Isolation and characteristics of the lactose-fermenting yeasts Candida kefyr

Screening of lactose-fermenting yeast strains has been conducted among 162 strains isolated from various plants and 28 strains isolated from cheese. Four yeast strains fermented lactose and were identified as Candida kefyr. The specific β-galactoside activity of the studied strains was 1501–2113 IU/g dry biomass. The ability of strains C. kefyr C24 and C30 to produce ethanol from lactose was significantly inhibited by the increase in substrate concentration (100 g/L).     

Selection of lactose-fermenting yeast for ethanol production from whey

Summary Candida pseudotropicalis ATCC 8619 was selected among nine strains of lactose-fermenting yeasts on the basis of its ability to ferment concentrated whey. In 28% (wt/vol) deproteinized whey solutions it produced an average of 12.4% (vol/vol) ethanol. This yeast could be used in a process for whey treatment.     

Selection of methods of detecting lactose-fermenting Salmonella strains and evaluating their usefulness for diagnostic studies

Salmonella rods of subspecies I, lactose-fermenting were first isolated in Poland in 1980. They were isolated from a plus sample taken from a brain abscess of a child. Next strains were isolated from faeces of newborn and hospitalized children. Growth characteristic of colonies of lactose-fermenting Salmonella strains on selective-differentiating media (Mac Conkey's Levine, SS, So?tys) recommended for inoculation of clinical material resembled Escherichia coli. So far these type of colonies were omitted in diagnostic examinations. Lactose-fermenting variants showed on Bismuth sulfate agar "Difco" (WB) typical for Salmonella growth pattern. They grew on this medium after 48 hr of incubation in a form of black, medium sized colonies, with some metallic brilliance and characteristic blackening of the medium undercolonies. Precise knowledge of biochemical properties of lactose-fermenting Salmonella allows to supplement so far used diagnostic scheme with additional tests permitting differentiation of lactose-fermenting variants of Salmonella from the other members of Enterobacteriaceae family. Taking into consideration biochemical variants in diagnostic procedure i.e. lactose-fermenting Salmonella, allowedns to isolate in the years 1983-1985 lactose-positive strains in 1305 out of 2773 (47%) individuals positive for S. agona. In 1987, 246 persons (28.3%) out of 869 with lactose-fermenting Salmonella of various serotypes were simultaneously infected with lactose-negative variant. Lactose-fermenting strains of Salmonella belonged most frequently to the following genera: S. agona, S. enteritidis, S. oranienburg, S. typhimurium, and S. goldcoast. It was found that the modified diagnostic procedure makes possible the isolation and the identification of lactose-positive varians of Salmonella.     

Selection and fermentation properties of cryophilic wine yeasts

Two cryophilic strains, YM-84 and YM-126, were selected by a double-layer agar fermenting technique from 100 strains of the wine yeast, Saccharomyces cerevisiae. The viability (specific growth rate) and fermentability of the two selected strains at low temperatures (7 and 13°C) were superior to those of wine yeast strains W3 and OC-2, indicating the usefulness of the two strains as cryophilic wine yeasts. Experiments using the two selected strains at intermediate temperatures (22 and 30°C) showed that their fermentation ceased prematurely and their ethanol yields were reduced.     

Selection of distiller's yeasts with particular respect to non-saccharomyces strains

The investigations demonstrate that in addition to Saccharomyces also strains Candida and Hansenula can be used for ethanol production. Their efficiences are at the standard level and the first phase of fermentation is considerably accelerated with results in suppression of bacterial contamination in cold mashing. In the future efficiency optimisations will be expected based on mixed strain populations as well as on technological improvement. Furthermore, genetic modifications present actually a real prospect to secure “made-to-measure” strains for special processes of corn mash fermentation.     

A taxonomic study of soil yeasts

Summary Eighty-four samples of Minnesota soils were collected in the spring of the year. All samples yielded yeasts, and a total of 180 cultures were isolated of which 117 were studied taxonomically. Twenty-five cultures were black yeasts. Approximately one-third of the isolates were spore forming yeasts and members of the oxidative and film forming generaHansenula, Pichia, andDebaryomyces, of which a new species (D. mrakii) has been described. Not a single culture ofSaccharomyces was isolated. Perhaps later in the season when fruit was abundant members of this genus would have been present.The greatest number of imperfects belonged to the genusCandida (32 cultures), althoughPullularia (25 cultures),Rhodotorula (20 cultures), andTorulopsis (16 cultures) were well represented.The cultures ofPullularia darkened very slowly if at all, in the asbence of excess sugar. Under these circumstances they were very similar toT. pullulans.A single culture ofTrichisporon cerebriformis was isolated. The culture isolated formed both blastospores and arthrospores.I would like to express my appreciation to Dr.C. E. Skinner of Washington State College, for his help and guidance throughout this entire problem. I would also like to thank Dr.E. M. Mark, of the University of California, Berkeley, for the privilege of working under his direction, and in his laboratory during the study.     

Comparative study on synteny between yeasts and vertebrates

We studied synteny conservation between 18 yeast species and 13 vertebrate species in order to provide a comparative analysis of the chromosomal plasticity in these 2 phyla. By computing the regions of conserved synteny between all pairwise combinations of species within each group, we show that in vertebrates, the number of conserved synteny blocks exponentially increases along with the divergence between orthologous protein and that concomitantly; the number of genes per block exponentially decreases. The same trends are found in yeasts but only when the mean protein divergence between orthologs remains below 36%. When the average protein divergence exceeds this threshold, the total number of recognizable synteny blocks gradually decreases due to the repeated accumulation of rearrangements. We also show that rearrangement rates are on average 3-fold higher in vertebrates than in yeasts, and are estimated to be of 2 rearrangements/Myr. However, the genome sizes being on average 200 times larger in vertebrates than in yeasts, the normalized rates of chromosome rearrangements (per Mb) are about 50-fold higher in yeast than in vertebrate genomes.     

Selection in yeast I: Assessing genetic stability and relative fitness of commercial yeasts

Summary The potential for changes in allele frequencies in yeast populations by selection was examined. Cells from the wine yeastSaccharomyces cerevisiae (strain Montrachet) were grown over a large number of generations using two different culturing techniques, each with two variations: serial transfers on WLN agar plates with and without UV irradiation, and continuous culture in autoclaved and in filter-sterilized grape must. A low frequency of variant isozyme patterns was found in samples taken at the end of the experiment. Growth rates in must and on agar plates were also examined, and it was found that all samples were faster-growing than the original strain, to varying degrees. Applications for the selection system developed are discussed.     

Novel enrichment technique for the isolation of highly potent catalase producing yeasts from soil1

Among the 252 soil isolates screened, about 40% were catalase-positive. A novel soil enrichment culture technique consisting of feeding decreasing concentrations of a rich nutrient medium along with increasing concentrations of H2O2 (up to 15% v/v) in a semi- continuous glass column reactor over 15 days resulted in the isolation of high catalase producing yeasts identified as Saccharomyces cerevisiae and Schizosaccharomyces pombe. These yeasts produced 2–3 times more of intracellular catalase (1630 to 2277 U/ml) than the other microbial isolates (15 to 820 U/ml) tested. The other microorganisms gradually disappeared as the concentration of H2O2 increased in the enrichment. The technique could be used for the isolation of exclusively yeasts that might have superior industrial properties such as high ethanol, flavour, protein and lipid production.     

Comparative study on the identification of food-borne yeasts

Morphologically distinct yeast colonies from partially and fully processed fruits and vegetables were isolated over a 3-year period. Identification of 239 strains was achieved by using standard methods, commercial identification kits (API 20C and API YEAST-IDENT), and a simplified system for food-borne yeasts. The identified strains of fruit origin represented 36 species belonging to 19 genera. Among strains of vegetable origin, 34 species representing 17 genera were identified. The simplified identification system and the conventional method provided the same results in 80% of the cases. The commercial identification kits were easy to use but were not appropriate for food-borne yeast species. Computer-assisted identification was helpful.     

Selection and preliminary characterization of β-glycosidases producer Patagonian wild yeasts

Four pre-selected indigenous yeast strains belonging to Candida guilliermondii (V₂ and V₅), Candida pulcherrima (V₆) and Kloeckera apiculata (V₉), were used as β-glucosidase (βGL) and β-xylosidase (βXL) sources. The optimization of yeast culture conditions was carried out and the effects of oenological parameters on β-glycosidase activities were evaluated. C. guilliermondii V₂ and C. pulcherrima V₆ strains were selected. These strains showed intracellular (C. pulcherrima V₆) and parietal (C. guilliermondii V₂) constitutive βGL and βXL. The enzymatic activities were active at pH, glucose, ethanol and SO₂ concentrations usually found in winemaking and they were able to release monoterpenols and alcohols from grape juice glycoside extracts. Additionally, these yeast strains were not able to produce volatile acidity and off flavour. Regional ecological relevance of these species was also discussed. Our results evidence that the selected C. guilliermondii V₂ and C. pulcherrima V₆ strains have interesting oenological characteristics and allow us to think in their potential application in winemaking.     

Selection and hybridization of wine yeasts for improved winemaking properties: Fermentation rate and aroma productivity

Three strains out of thirty-one wine yeasts (Saccharomyces cerevisiae) were selected for their good winemaking properties (fermentation rate, tolerance of sulfur dioxide, aroma productivity, wine quality) and genetic markers (KHR killer activity, galactose assimilation), and used for hybridization by spore to spore mating. Of the twelve hybrids produced, two, Hy17-108 (RIFY 1001 × RIFY 1067) and Hy41-308 (RIFY 1001 × RIFY 1065), were selected on the basis of a fermentation test. In experimental winemaking the two hybrids demonstrated improved aroma productivity for higher alcohols, aromatic esters and/or fatty acids, while their fermentation rate was nearly the same as that of the parental strains.     

Possible interference of lactose-fermenting marine vibrios in coliform -D-galactosidase assays

C.M. DAVIES, S.C. APTE AND S.M. PETERSON. 1995. An investigation into possible interferences in β-D-galactosidase-based assays for coliform bacteria in marine waters was carried out. A rapid instrumental fluorescence assay for β-D-galactosidase activity, using 4-methylumbelliferyl-β-D-galactosidase as a substrate, was used to investigate activities of this enzyme in non-coliform bacterial isolates from coastal waters. Only 2% of isolates showed slight enzyme activity after a 1-h incubation period at 44.5βC. At a lower incubation temperature of 20βC, 51% and 94% of the isolates showed some enzyme activity within 6 h and 48 h, respectively. Fifty-nine out of 67 of these isolates were identified as Vibrio species. A lac⁺ strain of Vibrio vulnificus was found to produce β-D-galactosidase which caused significant false-positive reactions in the Colilert-Marine Water assay when present at concentrations of 10 cfu ml⁻¹ or greater. This interference could be overcome by addition of the vibriostatic agent O/129. The high fluorescence of this reagent, however, precluded the simultaneous determination of Escherichia coli in the Colilert test and also its use in instrumental fluorescence assays. It was concluded that in assays employing high temperatures and short incubation times, Vibrio species are unlikely to cause significant interferences.     

Distribution of Vibrio vulnificus and other lactose-fermenting vibrios in the marine environment

During the summer of 1981, 3,887 sucrose-negative vibrios were isolated from seawater, sediment, plankton, and animal samples taken from 80 sites from Miami, Fla., to Portland, Maine. Of these, 4.2% were able to ferment lactose. The lactose-positive strains isolated from the various samples correlated positively with pH and turbidity of the water, vibrios in the sediment and oysters, and total bacterial counts in oysters. Negative correlations were obtained for water salinity. Numerical taxonomy was performed on 95 of the lactose-fermenting environmental isolates and 23 reference strains. Five clusters resulted, with the major cluster containing 33 of the environmental isolates and all of the Vibrio vulnificus reference strains. The 33 isolates, which produced an acid reaction in lactose broth within hours of initial inoculation, represented 20% of all lactose-fermenting vibrios studied. These isolates were nearly identical phenotypically to clinical strains of V. vulnificus studied by the Centers for Disease Control, Atlanta, Ga., and by our laboratory, and their identification was confirmed by DNA-DNA hybridization studies. V. vulnificus was isolated from all sample types and from Miami to Cape Cod, Mass., and comparison of the environmental parameters of the eight subsites yielding this species with those of all 80 subsites revealed no significant differences. The majority of the isolates were obtained from animals, with clams providing most (84%) of these. On injection into mice, 82% of the V. vulnificus isolates resulted in death. Members of the remaining four clusters contained strains which differed from V. vulnificus in such phenotypic traits as luminescence and in urease or H(2)S production. None of the other reference cultures, including nine other Vibrio species, were contained in the remaining clusters, and these isolates could not be identified. Most of these were also lethal for mice. Phenotypic differences, potential pathogenicity, and geographic distribution of the five clusters were examined. It is concluded that V. vulnificus is a ubiquitous organism, both geographically and in a variety of environmental sources, although it occurs in relatively low numbers. The public health significance of this organism and of the other unidentified lactose-fermenting Vibrio species is discussed.     

Selection and identification of autochthonous yeasts in Slovakian wine samples using a rapid and reliable three-step approach

Aims: The investigation of yeast microflora during the must fermentation of two wine varieties (Frankovka modra – Blaufränkisch and Veltlinske zelene – Grüner Veltliner) from two consecutive vintages was performed using a three‐step approach. Methods and Results: The investigation strategy consisted of the combination of yeast cultivation, selection of the isolated yeasts based on the amplification of internal transcribed spacer 2 using a fluorescence‐labelled primer (f‐ITS‐PCR) and a final identification step based on amplification and sequencing of the ITS1‐5.8S rDNA‐ITS2 region of the selected yeasts. By this three‐step approach, it was possible to screen 433 yeasts isolates that belonged to 13 different species. Conclusions: The f‐ITS‐PCR allowed the unambiguous differentiation of all isolated yeast species that produced their typical f‐ITS‐PCR profile. Significance and Impact of the Study: This is one of few reports that treat the yeast diversity in Slovakian wines and in two varieties largely cultivated in Central Europe. The three‐step approach permitted the rapid and reliable identification of isolated yeasts. The f‐ITS‐PCR with its good discrimination power can represent a suitable molecular tool for the selection of yeast members recovered from food or other environments.