Pure Yeast Cultures Research Articles

Conceptual framework for the assessment of the nutritional and biological value of the plant Galega orientalis (LAM)

Conducted a comprehensive comparative study of the nutritional and biological value Galega orientalis (Lam): the solubility of the protein in vitro – Galega’s seeds, leaves, hay, which was harvested in the phase of budding – beginning of flowering. It is believed to be the essential influence of feed type on the solubility of the protein. According to the conducted studies it has revealed that the solubility of the feed protein is in the range from 37.0% to 50.0%. The rate of solubility of Galega orientalis (Lam) seed protein is 50.0%, litestock – 40.0%, and hay – 37.0% from its total. The research results indicate that the protein of Galega orientalis (Lam) refers to the feed with the average level of solubility. It has researched qualitative assessment of the biologically active substances content in organs of Galega orientalis’ (Lam) plants. It has identified the highest content of flavonoids and saponins in rosette leaves and ascorbic acid in the organs of plants and petals. There is the high content of tannins found in the petals of Galega orientalis (Lam). There is a high content of coumarins and water-soluble polysaccharides in the roots of the studied plants. Phytochemical studies of the plant Galega orientalis (Lam) indicates the absence in it of cardiac glycosides and antrahlikozydiv. It was first conducted testing on antimicrobial activity of a plant Galega orientalis (Lam) on the growth pure cultural of bacteria gram positive (Micrococcus luteus), gram negative (Escherichia coli XL1, DH5) and yeasts (Saccharomyces cerevisiae W303). The material for the study was dried vegetative mass Galega orientalis (Lam) in the phase of budding and early flowering. As a result of the experiment was revealed that 20% concentration of aqueous extract of this plant had inhibitory effects on the growth of pure cultures of bacteria and yeast. It has grounded reasonably possible relationship between the antimicrobial activity of the extract of this plant and the lack of bloating in cows. It has outlined prospects of further researches of this problem.

Improved ethyl caproate production of Chinese liquor yeast by overexpressing fatty acid synthesis genes with OPI1 deletion.

During yeast fermentation, ethyl esters play a key role in the development of the flavor profiles of Chinese liquor. Ethyl caproate, an ethyl ester eliciting apple-like flavor, is the characteristic flavor of strong aromatic liquor, which is the best selling liquor in China. In the traditional fermentation process, ethyl caproate is mainly produced at the later fermentation stage by aroma-producing yeast, bacteria, and mold in a mud pit instead of Saccharomyces cerevisiae at the expense of grains and fermentation time. To improve the production of ethyl caproate by Chinese liquor yeast (S. cerevisiae) with less food consumption and shorter fermentation time, we constructed three recombinant strains, namely, α5-ACC1ΔOPI1, α5-FAS1ΔOPI1, and α5-FAS2ΔOPI1 by overexpressing acetyl-CoA carboxylase (ACC1), fatty acid synthase 1 (FAS1), and fatty acid synthase 2 (FAS2) with OPI1 (an inositol/choline-mediated negative regulatory gene) deletion, respectively. In the liquid fermentation of corn hydrolysate, the contents of ethyl caproate produced by α5-ACC1ΔOPI1, α5-FAS1ΔOPI1, and α5-FAS2ΔOPI1 increased by 0.40-, 1.75-, and 0.31-fold, correspondingly, compared with the initial strain α5. The contents of other fatty acid ethyl esters (FAEEs) (C8:0, C10:0, C12:0) also increased. In comparison, the content of FAEEs produced by α5-FAS1ΔOPI1 significantly improved. Meanwhile, the contents of acetyl-CoA and ethyl acetate were enhanced by α5-FAS1ΔOPI1. Overall, this study offers a promising platform for the development of pure yeast culture fermentation of Chinese strong aromatic liquor without the use of a mud pit.

Raman spectroscopy and determination of soil yeasts

In this paper, two isolates of yeasts, members of the genus Candida, have been examined using Raman spectroscopy. The yeasts were isolated from soils sampled at the town park in Tivat (Montenegro) and experimental farm Radmilovac (Serbia), by the accumulation method. The pure cultures of yeasts were identified by the API 20C AUX system. The presence of Candida guilliermondii and Candida utilis was noted. The Raman spectra, originating from lipids, amides, proteins, carbohydrates, aromatic amino acids and the nucleotide bases, were analyzed in the wide range of the wave numbers from 500 to 3200 cm-1. It was noticed that peaks which correspond to the lipid fraction were more pronounced in the case of Candida guilliermondii compared to Candida utilis. This may represent a specific response of the yeast species to stress conditions.

Non-Alcoholic, Naturally-Carbonated Beverage from Vitis Vinifera Using Saccharomyces Cerevisae Isolated from Cheese Whey

In the present work a pure yeast culture was isolated from cheese whey and identified by 26S rDNA sequencing as Saccharomyces cerevisiae NRRL Y-12632. This yeast was used to produce low alcoholic self carbonated beverage from grape juice, grape: Kinnow juice (2:1) and grape: Pomegranate juice (2:1) under optimized conditions. The physicochemical properties of 3 days fermented grape: Pomegranate beverage (2:1) were pH 3.87, TSS 15°B, CO2 content 0.09 bar, acidity 0.24%, sugar-acid ratio 63.34% and alcohol 0.06% (v/v). The changes in the physicochemical properties recorded were pH 3.08, TSS 14.8°B, CO2 content 0.14 bar, acidity 0.35%, sugar-acid ratio 42.04% and alcohol 0.10% (v/v) after storage for 30 days at 4°C without adding any preservative. Among the four beverages, grape: Pomegranate beverage (2:1) was highly acceptable when compared on 9 point hedonic scale. Naturally produced CO2 was found to give tangy taste to the beverage along with antimicrobial properties. This technology can be a useful processing technology for perishable fruits and vegetables and can also help to overcome their seasonal overabundance.

COMPARATIVE ASSESSMENT OF THE LABORATORY SELECTED AND ACTIVE DRIED SACCHAROMYCES CEREVISIAE YEAST CULTURE IN BIOTECHNOLOGY OF THE BRANDY PRODUCTION

<p>Samples from different industrial grape cultivars were collected during the vintage season from the vineyard of the winery (the «Shabo» winery Company, located in the Odesa region, Ukraine). The following industrial cultivars of grapes were selected for the research: Chardonnay, Cabernet Sauvignon, Merlot, Sauvignon, Riesling Rhenish, Aligote, Rkatsiteli, Bastardo, Traminer, Telti Kuruk, Grinosh.</p> <p>The grape cultivars were cultivated on the sandy soils in the district located between the Black Sea and the Dnestrovsky estuary. Grape must derived from different grape cultivars was placed into sterile glass flasks to half of the 450ml flask volume. Each flask was carefully closed with a rubber stopper with an injection needle in it. During the fermentation process, it was necessary to remove carbon dioxide, which was present as a result of active anaerobic fermentation processes in the grape must. At the end of grape must fermentation, pure yeast cultures were isolated using traditional microbiological methods by consistent inoculation of a sample into a Petri dish with a few modifications of nutrient selective agar for yeast isolation and cultivation. Primary yeast isolation was carried out using Inhibitory Mold Agar medium (Becton Dickinson Company, USA).</p> <p>The yeast culture morphological properties were analyzed after the primary yeast culture isolation. Yeasts were identified by polymerase chain reaction (PCR) using universal yeast primers. After yeast culture identification, the next step in yeast cultivation was carried out on Wort Agar medium (Becton Dickinson Company, USA). Each isolated, and identified yeast culture was deposited in the Genebank of Japan, MAFF culture Collection, Tsukuba, Ibaraki, Japan and (NCYC) - Yeast Culture Collection (National Collection of Yeast Cultures, Institute of Food Research, Norwich, United Kingdom). Each yeast culture was tested for technological characteristics such as growth resistance to high temperature (+42°C) and low temperature (+6°C), growth at low pH 2.6–3.0 (acid resistance), growth in the presence of 5, 10, and 15% ethanol (ethanol resistance), and growth in the presence of high concentration potassium bisulfite (bisulfite resistance). Hydrosulfide synthesis (H<sub>2</sub>S gassing production) was studied in addition.</p> <p>Parameters of cellular metabolism in yeast suspension, such as concentration of nitrogen, protein, triglicerides, enzymatic activity and total sugar (which include glucose, fructose, and galactose) were determined. Macro- and micro-element concentrations in fermented grape must, which contained pure yeast culture was determined and included: potassium, sodium, calcium, phosphorus, magnesium, iron, chlorides. In addition to identifying parameters of macro- and micro- element concentration in grape must during and following fermentation based on a principle of photometric analysis, carried out using a biochemical analyser Respons-920 (DiaSys Diagnostic Systems GmbH, Germany).</p> <p>Laboratory selected <em>Saccharomyces cerevisiae </em>wine yeast showed high enzymatic activity with short lag phase. Since of fermentation started on third day concentration of Triglicerides, Protein (total), Potassium and Sodium increased and then level of Protein (total) on the 5th day of fermentation twice decreased. Trigliceride concentration on the 5th day of fermentation continued to increase. Concentration of Iron on the 5th day of fermentation increase in geometrical progression, concentration increase in 4-5 times. Contrary Chloride concentration on the 5th day of fermentation decreased in 3-4 times. Enzymatic activity on 3rd day of fermentation maximal for Lactate Dehydrogenase, Alanine aminotransferase, Aspartate aminotransferase, Phosphatase. Since of 5th day of fermentation Enzymatic activity for Lactate Dehydrogenase, Alanine aminotransferase, Aspartate aminotransferase 3-4 times. Especially level of Phosphatase activity very decreased in 6-7 times. Comparative assessment between our Laboratory selected <em>Saccharomyces cerevisiae</em> yeast culture and Dry active commercial <em>Saccharomyces cerevisiae</em> yeast culture did not showed any difference in enzymatic activity. Both groups showed high enzymatic activity on the third day from the start of fermentation and decreasing on the fifth day since of fermentation started.</p> <p><em> Key words: wine yeast, enzymatic activity, cellular metabolism</em><em>, </em><em>Saccharomyces cerevisiae</em><em>.</em><em></em></p>

Genetic and physiological characterization of yeast isolated from ripe fruit and analysis of fermentation and brewing potential

‘Wild’ and spontaneously fermented beers are growing in popularity in the craft beer industry. Most of these beers are fermented by the use of either pure cultures of unconventional yeast and bacteria or spontaneous fermentation using mixed local microflora. This study examined the potential of using pure strains of new isolates of wild yeast in the fermentation of a unique beer. The microbial communities from the fruit of pindo palm, loquat, hackberry and blackberry were collected in liquid culture, then plated for isolation. Ten isolates were selected for further analysis. Strains were identified by restriction fragment length polymorphism (RFLP) analysis and analysed for growth in a simple liquid media, fermentation in a complex media, alcohol tolerance and acid tolerance. Despite identification of some strains as the same species, they displayed a wide range of physiological properties. All strains were tolerant of pH values as low as 2.4, but none were tolerant of pH 1.9. Alcohol tolerance of different strains varied from 6 to 12%. Several strains had properties that suggest potential as primary fermenters, including the alcohol fermentation of a beer wort. Organoleptic properties of beers fermented with several of the strains demonstrated potential for commercial brewing. Copyright © 2014 The Institute of Brewing & Distilling

The isolation of novel marine yeasts; a new procedure

The recent research on marine yeasts highlighted that they are able to produce many bioactive substances including enzymes, amino acids, killer toxins, and vitamin C with many potential applications in food, pharmaceutical, fuel, cosmetics and chemical industries [1]. However, the employment of marine yeast in research and industry is limited due to the lack of suitable isolation methods. Current methods suffer from fungus interference and/or low number of yeast isolates. In this report, a novel 3-step isolation method has been developed. Samples were taken from seawater, sea sand or seaweed which could potentially contain marine yeasts. These samples were enriched using a nutrient rich media with antibiotics for three cycles as the first step. In the second step, single colonies with different morphologies were isolated on plates of agar medium lacking antibiotic after 24–48h of incubation at 30 ◦C. These isolates were further confirmed to be pure cultures of yeast by streaking them on agar plates and checking their purity and morphology under a microscope. In this study, 14 samples from different marine habitats in Egypt, UK and USA were used for the isolation. 122 isolates were obtained and confirmed to be pure yeast cultures. Our 3-step isolation method for marine yeast has demonstrated high level of efficiency comparing with two reference methods.

Dynamic model‐based analysis of furfural and HMF detoxification by pure and mixed batch cultures of S. cerevisiae and S. stipitis

Inhibitory compounds that result from biomass hydrolysis are an obstacle to the efficient production of second-generation biofuels. Fermentative microorganisms can reduce compounds such as furfural and 5-hydroxymethyl furfural (HMF), but detoxification is accompanied by reduced growth rates and ethanol yields. In this study, we assess the effects of these furan aldehydes on pure and mixed yeast cultures consisting of a respiratory deficient mutant of Saccharomyces cerevisiae and wild-type Scheffersomyces stipitis using dynamic flux balance analysis. Uptake kinetics and stoichiometric equations for the intracellular reduction reactions associated with each inhibitor were added to genome-scale metabolic reconstructions of the two yeasts. Further modification of the S. cerevisiae metabolic network was necessary to satisfactorily predict the amount of acetate synthesized during HMF reduction. Inhibitory terms that captured the adverse effects of the furan aldehydes and their corresponding alcohols on cell growth and ethanol production were added to attain qualitative agreement with batch experiments conducted for model development and validation. When the two yeasts were co-cultured in the presence of the furan aldehydes, inoculums that reduced the synthesis of highly toxic acetate produced by S. cerevisiae yielded the highest ethanol productivities. The model described here can be used to generate optimal fermentation strategies for the simultaneous detoxification and fermentation of lignocellulosic hydrolysates by S. cerevisiae and/or S. stipitis.

Multi-enzyme production by pure and mixed cultures of Saccharomyces and non-Saccharomyces yeasts during wine fermentation

Saccharomyces and non-Saccharomyces yeasts release enzymes that are able to transform neutral compounds of grape berries into active aromatic compounds, a process that enhances the sensory attributes of wines. So far, there exists only little information about enzymatic activity in mixed cultures of Saccharomyces and non-Saccharomyces during grape must fermentations. The aim of the present work was to determine the ability of yeasts to produce extracellular enzymes of enological relevance (β-glucosidases, pectinases, proteases, amylases or xylanases) in pure and mixed Saccharomyces/non-Saccharomyces cultures during fermentation. Pure and mixed cultures of Saccharomyces cerevisiae BSc562, Hanseniaspora vinae BHv438 and Torulaspora delbrueckii BTd259 were assayed: 1% S. cerevisiae/99% H. vinae, 10% S. cerevisiae/90% H. vinae, 1% S. cerevisiae/99% T. delbrueckii and 10% S. cerevisiae/90% T. delbrueckii. Microvinifications were carried out with fresh must without pressing from Vitis vinifera L. c.v. Pedro Jiménez, an autochthonous variety from Argentina. Non-Saccharomyces species survived during 15–18days (BTd259) or until the end of the fermentation (BHv438) and influenced enzymatic profiles of mixed cultures. The results suggest that high concentrations of sugars did not affect enzymatic activity. β-Glucosidase and pectinase activities seemed to be adversely affected by an increase in ethanol: activity diminished with increasing fermentation time. Throughout the fermentation, Saccharomyces and non-Saccharomyces isolates assayed produced a broad range of enzymes of enological interest that catalyze hydrolysis of polymers present in grape juice. Vinifications carried out by a pure or mixed culture of BTd259 (99% of T. delbrueckii) showed the highest production of all enzymes assayed except for β-glucosidase. In mixed cultures, S. cerevisiae outgrew H. vinae, and T. delbrueckii was only detected until halfway the fermentation process. Nevertheless, their secreted enzymes could be detected throughout the fermentation process.Our results may contribute to a better understanding of the microbial interactions and the influence of some enzymes on vinification environments.

Competition between species can stabilize public-goods cooperation within a species.

Competition between species is a major ecological force that can drive evolution. Here, we test the effect of this force on the evolution of cooperation within a species. We use sucrose metabolism of budding yeast, Saccharomyces cerevisiae, as a model cooperative system that is subject to social parasitism by cheater strategies. We find that when cocultured with a bacterial competitor, Escherichia coli, the frequency of cooperator phenotypes in yeast populations increases dramatically as compared with isolated yeast populations. Bacterial competition stabilizes cooperation within yeast by limiting the yeast population density and also by depleting the public goods produced by cooperating yeast cells. Both of these changes induced by bacterial competition increase the cooperator frequency because cooperator yeast cells have a small preferential access to the public goods they produce; this preferential access becomes more important when the public good is scarce. Our results indicate that a thorough understanding of species interactions is crucial for explaining the maintenance and evolution of cooperation in nature.

Potential spoilage non-Saccharomyces yeasts in mixed cultures with Saccharomyces cerevisiae

With the aim of exploring the possibility to improve wine quality through the utilization of wine-related yeasts generally considered as spoilage, mixed cultures of Saccharomyces cerevisiae with Hanseniaspora osmophila, Pichia fermentans, Saccharomycodes ludwigii and Zygosaccharomyces bailii were inoculated in grape juice. All the fermentations got to completion and most of the compounds normally produced at high concentrations by pure cultures of non-Saccharomyces yeasts, and considered detrimental for wine quality, did not reach the threshold taste level in mixed fermentations with S. cerevisiae. Interestingly, the association of S. cerevisiae with P. fermentans, S. ludwigii and Z. bailii produced significant increases in the production of polysaccharides as compared to pure cultures of S. cerevisiae. Since polysaccharides improve wine taste and body, and exert positive effects on aroma persistence and protein and tartrate stability, a possible use for these yeasts can be envisaged in mixed starter cultures with S. cerevisiae for the enhancement of the final quality of wine.

Use of variability of the fungal ITS2 intergenic region for the identification of medically important yeast species

AbstractYeasts are common fungal opportunistic pathogens in humans playing a significant role in the morbidity and mortality of immunocompromised patients. Number of resistant yeast species is responsible for infections and consequent infectious complications, but the final microbiological diagnosis can be affected by variability of their phenotype and may result in incorrect identification. For the purposes of this study, advanced applications of molecular genetic methods based on certain up-to-date knowledge of fungal internal transcribed spacer (ITS) regions have been employed, which could support a possibility of universal application of such methods for identification of any pure yeast culture. In this study, the targeted DNA was amplified by a couple of primers, and the products of PCR reaction were divided by capillary electrophoresis. In the cases, in which the measured sizes of fragments did not correspond with the anticipated sizes, fragments were used for the sequencing analysis and compared to the nucleotide databases using the BLAST tool. Out of 208 isolates, 7.2% (n = 16) of cases occurred to be incorrectly determined, particularly in the group of non-albicans Candida species accounting for as many as 21.7%.

Application of pure and mixed probiotic lactic acid bacteria and yeast cultures for oat fermentation

AbstractFermentation of a prebiotic containing oat substrate with probiotic lactic acid bacteria and yeast strains is an intriguing approach for the development of new synbiotic functional products. This approach was applied in the present work by using pure and mixed microbial cultures to ferment a heat‐treated oat mash. Results show that the strains studied were appropriate for oat fermentation and the process could be completed for 6–10 h depending on the strain. The viable cell counts achieved within this time were above the required levels of 106–107 cfu ml−1 for probiotic products. Both single lactic acid bacteria strains and mixed cultures of the same strains with yeast were found suitable for oat fermentation. However, the pure LAB cultures attributed better flavour and shelf life of the oat drinks. The content of the prebiotic oat component beta‐glucan remained within 0.30–0.36% during fermentation and storage of the drinks obtained with each of the strains used. Thus, these products would contribute diet with the valuable functional properties of beta‐glucan. Also, the viability of pure and mixed cultures in the oat products was good: levels of cell counts remained above the required numbers for probiotic products throughout the estimated shelf‐life period. Copyright © 2005 Society of Chemical Industry

Alcohols, esters and heavy sulphur compounds production by pure and mixed cultures of apiculate wine yeasts

Strains of Hanseniaspora uvarum, Hanseniaspora guilliermondii and Saccharomyces cerevisiae were used as pure or mixed starter cultures in commercial medium, in order to compare their kinetic parameters and fermentation patterns. In pure and mixed cultures, yeasts presented similar ethanol yield and productivity. Pure cultures of H. uvarum and S. cerevisiae showed a specific growth rate of 0.38 h −1; however, this value decreased when these yeasts were grown in mixed cultures with H. guilliermondii. The specific growth rate of pure cultures of H. guilliermondii was 0.41 h −1 and was not affected by growth of other yeasts. H. guilliermondii was found to be the best producer of 2-phenylethyl acetate and 2-phenylethanol in both pure and mixed cultures. In pure cultures, H. uvarum led to the highest contents of heavy sulphur compounds, but H. guilliermondii and S. cerevisiae produced similar levels of methionol and 2-methyltetrahydrothiophen-3-one. Growth of apiculate yeasts in mixed cultures with S. cerevisiae led to amounts of 3-methylthiopropionic acid, acetic acid-3-(methylthio)propyl ester and 2-methyltetrahydrothiophen-3-one similar to those obtained in a pure culture of S. cerevisiae; however, growth of apiculate yeasts increased methionol contents of fermented media.

Contaminant yeast detection in industrial ethanol fermentation must by rDNA-PCR

The present work focuses on the possibility to use conserved primers that amplify yeast ITS1-5.8S-ITS2 ribosomal DNA locus (rDNA) to detect the presence of non-Saccharomyces cerevisiae yeast in fermentation must of bioethanol fermentation process. Total DNA was extracted from pure or mixed yeast cultures containing different cell concentrations and different contaminant/fermenting yeast concentrations and submitted to PCR. Upon improvement of detection limits and DNA extraction protocol, must samples of distillery were checked for the presence of contaminant yeast. Contaminant rDNA bands were detected only in industrial samples during contamination episodes, but not in noncontaminated must. The method described here could detect the presence of contaminant yeast from industrial must in eight hours after sampling. The improved procedure may help to avoid severe contamination episodes at fermentation industries by decreasing the detection time from 5 days to 8 h and possible quantification of contaminant yeasts that can impose economical loss to the process.

Degradation of HMW Glutenins During Wheat Sourdough Fermentations

ABSTRACTBakeries use sourdoughs to improve bread properties such as flavor and shelf life. The degradation of gluten proteins during fermentation may, however, crucially alter the gluten network formation. We observed changes that occurred in the HMW glutenins during wheat sourdough fermentations. As fermentation starters, we used either rye sourdough or pure cultures of lactobacilli and yeast. In addition, we incubated wheat flour (WF) in the presence of antibiotics under different pH conditions. The proteolytic activities of cereal and sourdough‐derived proteinases were studied with edestin and casein. During sourdough fermentations, most of the highly polymerized HMW glutenins degraded. A new area of alcohol‐soluble proteins (≈30.000 MW) appeared as a result of the proteolytic breakdown of gluten proteins. Very similar changes were observable as WF was incubated in the presence of antibiotics at pH 3.7. Cereal and sourdough‐derived proteinases hydrolyzed edestin at pH 3.5 but showed no activity at pH 5.5. An aspartic proteinase inhibitor (pepstatin A) arrested 88–100% of the activities of sourdough enzymes. According to these results, the most active proteinases in wheat sourdoughs were the cereal aspartic proteinases. Acidic conditions present in sourdoughs create an ideal environment for cereal aspartic proteinases to be active against gluten proteins.

Fermentation behaviour of controlled mixed and sequential cultures of Candida cantarellii and Saccharomyces cerevisiae wine yeasts

Behaviour of Candida cantarellii and Saccharomyces cerevisiae strains during the fermentation of Syrah grape must using pure, mixed and sequential yeast cultures was studied. Different kinds of inocula have been tested according to the type of culture. Inocula proportions used in mixed C. cantarellii and S. cerevisiae strains reflect the population levels in natural grape microbiota. Biomass evolution of both strains was analysed in relation to different byproduct levels. Saccharomyces cerevisiae overcame C. cantarellii in the different co-culture assays at 48 h of fermentation. The final concentration of ethanol was similar in mixed and both sequential tests and higher (from 7.8 to 10.6%) than in S. cerevisiae pure culture. In mixed and sequential cultures, the glycerol content of the final products was 44.3 to 52.8% higher than the one obtained with pure S. cerevisiae fermentation. Wine analytical profiles of experiments that involved S. cerevisiae and C. cantarellii strains differed from the pure ones mainly in acetoin, propanol and succinic acid contents. From an enological point of view, analysed byproducts are relevant. Considering this, mixed assay and the inoculation of S. cerevisiae after 3 days of pure C. cantarellii fermentation appear to be the more appropriate options to develop the particular characteristics of Syrah wines.

A genetic study of natural flor strains of Saccharomyces cerevisiae isolated during biological ageing from Sardinian wines.

In this study, three flor strains of Saccharomyces cerevisiae were genetically characterized. They were isolated from biofilms on Sardinian sherry-like wines produced at family-run wineries where pure cultures of yeasts were not used. The study aimed to investigate the life cycle of these naturally-occurring flor strains, using a genetic procedure supplemented by analysis of subsequent meiotic generations. A semi-homothallic life cycle was found in three strains that could be helpful in a genetic improvement programme.

Response of the aroma fraction in sherry wines subjected to accelerated biological aging.

The effect of an acceleration assay, carried out with a periodic aeration and an increased surface/volume ratio, on various aroma compounds of "fino" Sherry wines aging under a veil of a pure culture of Saccharomyces cerevisiae race capensis G1 flor film yeast was studied. The results were subjected to multifactor analysis of variance, and the compounds simultaneously depending on acceleration conditions and aging time at p < 0.01 were subjected to principal component analysis. The first component, accounting for 86.14% of the overall variance, was mainly defined by acetaldehyde and its derivatives 1,1-diethoxyethane and acetoin. These compounds reached higher concentrations in accelerated aging wines in a shorter time than they did in control wines, and no browning problems were detected. Taking into account that these compounds can be used as indicators for biological aging of "fino" Sherry wines, the acceleration condition assayed can be applied to shorten the time of this process.

Rapid detection of microbial contamination in grape juice by flow cytometry

&lt;p style="text-align: justify;"&gt;This study presents an application of flow cytometry to evaluate rapidly the viable micro-organisms in grape juice. In this method, viable cells are firstly specitically labelled with a fluorescent reagent. The sample is then injected into the flow cytometer where the labelled micro-organisms are individually illuminated by a laser beam. The emission of fluorescence is measured. The system counts the number of fluorescent events and prints out a histogram of the fluorescence intensity which is characteristic of the micro-organism being analysed. In laboratory conditions, preliminary trials have been undertaken with an artificially inoculated grape juice with pure yeast and bacteria cultures. This method succeeded in counting simultaneously yeasts and bacteria within 15 minutes, with a high degree of sensitivity, 5.10&lt;sup&gt;3&lt;/sup&gt; yeasts perml and 5.10&lt;sup&gt;4&lt;/sup&gt; bacteria per ml. This technique can also be applied to the detection of mould contamination and the test has been done with &lt;em&gt;Botrytis&lt;/em&gt; spores. The method makes direct cell counts possible and is capable of analysing 30 samples per hour. It can be automatised and easily used in industrial laboratory. During the last harvest, more than a thousand of must samples were controled using this technique. The results let to determine the yeast contamination level of a grape juice tank even before unloading. The results obtained by flow cytometry were compared to the plate count reference method. The correlation between cytometry and count by plate culture was 99 p. cent for the threshold of 5.1 0&lt;sup&gt;4&lt;/sup&gt; yeasts/ml which seemed to point out a high contamination. By using this flow cytometry method during the harvest period, the results were supplied in real time. This allowed a rapid selection of the musts, depending upon the scale of their contamination and improved the quality of the wine by corrective actions.&lt;/p&gt;