from the same ascus was found, leading to a very low frequency of self spore clone mating. This mating restriction slowed down the LOH process of the yeast population, maintaining the heterozygote frequency higher than would be expected assuming a fully random mating of the haploid yeasts or according to the Mortimer genome renewal proposal. The observed LOH occurs because of the linkage of the locus MAT to the chromosome III centromere, without the necessity for self spore clone mating or the high frequency of gene conversion and rapid asymmetric LOH observed in genetically unstable yeasts. This phenomenon is enough in itself to explain the high level of homozygosis found in natural populations of wine yeasts. The LOH process for centromere-linked markers would be slower than that for the nonlinked markers, because the linkage decreases the frequency of newly originated heterozygous yeasts after each round of sporulation and mating. This phenomenon is interesting in yeast evolution and may cause important sudden phenotype changes in genetically stable wine yeasts.     

Analysis of Homothallic Saccharomyces cerevisiae Strain Mating during Must Fermentation

Genetic instability and genome renewal may cause loss of heterozygosity (LOH) in homothallic wine yeasts (Saccharomyces cerevisiae), leading to the elimination of the recessive lethal or deleterious alleles that decrease yeast fitness. LOH was not detected in genetically stable wine yeasts during must fermentation. However, after sporulation, the heterozygosity of the new yeast population decreased during must fermentation. The frequency of mating between just-germinated haploid cells from different tetrads was very low, and the mating of haploid cells from the same ascus was favored because of the physical proximity. Also, mating restriction between haploid cells from the same ascus was found, leading to a very low frequency of self spore clone mating. This mating restriction slowed down the LOH process of the yeast population, maintaining the heterozygote frequency higher than would be expected assuming a fully random mating of the haploid yeasts or according to the Mortimer genome renewal proposal. The observed LOH occurs because of the linkage of the locus MAT to the chromosome III centromere, without the necessity for self spore clone mating or the high frequency of gene conversion and rapid asymmetric LOH observed in genetically unstable yeasts. This phenomenon is enough in itself to explain the high level of homozygosis found in natural populations of wine yeasts. The LOH process for centromere-linked markers would be slower than that for the nonlinked markers, because the linkage decreases the frequency of newly originated heterozygous yeasts after each round of sporulation and mating. This phenomenon is interesting in yeast evolution and may cause important sudden phenotype changes in genetically stable wine yeasts.     

Niche construction initiates the evolution of mutualistic interactions

Niche construction theory explains how organisms' niche modifications may feed back to affect their evolutionary trajectories. In theory, the evolution of other species accessing the same modified niche may also be affected. We propose that this niche construction may be a general mechanism driving the evolution of mutualisms. Drosophilid flies benefit from accessing yeast‐infested fruits, but the consequences of this interaction for yeasts are unknown. We reveal high levels of variation among strains of Saccharomyces cerevisiae in their ability to modify fruits and attract Drosophila simulans. More attractive yeasts are dispersed more frequently, both in the lab and in the field, and flies associated with more attractive yeasts have higher fecundity. Although there may be multiple natural yeast and fly species interactions, our controlled assays in the lab and field provide evidence of a mutualistic interaction, facilitated by the yeast's niche modification.     

Pulmonary cryptococcosis due to a capsule-deficient strain confused with metastatic lung cancer

A patient with hepatocellular cancer developed pulmonary cryptococcosis due to infection with a capsule-deficient Cryptococcus neoformans. Pulmonary lesions initially diagnosed as metastatic cancer by chest x-ray film and CT scan were subsequently found to be fungal granulomas by autopsy. Although morphologic studies of the fungi were insufficient to render a specific mycologic diagnosis because of the absence of encapsulated yeasts, fluorescent antibody studies confirmed the diagnosis of cryptococcosis. The use of various stains and electron microscopy for the pathological differential diagnosis of cryptococcosis caused by capsule-deficient yeasts is discussed. This revised version was published online in August 2006 with corrections to the Cover Date.     

Gut-associated yeast in bark beetles of the genus Dendroctonus Erichson (Coleoptera: Curculionidae: Scolytinae)

Scolytine bark beetles are the most destructive pests of conifers; they sometimes aggregate in such large numbers that they actually kill their hosts. They maintain close relationships with yeasts and fungi, in particular those that are assumed to aid in digestive, detoxification processes and pheromone production. In this study, 403 yeast strains were isolated from the guts, ovaries, eggs and frass of nine bark beetle species in the genus Dendroctonus Erichson. The beetles were collected from 10 conifer species at 34 locations in Mexico, Guatemala and the USA. Yeast identification was based on partial DNA sequences from 18S rDNA, 26S rDNA and internal transcribed spacer (ITS1), as well as morphological and physiological characteristics. A combined phylogenetic analysis delimited 11 clades with sequences similar to Candida arabinofermentans , C. ernobii , C. membranifaciens (including C. lessepsii , Pichia mexicana and P. scolyti ), C. oregonensis , C. piceae , Kuraishia capsulata (including K. capsulata and K. cf. molischiana ), Pichia americana , P. canadensis , P. glucozyma , P. guilliermondii and an undescribed species of Candida . Nucleotide divergences between the major clades were at least 5% while, with the exception of 30 isolates, yeasts within clades differed from named reference species at fewer than 1% of the nucleotide sites. There do not appear to be obligate relationships between particular yeasts and specific anatomical partitions, nor between particular yeasts and bark beetle species. Some yeasts do appear to be preferentially associated with bark beetles feeding on different conifer genera and therefore host plant defences may limit yeast community diversity in Dendroctonus .  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 98 , 325–342.     

Relation of the radiosensitivity of yeast cells to the LET of the radiation. Effect of additional growth and its contribution to resistance and the shape of the survival curve of diploid cells

A study was made of the contribution of the additional growth effect to the survival rate of Saccharomyces cerevisiae diploid yeasts and its influence on the survival curve shape after exposure to ionizing radiation of different linear energy transfer (LET). The effect of the additional growth was shown to be the only factor responsible for a sigmoid shape of the survival curve of diploid yeasts affected by high LET radiation. As LET increased the contribution of the additional growth effect to the survival rate decreased: this was due to the pattern of cell distribution by the number of radiation lesions that altered with alterations in LET.     

The vectoring of cactophilic yeasts by Drosophila

Summary At two locations in the Sonoran Desert, yeasts were sampled from species of Drosophila, the flies' cactus hosts, and other neighboring sources of cactophilic yeasts to determine the relation between the yeasts vectored by the fly and the yeasts found in their breeding sites. D. mojavensis, D. nigrospiracula, and D. mettleri vectored yeast assemblages significantly more similar to the yeast species found on the rot from which the flies were collected than to the yeasts found on other rots from the flies host cactus or other rotting cactus at the same site. Rots with Drosophila had fewer yeast species than those without flies, suggesting that flies were associated with younger rots. Rots with flies and the Drosophila also had more yeast species with the capability to produce ethyl acetate than rots without flies. The results support the contention that cactophilic Drosophila feed on a subset of the yeasts available in an area, and may act to maintain differences among the yeast communities found on different species of cactus.