Corrigendum: Flor Yeast Diversity and Dynamics in Biologically Aged Wines

International audience; [This corrects the article DOI: 10.3389/fmicb.2018.02235.].

Influence of nitrogen status in wine alcoholic fermentation

Nitrogen is an essential nutrient for yeast during alcoholic fermentation. Nitrogen is involved in the biosynthesis of protein, amino acids, nucleotides, and other metabolites, including volatile compounds. However, recent studies have called several mechanisms that regulate its role in biosynthesis into question. An initial focus on S. cerevisiae has highlighted that the concept of "preferred" versus "non-preferred" nitrogen sources is extremely variable and strain-dependent. Then, the direct involvement of amino acids consumed in the formation of proteins and volatile compounds has recently been reevaluated. Indeed, studies have highlighted the key role of lipids in nitrogen regulation in…

Tyrosine-containing peptides are precursors of tyramine produced by Lactobacillus plantarum strain IR BL0076 isolated from wine

Abstract Background Biogenic amines are molecules with allergenic properties. They are found in fermented products and are synthesized by lactic acid bacteria through the decarboxylation of amino acids present in the food matrix. The concentration of biogenic amines in fermented foodstuffs is influenced by many environmental factors, and in particular, biogenic amine accumulation depends on the quantity of available precursors. Enological practices which lead to an enrichment in nitrogen compounds therefore favor biogenic amine production in wine. Free amino acids are the only known precursors for the synthesis of biogenic amines, and no direct link has previously been demonstrated between …

Design and Performance Testing of a DNA Extraction Assay for Sensitive and Reliable Quantification of Acetic Acid Bacteria Directly in Red Wine Using Real Time PCR

International audience; Although strategies exist to prevent AAB contamination, the increased interest for wines with low sulfite addition leads to greater AAB spoilage. Hence, there is a real need for a rapid, specific, sensitive, and reliable method for detecting these spoilage bacteria. All these requirements are met by real time Polymerase Chain Reaction (or quantitative PCR; qPCR). Here, we compare existing methods of isolating DNA and their adaptation to a red wine matrix. Two different protocols for isolating DNA and three PCR mix compositions were tested to select the best method. The addition of insoluble polyvinylpolypyrrolidone (PVPP) at 1% (v/v) during DNA extraction using a pro…

Color Stabilization of Apulian Red Wines through the Sequential Inoculation of Starmerella bacillaris and Saccharomyces cerevisiae

Mixed fermentation using Starmerella bacillaris and Saccharomyces cerevisiae has gained attention in recent years due to their ability to modulate the qualitative parameters of enological interest, such as the color intensity and stability of wine. In this study, three of the most important red Apulian varieties were fermented through two pure inoculations of Saccharomyces cerevisiae strains or the sequential inoculation of Saccharomyces cerevisiae after 48 h from Starmerella bacillaris. The evolution of anthocyanin profiles and chromatic characteristics were determined in the produced wines at draining off and after 18 months of bottle aging in order to assess the impact of the different f…

Polyphenols in kombucha: Metabolomic analysis of biotransformations during fermentation

Kombucha is a non-alcoholic beverage made of sugared tea that is transformed by a symbiotic consortium of yeasts and bacteria. This beverage is increasingly produced at industrial scale, but its quality standards remain to be defined. Metabolomics analysis was carried out using FT-ICR-MS (Fourier Transform-Ion Cyclotron Resonance-Mass Spectrometry) to understand the chemical transformations induced by the production phases and the type of tea on the non-volatile compounds of kombucha.

Relationship between ethanol tolerance, lipid composition and plasma membrane fluidity inSaccharomyces cerevisiaeandKloeckera apiculata

The lipid composition of a strain of each of two yeasts, Saccharomyces csrevisiae and Kloeckera apiculata, with different ethanol tolerances, was determined for cells grown with or without added ethanol. An increase in the proportion of ergosterol, unsaturated fatty acid levels and the maintenance of phospholipid biosynthesis seemed to be responsible for ethanol tolerance. The association of ethanol tolerance of yeast cells with plasma membrane fluidity, measured by fluorescence anisotropy, is discussed. We propose that an increase in plasma membrane fluidity may be correlated with a decrease in the sterol: phospholipid and sterol: protein ratios and an increase in unsaturation index.

Wine microbiome : A dynamic world of microbial interactions

International audience; Most fermented products are generated by a mixture of microbes. These microbial consortia perform various biological activities responsible for the nutritional, hygienic, and aromatic qualities of the product. Wine is no exception. Substantial yeast and bacterial biodiversity is observed on grapes, and in both must and wine. The diverse microorganisms present interact throughout the winemaking process. The interactions modulate the hygienic and sensorial properties of the wine. Many studies have been conducted to elucidate the nature of these interactions, with the aim of establishing better control of the two fermentations occurring during wine processing. However, …

Microbial Dynamics between Yeasts and Acetic Acid Bacteria in Kombucha: Impacts on the Chemical Composition of the Beverage.

Kombucha is a traditional low-alcoholic beverage made from sugared tea and transformed by a complex microbial consortium including yeasts and acetic acid bacteria (AAB). To study the microbial interactions and their impact on the chemical composition of the beverage, an experimental design with nine couples associating one yeast strain and one AAB strain isolated from original black tea kombucha was set up. Three yeast strains belonging to the genera Brettanomyces, Hanseniaspora, and Saccharomyces and three strains of Acetobacter and Komagataeibacter species were chosen. Monocultures in sugared tea were analyzed to determine their individual microbial behaviors. Then, cultivation of the ori…

Using CRISPR/Cas platform for Genetic Modification of Commercial Saccharomyces cerevisiae strains

International audience; Traditional wine fermentation is a complex microbial process initiated by various yeast species classified as Saccharomyces and non-Saccharomyces species.To better understand the different interactions occurring within wine fermentations and track a specific yeast population, we wish to obtain GFP-tagged yeast cells that stably expres fluorescence signal without compromising the fermentative capability of the strain.To this end, the CRISPR/Cas system was investigated to genetically modify the commercial Saccharomyces Saccharomyces cerevisiae diploid strain Lalvin EC 1118

Sorption of wine volatile phenols by yeast lees

The capacity of Saccharomyces cerevisiae yeast lees to sorb 4-ethylguaiacol and 4-ethylphenol was investigated in a synthetic medium and in wine. Active dried yeast was more effective when volatile phenols were diluted in red wine. Partition coefficients between wine model solution and wine yeast lees were determined and compared with those measured for dried active yeast. They showed a larger affinity of volatile phenols for wine yeast lees than for dried active yeast. The effect of yeast lees on volatile phenol sorption was sensitive to yeast autolysis level and to physicochemical parameters, such as ethanol content, temperature and pH. These results could be applied in the technology of …

Non-Botrytis grape-rotting fungi responsible for earthy and moldy off-flavors and mycotoxins

Abstract The grape microflora is complex and includes filamentous fungi, yeasts and bacteria with different physiological characteristics and effects on wine production. Most studies have focused on the wine microbiota, but a few studies have reported the ecology of grape microorganisms. Some of these organisms — such as non-Botrytis bunch rotting fungi, which greatly influence the safety or sensory quality of wine, due to the production of mycotoxins and off-flavors, respectively — are considered to be spoilage agents. We review here the diversity of filamentous fungi on grapes and the factors influencing their development, such as grape ripening stage, environmental factors (climate, rain…

Yeast autolysis in sparkling wine – a review

Sparkling wine produced by the traditional methode champenoise requires a second in-bottle alcoholic fermentation of a base wine, leading to the sparkling wine. This second fermentation is followed by prolonged ageing in contact with yeast cells (lees). The autolysis of yeast occurs during the ageing of sparkling wines. During this process, the yeast releases different compounds that modify the organoleptic properties of the wine. The ageing period is required to give these wines their roundness and characteristic aroma and flavour. Autolysis products also influence the foaming properties of sparkling wine. Yeast autolysis is characterised by the hydrolysis of intracellular biopolymers by y…

Biogenic amines in fermented foods

Food-fermenting lactic acid bacteria (LAB) are generally considered to be non-toxic and non-pathogenic. Some species of LAB, however, can produce biogenic amines (BAs). BAs are organic, basic, nitrogenous compounds, mainly formed through decarboxylation of amino acids. BAs are present in a wide range of foods, including dairy products, and can occasionally accumulate in high concentrations. The consumption of food containing large amounts of these amines can have toxicological consequences. Although there is no specific legislation regarding BA content in many fermented products, it is generally assumed that they should not be allowed to accumulate. The ability of microorganisms to decarbox…

Comparative morphological characteristics of three Brettanomyces bruxellensis wine strains in the presence/absence of sulfur dioxide

International audience; The red wine spoilage yeast Brettanomyces bruxellensis has been the subject of numerous investigations. Some of these studies focused on spoilage mechanisms, sulfur dioxide tolerance and nutrient requirements. Pseudomycelium formation, although a striking feature of this species, has however been poorly investigated. Furthermore, literature regarding the induction mechanism of pseudomycelium formation in this yeast is limited and lacks clarity, as results published are contradictory. This study elucidates this phenomenon among strains from geographically different areas. Potential environmental cues were investigated, to attain a better understanding of this mechanis…

PCR ITS-RFLP: A useful method for identifying filamentous fungi isolates on grapes.

Restriction digestion analysis of the ITS products was tested as an easy method to identify isolates of filamentous fungi on grapes. Endonucleases SduI, HinfI, MseI, HaeIII were used. Endonucleases BfmI, Cfr9I, Hpy188I, MaeII or PspGI were used as necessary to complete discrimination. The 43 species studied generated 42 different composite profiles. Only the species P. thomii and P. glabrum gave the same composite profile. 96.3% strains tested could be differentiated to the species level with only four enzymes. Hundred ninety nine strains of filamentous fungi were isolated from various vineyards in Burgundy and identified by this method. Penicillium (58.5%) was the genus the most frequently…

The yeast Starmerella bacillaris (synonym Candida zemplinina) shows high genetic diversity in winemaking environments.

International audience; The yeast Candida zemplinina (Starmerella bacillaris) is frequently isolated from grape and wine environments. Its enological use in mixed fermentation with Saccharomyces cerevisiae has been extensively investigated these last few years, and several interesting features including low ethanol production, fructophily, glycerol and other metabolites production, have been described. In addition, molecular tools allowing the characterization of yeast populations have been developed, both at the inter- and intraspecific levels. However, most of these fingerprinting methods are not compatible with population genetics or ecological studies. In this work, we developed 10 micr…

An improved protocol for electroporation ofOenococcus oeniATCC BAA-1163 using ethanol as immediate membrane fluidizing agent

Aims:  To finalize an effective and reproducible electroporation procedure to transform Oenococcus oeni ATCC BAA-1163 strain. Methods and Results:  The vector pGID052 was selected to optimize the electroporation procedure. Transformation efficiency was 5·8 × 103 per μg of DNA. Transformation was improved when competent cells were prepared with exponential phase cultures; optimum electroporation parameters were an electric pulse of 12·5 kV cm−1, under a resistance of 200 Ω and the presence of 10% (v/v) ethanol in the electroporation buffer (EPB). Conclusions:  An effective protocol to transform O. oeni ATCC BAA-1163 strain by electroporation has been obtained by addition of ethanol to the EP…

Viable But Not Culturable (VBNC) state of Brettanomyces bruxellensis in wine: New insights on molecular basis of VBNC behaviour using a transcriptomic approach

International audience; The spoilage potential of Brettanomyces bruxellensis in wine is strongly connected with the aptitude of this yeast to enter in a Viable But Non Culturable (VBNC) state when exposed to the harsh wine conditions. In this work, we characterized the VBNC behaviour of seven strains of B. bruxellensis representing a regional intraspecific biodiversity, reporting conclusive evidence for the assessment of VBNC as a strain-dependent character. The VBNC behaviour was monitored by fluorescein diacetate staining/flow cytometry for eleven days after addition of 0.4, 0.6, 0.8, 1 and 1.2 mg/L of molecular SO2 (entrance in the VBNC state) and after SO2 removal (exit from the VBNC st…

Characterization of the Viable but Nonculturable (VBNC) State in Saccharomyces cerevisiae

The Viable But Non Culturable (VBNC) state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable populat…

The aroma glycosides composition of burgundy Pinot noir must

Research Note

Notations et écarts de rentabilité : le marché français avant l'euro

The main task of this paper is to confront two classical measures of default risk of the issuer, the rating and the spread. The first is attributed by agencies specialized in this activity (Standard and Poor's or Moody's) while the second results directly from the market price of the bond. This article studies this link over a period of two years for about forty French denominated bonds. Two measures of the spread are used and the results obtained show the very partial consideration of this information by the investors on the French bond market.

Origines des levures Saccharomyces et non-Saccharomyces présentes dans le moût : Vignoble ou cuverie ?

International audience; Les levures de l’espèce Saccharomyces cerevisiae ont longtemps été considérées comme les seules responsables du processus de fermentation. On sait aujourd’hui que différentes espèces de levures Saccharomyces et non-Saccharomyces (NS) sont présentes dans les moûts et qu’un certain nombre d’entre elles participe à la fermentation alcoolique (FA). Bien qu’il soit aujourd’hui largement admis que les principales sources de levures dans le moût de raisin sont les baies de raisin et le matériel de vinification (Fleet et Heard, 1993 ; Mortimer et Polsinelli, 1999) (figure 1), la part apportée par chacune de ces deux sources est encore mal définie. De plus, d’autres sources p…

Oligopeptide assimilation and transport by Oenococcus oeni

International audience; Aims: Oenococcus oeni is a slow-growing wine bacterium with a low growth yield. It thrives better on complex nitrogen sources than on free amino-acid medium. We aimed to characterize the oligopeptide use of this micro-organism. Methods and Results: Several peptides of two to eight amino-acid residues were able to provide essential amino acids. The disappearance of various peptides from extracellular medium was assessed with whole cells. Initial rates of utilization varied with the peptide, and free amino acids were released into the medium. Conclusions: Oenococcus oeni was able to transport the oligopeptides with two to five amino-acid residues tested and to hydrolys…

Hydrolysis and sorption by Saccharomyces cerevisiae strains of Chardonnay grape must glycosides during fermentation

Abstract Different Saccharomyces cerevisiae strains were tested for their ability to hydrolyse glycosyl–glucose (G–G) compounds present in Chardonnay must during alcoholic fermentation. The determination of glycosyl–glucose during the course of the alcoholic fermentation of Chardonnay grape must revealed that S. cerevisiae is able to hydrolyse G–G during alcoholic fermentation in wine conditions. The extent of hydrolysis vary from 17 to 57% of the initial glycoside concentration. In order to confirm that decrease in G–G concentration during alcoholic fermentation is due to hydrolytic action and not to sorption phenomenon, in the first experiment, an analysis of the osidic part of Chardonnay…

Effect of ethanol on membrane fluidity of protoplasts fromSaccharomyces cerevisiae andKloeckera apiculata grown with or without ethanol, measured by fluorescence anisotropy

Direct measurements of membrane fluidity by fluorescence anisotropy of protoplasts fromKloeckera apiculata andSaccharomyces cerevisiae, a low and a high ethanol tolerant strain respectively, are presented. The comparison of the behaviour of the two strains grown with or without ethanol enabled us to demonstrate the existing relationship between ethanol tolerance and membrane fluidity.

Wine Yeast Terroir: Separating the Wheat from the Chaff-for an Open Debate.

Wine terroir is characterized by a specific taste and style influenced by the cultivar of the fermented grapes, geographical factors such as the vineyard, mesoclimate, topoclimate, and microclimate, soil geology and pedology, and the agronomic approach used. These characteristics together define the concept of “terroir”. Thus, regional distinctive flavors in wine have been the subject of many studies aimed at better understanding the link between the wine and the vineyard. Indeed, the identification of key environmental elements involved in the regional variation of grape and wine quality characteristics is a critical feature for improving wine production in terms of consumer preference and…

Efficiency of French privatizations: a dynamic vision

The program of French privatizations is one of the principal worldwide programs as for the volume of the equity issues. A reading of the process of privatization through the corporate governance theory resulted in working out a model making it possible to take into account, on the one hand, the time dimension of the process of privatization, on the other hand, the contextual, organizational, governance and strategic variables which influence this process. After having replicated a certain number of traditional tests, we carried out a test of this model on a sample of 19 French privatized firms and on a seven years horizon, which made it possible to obtain the following conclusions. The favo…

Effect of ethanol on yeast film formation

<p style="text-align: justify;">In this study, we have investigated the influence of ethanol on yeast film formation and cell surface hydrophobicity (CSH). A yeast strain (P3) previously isolated from film yeast was grown in a medium containing increasing ethanol concentration ranging from 0 to 14 p. cent (v/v). It results from this study that up to 10 p. cent ethanol, the greater was the ethanol concentration, the greater was the growth of film. Using two different techniques (phase partition method, magnobead assay), we have shown that ethanol altered the CSH of the yeast. The measured hydrophobicity (p. cent) of cells grown without ethanol was 65 p. cent compared with 81 p. cent wi…

Vitamins in wine: Which, what for, and how much?

Vitamins are essential compounds to yeasts, and notably in winemaking contexts. Vitamins are involved in numerous yeast metabolic pathways, including those of amino acids, fatty acids, and alcohols, which suggests their notable implication in fermentation courses, as well as in the development of aromatic compounds in wines. Although they are major components in the course of those microbial processes, their significance and impact have not been extensively studied in the context of winemaking and wine products, as most of the studies focusing on the subject in the past decades have relied on relatively insensitive and imprecise analytical methods. Therefore, this review provides an extensi…

Biochemical aspects of stuck and sluggish fermentation in grape must

Recently a number of studies have focused on the factors responsible for the occurrence of stuck and sluggish fermentations. Results from these studies indicate that together with nutritional deficiencies and inhibitory substances, technological practices could lead to such situations. This review explains, from a biochemical point of view, the influence of nutritional deficiencies, inhibitory substances and technological practices on yeast cell development and physiology and the fermentation process.

Yeast–yeast interactions revealed by aromatic profile analysis of Sauvignon Blanc wine fermented by single or co-culture of non-Saccharomyces and Saccharomyces yeasts

International audience; There has been increasing interest in the use of selected non-Saccharomyces yeasts in co-culture with Saccharomyces cerevisiae. The main reason is that the multistarter fermentation process is thought to simulate indigenous fermentation, thus increasing wine aroma complexity while avoiding the risks linked to natural fermentation. However, multistarter fermentation is characterised by complex and largely unknown interactions between yeasts. Consequently the resulting wine quality is rather unpredictable. In order to better understand the interactions that take place between non-Saccharomyces and Saccharomyces yeasts during alcoholic fermentation, we analysed the vola…

Molecular Cloning, Heterologous Expression, and Characterization of Ornithine Decarboxylase from Oenococcus oeni

International audience; Ornithine decarboxylase (ODC) is responsible for the production of putrescine, the major biogenic amine found in wine. Oenococcus oeni is the most important lactic acid bacterium in the winemaking process and is involved in malolactic fermentation. We report here the characterization of ODC from an O. oeni strain isolated from wine. Screening of 263 strains isolated from wine and cider from all over the world revealed that the presence of the odc gene appears to be strain specific in O. oeni. After cloning, heterologous expression in Escherichia coli, and characterization, the enzyme was found to have a molecular mass of 85 kDa and a pI of 6.2 and revealed maximal ac…

Characterization of EprA, a major extracellular protein of Oenococcus oeni with protease activity

International audience; Extracellular proteins from Oenococcus oeni. a wine-making bacterium, were isolated during growth on media differing by their nitrogen content. Analysis by two-dimensional electrophoresis revealed a low number of protein signals. Among the main spots, one signal corresponded to a single protein, which contained a lysine repeat domain characteristic of cell-wall hydrolases. We demonstrated that this major protein, named EprA, was able to hydrolyse several proteins. The heterologous production of this protein in Escherichia coli confirmed the protease activity of EprA. With a MW of 21.3 kDa and a pl of 5.3, EprA presents optimal activity at pH 7.0 and 45 degrees C. Thi…

Diversité des Brettanomyces et de leur résistance au SO2. Les nouvelles avancées vers une meilleure gestion du SO2 en vinification.

International audience; Des recherches ont été menées par le Groupe National « Lutte contre Brettanomyces » et plus particulièrement sur la relation SO2 et Brettanomyces bruxellensis afin d’approfondir les connaissances sur le comportement de la levure et d’apporter des données essentielles à une bonne gestion du risque.Une grande diversité de la levure Brettanomyces a été mise en évidence (identification de différents groupes génétiques) ainsi que des comportements différents vis-à-vis du SO2 : sensibles, tolérants ou résistant. Grâce à la mise au point d’un outil prédictif (TYP \ Brett), les professionnels pourront connaître le groupe génétique pour mieux intervenir.Ces travaux ont égalem…

Yeasts and Sparkling Wine Production

Sparkling wines are produced in many different countries. Among sparkling wines, some are produced from Charmat or bottle fermentation. In this chapter, the practical methods and principles of producing quality sparkling wine is given in details including each step of the production from viticultural parameters, base wine production up to secondary fermentation. After the second fermentation in bottle, the wine is aged on lees. During aging, yeast autolysis occurred. The mechanisms of yeast autolysis and the compounds released are presented. The consequences of yeast autolysis on wine quality are detailed. Finally the riddling and disgorging operation are presented

Protease A activity and nitrogen fractions released during alcoholic fermentation and autolysis in enological conditions

Determination of protease A activity during alcoholic fermentation of a synthetic must (pH 3.5 at 25 degrees C) and during autolysis showed that a sixfold induction of protease A activity occurred after sugar exhaustion, well before 100% cell death occurred. A decrease in protease A activity was observed when yeast cell autolysis started. Extracellular protease A activity was detected late in the autolysis process, which suggests that protease A is not easily released. Evolution of amino acids and peptides was determined during alcoholic fermentation and during autolysis. Amino acids were released in early stationary phase. These amino acids were subsequently assimilated during the fermenta…

Biovi : a research program for reducing chemical input in vine and wine

Decrease of chemical inputs during vine management and winemaking is of great importance from a political and societal point of view. In our ongoing project we propose alternative tools to chemicals in the vineyard and the cellar. We have compared a conventional vineyard protection strategy to an alternative strategy using copper and biocontrol products (Biocontrol) against downy and powdery mildews. Both strategies were compared regarding sanitary quality, berries and/or must enological parameters, and physical, biochemical and biological characteristics (berry surface observation, proteomic, metabolomic, volatilomic, metagenomic analyses). Musts obtained with both strategies were then use…

Characterization of the "viable but nonculturable" (VBNC) state in the wine spoilage yeast Brettanomyces.

Although the viable but not culturable (VBNC) state has been studied in detail in bacteria, it has been suggested that maintenance of viability with loss of culturability also exists in eukaryotic cells, such as in the wine spoilage yeast Brettanomyces. To provide conclusive evidence for the existence of a VBNC state in this yeast, we investigated its capacity to become viable and nonculturable after sulfite stress, and its ability to recover culturability after stressor removal. Sulfite addition induced loss of culturability but maintenance of viability. Increasing the medium pH to decrease the concentration of toxic SO(2) allowed yeast cells to become culturable again, thus demonstrating …

Polyphenols in kombucha: impact of infusion time on extraction and investigation of their behavior during "fermentation"

Kombucha is a non-alcoholic beverage made of sugared tea that is transformed by a symbiotic consortium of yeasts and bacteria. Polyphenols are expected to be responsible of several health benefits attributed to kombucha consumption, among other metabolites. Evidence point at an enhancement of tea polyphenol bioactivity during kombucha elaboration. This study investigated the impact of tea infusion time and of kombucha "fermentation", on the tea polyphenols and the color. Most of the present experiments have been carried out during a 3 rd year Bachelor training by Shane Fennell from Carlow Institute of Technology (Ireland).

Petit traité de mythologie oenologique

La 4ème de couv. indique :"Cet ouvrage s'adresse à tous les amateurs de vins, les experts, les oenophiles curieux et passionnés. Peut-être vous êtes-vous posé des questions sur l'existence ou non de vins de femmes ? Est-ce que la forme du verre a une incidence sur les perceptions sensorielles ? Les guides sont-ils des conseillers efficaces ? Un vin médaillé est-il bon ? Est-ce que la couleur influence notre jugement ? Il y a également les questions que vous ne vous êtes pas posé mais auxquelles vous trouverez les réponses dans ce livre. Vos serviteurs se sont interrogés sur un grand nombre d'allégations, d'idées reçues, d'à priori, de mythes et apportent grâce à l'aide des étudiants du Dipl…

Flor yeasts of Saccharomyces cerevisiae--their ecology, genetics and metabolism.

Abstract The aging of certain white wines is dependent on the presence of yeast strains that develop a biofilm on the wine surface after the alcoholic fermentation. These strains belong to the genus Saccharomyces and are called flor yeasts. These strains possess distinctive characteristics compared with Saccharomyces cerevisiae fermenting strain. The most important one is their capacity to form a biofilm on the air–liquid interface of the wine. The major gene involved in this phenotype is FLO11, however other genes are also involved in velum formation by these yeast and will be detailed. Other striking features presented in this review are their aneuploidy, and their mitochondrial DNA polym…

Directed evolution of oenococcus oeni to improve acid-tolerance reveals fixed beneficial mutations in the citrate locus

Chemical Transfers Occurring Through Oenococcus oeni Biofilm in Different Enological Conditions

International audience; Chardonnay wine malolactic fermentations were carried out to evaluate the chemical transfers occurring at the wood/wine interface in the presence of two different bacterial lifestyles. To do this, Oenococcus oeni was inoculated into must and wine in its planktonic and biofilm lifestyles, whether adhering or not to oak chips, leading to three distinct enological conditions: (i) post-alcoholic fermentation inoculation in wine in the absence of oak chips, (ii) post-alcoholic fermentation inoculation in wine in the presence of oak chips, and (iii) co-inoculation of both Saccharomyces cerevisiae and O. oeni directly in Chardonnay musts in the presence of oak chips. Classi…

PASSAGE OBLIGATOIRE AUX NORMES COMPTABLES IAS/IFRS, CONTRAINTES EN LIQUIDITE ET RATIONNEMENT DU CREDIT : UNE ETUDE EMPIRIQUE DANS L'INDUSTRIE BANCAIRE EUROPENNE

Financial theory indicates that banks dependent on external resources and/or financially fragile have more difficulties in refinancing their operations of credit supply, due to the informational problems they face and/or they cause. In this context, this study tests the hypothesis that the mandatory adoption by banks of the IAS/IFRS accounting standards, known to be of higher quality, leads to an increase in the quantity of loans granted by banks constrained in liquidity, all else equal. Based on a sample of European banks, between 2003 and 2008, we obtain results in favour of this hypothesis.

Diversity of yeast strains of the genus Hanseniaspora in the winery environment: What is their involvement in grape must fermentation?

Isolated yeast populations of Chardonnay grape must during spontaneous fermentation were compared to those isolated on grape berries and in a winery environment before the arrival of the harvest (air, floor, winery equipment) and in the air through time. Two genera of yeast, Hanseniaspora and Saccharomyces, were isolated in grape must and in the winery environment before the arrival of the harvest but not on grape berries. The genus Hanseniaspora represented 27% of isolates in the must and 35% of isolates in the winery environment. The isolates of these two species were discriminated at the strain level by Fourier transform infrared spectroscopy. The diversity of these strains observed in t…

Cyclopropane Fatty Acid Synthase from Oenococcus oeni: Expression in Lactococcus lactis subsp. Cremoris and Biochemical Characterization

Bacterial cyclopropane fatty acid synthases (CFA synthases) catalyze the transfer of a methyl group from S-adenosyl-L-methionine (AdoMet) to the double bond of a lipid chain, thereby forming a cyclopropane ring. CFAs contribute to resistance to acidity, dryness, and osmotic imbalance in many bacteria. This work describes the first biochemical characterization of a lactic acid bacterium CFA synthase. We have overexpressed Oenococcus oeni CFA synthase in E. coli in order to purify the enzyme. The optimum cyclopropanation activity was obtained at pH 5.6 and 35.8 °C. The high K(m) (AdoMet) value obtained (2.26 mM) demonstrates the low affinity of O. oeni enzyme toward the L. lactis subsp. cremo…

Inoculation of Torulaspora delbrueckii as a bio-protection agent in winemaking

International audience; In oenology, bio-protection consists in adding bacteria, yeasts or a mixture of microorganisms on grape must before fermentation in order to reduce the use of chemical compounds such as sulphites. More particularly, non-Saccharvinyces yeasts are used as a total or partial alternative to sulphites. However, scientific data capable of proving the effectiveness of adding these yeasts on grape must is lacking. This study reports the analysis of antimicrobial and antioxidant effects of one non-Saccharamyces yeast, Torulaspora delbruecicii, inoculated at the beginning of the white winemaldng process in two Burgundian wineries as an alternative to sulphiting. The implantati…

Peptidases specific for proline-containing peptides and their unusual peptide-dependent regulation in Oenococcus oeni

International audience; Growth of the lactic acid bacterium (LAB) Oenococcus oeni, which is involved in malolactic fermentation during the winemaking process, is stimulated by peptides originating from yeast. In this study, we investigated the impact of peptides on O. oeni growth, peptidase activity and the expression of genes encoding the studied peptidases. Low levels of PepN activity and very high levels of PepI activity were observed in O. oeni, whereas levels of PepX activity were intermediate. The level of biosynthesis of these O. oeni peptidases was shown to depend on peptides present in the culture medium. These results were confirmed by transcriptional analyses of putative pep gene…

Intraspecific biodiversity and 'spoilage potential' of Brettanomyces bruxellensis in Apulian wines

Abstract The yeast Brettanomyces bruxellensis, generally considered the main oenological spoilage microbe, is able to survive during the winemaking process and it confers off-odors to wine, in reason of its ability to produce considerable amounts of volatile phenols. Forty-eight isolates of B. bruxellensis, obtained from several wines collected in Apulia (Southern Italy), were genetically characterized using an integrated approach, including a strain biodiversity analysis by Sau-PCR. Furthermore, the production of volatile phenols was assessed in wine and in synthetic medium, confirming the oenological spoilage potential of the analysed strains. Our findings indicate a remarkable genetic va…

Effect of Production Phase on Bottle-Fermented Sparkling Wine Quality

This review analyzes bottle-fermented sparkling wine research at each stage of production by evaluating existing knowledge to identify areas that require future investigation. With the growing importance of enological investigation being focused on the needs of the wine production industry, this review examines current research at each stage of bottle-fermented sparkling wine production. Production phases analyzed in this review include pressing, juice adjustments, malolactic fermentation (MLF), stabilization, clarification, tirage, lees aging, disgorging, and dosage. The aim of this review is to identify enological factors that affect bottle-fermented sparkling wine quality, predominantly …

Persistence of Two Non-Saccharomyces Yeasts (Hanseniaspora and Starmerella) in the Cellar

International audience; Different genera and/or species of yeasts present on grape berries, in musts and wines are widely described. Nevertheless, the community of non-Saccharomyces yeasts present in the cellar is still given little attention. Thus it is not known if the cellar is a real ecological niche for these yeasts or if it is merely a transient habitat for populations brought in by grape berries during the winemaking period. This study focused on three species of non-Saccharomyces yeasts commonly encountered during vinification: Starmerella bacillaris (synonymy with Candida zemplinina), Hanseniaspora guilliermondii and Hanseniaspora uvarum. More than 1200 isolates were identified at …

Alteration in membrane fluidity and lipid composition, and modulation of H(+)-ATPase activity in Saccharomyces cerevisiae caused by decanoic acid.

Decanoic acid, a lipophilic agent, inhibited in vitro the plasma membrane H+-ATPase of Saccharomyces cerevisiae grown in YPD medium. Conversely, when decanoic acid (35 μM) was present in the growth medium, the measured H+-ATPase activity was four times higher than that of control cells. K m, and pH and orthovanadate sensitivity were the same for the two growth conditions, which indicated that H+-ATPase activation was not due to conformational changes in the enzyme. The activation process was not entirely reversible which showed that plasma membrane H+-ATPase activation is due to several mechanisms. 1,6-diphenyl-1,3,5-hexatriene anisotropy performed on protoplasts from cells grown in YPD rev…

High-throughput sequencing of amplicons for monitoring yeast biodiversity in must and during alcoholic fermentation

Abstract We compared pyrosequencing technology with the PCR-ITS-RFLP analysis of yeast isolates and denaturing gradient gel electrophoresis (DGGE). These methods gave divergent findings for the yeast population. DGGE was unsuitable for the quantification of biodiversity and its use for species detection was limited by the initial abundance of each species. The isolates identified by PCR-ITS-RFLP were not fully representative of the true population. For population dynamics, high-throughput sequencing technology yielded results differing in some respects from those obtained with other approaches. This study demonstrates that 454 pyrosequencing of amplicons is more relevant than other methods …

Membres du comité d'organisation scientifique

The chemodiversity of wines can reveal a metabologeography expression of cooperage oak wood

Wine chemical compositions, which result from a complex interplay between environmental factors, genetic factors, and viticultural practices, have mostly been studied using targeted analyses of selected families of metabolites. Detailed studies have particularly concerned volatile and polyphenolic compounds because of their acknowledged roles in the organoleptic and therapeutic properties. However, we show that an unprecedented chemical diversity of wine composition can be unraveled through a nontargeted approach by ultrahigh-resolution mass spectrometry, which provides an instantaneous image of complex interacting processes, not easily or possibly resolvable into their unambiguous individu…

Microbiological and technological parameters impacting the chemical composition and sensory quality of kombucha

International audience; Kombucha is a beverage made from sugared tea transformed by yeasts and acetic acid bacteria. Being originally homemade, it has become an industrially produced soft drink whose quality standards are poorly defined and whose production process is still not fully controlled. Based on current knowledge in beverages, links between kombucha's chemical composition and sensorial compounds are drawn. Macromolecules create turbidity, whereas uncharacterized tea pigments derivatives participate in the color. Residual sugars bring sweetness and organic acids produced by acetic acid bacteria form its characteristic sour taste. Acetic acid is also part of its aroma profile, althou…

Development of  a qPCR assay for specific quantification of Botrytis cinerea on grapes

The aim of this study was to develop a system for rapid and accurate real-time quantitative PCR (qPCR) identification and quantification of Botrytis cinerea , one of the major pathogens present on grapes. The intergenic spacer (IGS) region of the nuclear ribosomal DNA was used to specifically detect and quantify B. cinerea . A standard curve was established to quantify this fungus. The qPCR reaction was based on the simultaneous detection of a specific IGS sequence and also contained an internal amplification control to compensate for variations in DNA extraction and the various compounds from grapes that inhibit PCR. In these conditions, the assay had high efficiency (97%), and the limit o…

Foodomics assessed by Fourier transform mass spectrometry

Abstract With its ultrahigh resolution power and excellent mass accuracy, Fourier transform ion cyclotron resonance mass spectrometry is the perfct tool to analyze the “foodome” as the set of all polar small molecular compounds present in an investigated food sample. We present here selected applications of this technique in unraveling the metabolome in complex foods (thermally affected, fermentation products, distillates) such exemplified with Maillard reaction products or with wines and spirits. Technical considerations for high resolution mass analyzers are provided from a practical point of view. Moreover, aspects of food safety and nutritional quality are covered by presenting applicat…

L'uB au coeur de la viticulture durable

SPEIPMUB; Ancrée au cœur d’une région viticole historique, l’université de Bourgogne soutient de multiples projets de recherche viti-vinicoles et s’investit dans la formation des futurs professionnels du secteur. Elle est d’ailleurs la seule université française à disposer d’un domaine expérimental, situé à Marsannay-la-Côte. En activité depuis 1955, il apporte à la fois un support pédagogique aux étudiants de l’institut Jules Guyot, qui y effectuent différentes manipulations en conditions réelles, et répond à des demandes d’expérimentations émanant des viticulteurs et vignerons de la région. Dans ce cadre, il teste de nouveaux produits, matériaux ou micro-organismes susceptibles de mieux p…

FT-IR spectroscopy : A powerful tool for studying the inter- and intraspecific biodiversity of cultivable non-Saccharomyces yeasts isolated from grape must

International audience; The efficiency of the FT-IR technique for studying the inter- and intra biodiversity of cultivable non-Saccharomyces yeasts (NS) present in different must samples was examined. In first, the capacity of the technique FF-IR to study the global diversity of a given sample was compared to the pyrosequencing method, used as a reference technique. Seven different genera (Aureobasidium, Candida, Cryptococcus, Hanseniaspora, Issatchenkia, Metschnikowia and Pichia) were identified by FT-IR and also by pyrosequencing. Thirty-eight other genera were identified by pyrosequencing, but together they represented less than 6% of the average total population of 6 musts. Among the sp…

The Antisense RNA Approach: a New Application for In Vivo Investigation of the Stress Response of Oenococcus oeni, a Wine-Associated Lactic Acid Bacterium

ABSTRACT Oenococcus oeni is a wine-associated lactic acid bacterium mostly responsible for malolactic fermentation in wine. In wine, O. oeni grows in an environment hostile to bacterial growth (low pH, low temperature, and ethanol) that induces stress response mechanisms. To survive, O. oeni is known to set up transitional stress response mechanisms through the synthesis of heat stress proteins (HSPs) encoded by the hsp genes, notably a unique small HSP named Lo18. Despite the availability of the genome sequence, characterization of O. oeni genes is limited, and little is known about the in vivo role of Lo18. Due to the lack of genetic tools for O. oeni , an efficient expression vector in O…

Quantification des Brettanomyces par qPCR

International audience; Brettanomyces bruxellensis est une levure d’altération du vin avec de faibles besoins nutritionnels, résistante à l’éthanol et aux faibles pH, lui permettant une implantation en vin durant ou après la fermentation alcoolique (Conterno et al., 2006). B. bruxellensis est capable de produire des phénols volatils (éthyl-4-phénol, éthyl-4-gaïacol et éthyl-4-catéchol) (Oelofse et al., 2008). Ces molécules volatiles odorantes amènent un caractère phénolé et animal au vin connu sous le nom de « caractère Brett ». D’autres molécules (2-acétyltétrahydropyridine et 2-éthyltétrahydropyridine) produites par B. bruxellensis sont également responsables d’une déviation organoleptiqu…

Cyclopropanation of Membrane Unsaturated Fatty Acids Is Not Essential to the Acid Stress Response of Lactococcus lactis subsp. cremoris

International audience

Specific Identification and Quantification of the Spoilage Microorganism Brettanomyces in Wine by Flow Cytometry: A Useful Tool for Winemakers

International audience; no abstract

Corrigendum to ‘Inoculation of Torulaspora delbrueckii as a bio-protection agent in winemaking’ [Food Research International, Inoculation of Torulaspora delbrueckii as a bio-protection agent in winemaking, Volume: 107 (2018), 451–461]

Cytofluorometric detection of wine lactic acid bacteria: application of malolactic fermentation to the monitoring

Abstract In this study we report for the first time a rapid, efficient and cost-effective method for the enumeration of lactic acid bacteria (LAB) in wine. Indeed, up to now, detection of LAB in wine, especially red wine, was not possible. Wines contain debris that cannot be separated from bacteria using flow cytometry (FCM). Furthermore, the dyes tested in previous reports did not allow an efficient staining of bacteria. Using FCM and a combination of BOX/PI dyes, we were able to count bacteria in wines. The study was performed in wine inoculated with Oenococcus oeni (106 CFU ml−1) stained with either FDA or BOX/PI and analyzed by FCM during the malolactic fermentation (MLF). The analysis …

Editorial for Special Issue “Yeast in Winemaking”

Yeast in winemaking was first studied for its role in alcoholic fermentation, and has led to the publication of a huge amount of scientific articles [...]

Shedding Light on the Formation and Structure of Kombucha Biofilm Using Two-Photon Fluorescence Microscopy

Kombucha pellicles are often used as inoculum to produce this beverage and have become a signature feature. This cellulosic biofilm produced by acetic acid bacteria (AAB) involves yeasts, which are also part of the kombucha consortia. The role of microbial interactions in thede novoformation and structure of kombucha pellicles was investigated during the 3 days following inoculation, using two-photon microscopy coupled with fluorescent staining. Aggregated yeast cells appear to serve as scaffolding to which bacterial cellulose accumulates. This initial foundation leads to a layered structure characterized by a top cellulose-rich layer and a biomass-rich sublayer. This sublayer is expected t…

Biodiversité intraspécifique chez Oenococcus oeni

National audience

L'efficacité des privatisations françaises : une vision dynamique à travers la théorie de la gouvernance

International audience; A reading of the process of privatization through the corporate governance theory leads to propose a model taking into account, on the one hand, the time dimension of the process of privatization, on the other hand, the contextual, organizational, governance and strategic variables which influence this process. After replicating some traditional tests, we test this model on a sample of French privatized firms and on a seven years horizon. The positive effect traditionally attributed to privatizations is not confirmed. The importance of the effect is subordinated to some of the suggested variables.; Une lecture du processus de privatisation à travers la théorie de la …

Unexpected high production of biohydrogen from the endogenous fermentation of grape must deposits

International audience; The aim of this work was to assess the performances of wine byproduct biomass for hydrogen production by dark fermentation. Grape must deposits from two grape varieties (Pinot Gris and Chardonnay) were considered, either with external microbial inoculum or without. We show that grape must residues contain endogenous microflora, well adapted to their environment, which can degrade sugars (initially contained in the biomass) to hydrogen without any nutrient addition. Indeed, hydrogen production during endogenous fermentation is as efficient as with an external heat-treated inoculum (2.5 ± 0.4 LH2.L-1reactor and 1.61 ± 0.41 molH2.mol-1consumed hexose, respectively) with…

Flor Yeast Diversity and Dynamics in Biologically Aged Wines

International audience; Wine biological aging is characterized by the development of yeast strains that form a biofilm on the wine surface after alcoholic fermentation. These yeasts, known as flor yeasts, form a velum that protects the wine from oxidation during aging. Thirty-nine velums aged from 1 to 6 years were sampled from "Vin jaune" from two different cellars. We show for the first time that these velums possess various aspects in term of color and surface aspects. Surprisingly, the heterogeneous velums are mostly composed of one species, S. cerevisiae. Scanning electron microscope observations of these velums revealed unprecedented biofilm structures and various yeast morphologies f…

Influence of yeast autolysis after alcoholic fermentation on the development of <em>Brettanomyces</em>/<em>Dekkera</em> in wine

<p style="text-align: justify;"><em>Brettanomyces</em>, a contaminant yeast, is relatively common in wines and mainly in red wines during barrel aging. The results presented here relate to the effects of yeast lees autolysis on the growth of <em>Brettanomyces</em>. Experiments were realised in a culture medium after alcoholic fermentation, in a hydroalcoholic wine-like solution and in a red wine. <em>Brettanomyces</em> was inoculated at low level : 10<sup>2</sup> cfu/ml and the growth was controlled by counting on agar appropriate medium. Yeast lees from <em>S. cerevisiae</em> were added to these media in the presence or abse…

Influence of cell-cell contact between L. thermotolerans and S. cerevisiae on yeast interactions and the exo-metabolome

International audience; Sequential fermentation of grape must inoculated with L. thermotolerans and then S. cerevisiae 24 h later (typical wine-making practice) was conducted with or without cell-cell contact between the two yeast species. We monitored cell viability of the two species throughout fermentation by flow cytometry. The cell viability of S. cerevisiae decreased under both conditions, but the decrease was greater if there was cell-cell contact. An investigation of the nature of the interactions showed competition between the two species for nitrogen compounds, oxygen, and must sterols. Volatile-compound analysis showed differences between sequential and pure fermentation and that…

Lack of correlation between trehalose accumulation, cell viability and intracellular acidification as induced by various stresses in Saccharomyces cerevisiae.

A pma1-1 mutant of Saccharomyces cerevisiae with reduced H+-ATPase activity and the isogenic wild-type strain accumulated high levels of trehalose in response to a temperature upshift to 40 éC and after addition of 10% ethanol, but only modest levels in response to a rapid drop in external pH and after addition of decanoic acid. There was, however, no correlation between the absolute levels of trehalose in the stressed cells and their viability. All these treatments induced a significant decrease in intracellular pH, and surprisingly, this decrease was very similar in both strains, indicating that intracellular acidification could not be the triggering mechanism for trehalose accumulation i…

Saccharomyces cerevisiae-Oenococcus oeni interactions in wine: current knowledge and perspectives.

Winemaking can be summarized as the biotransformation of must into wine, which is performed principally by Saccharomyces cerevisiae strains during the primary or alcoholic fermentation. A secondary fermentation, the so-called malolactic fermentation (MLF) is a biodeacidification that is often encouraged, since it improves wine stability and quality. Malolactic fermentation usually occurs either spontaneously or after inoculation with selected bacteria after alcoholic fermentation. The main organism responsible for MLF, the lactic acid bacterium Oenococcus oeni, develops in physicochemically harsh conditions, which may lead to MLF failure. Furthermore, yeast that ferment must before or toget…