Influence of different stress factors during the elaboration of grape must's pieddecuve on the dynamics of yeast populations during alcoholic fermentation.

The pieddecuve (PdC) technique involves using a portion of grape must to undergo spontaneous fermentation, which is then used to inoculate a larger volume of must. This allows for promoting autochthonous yeasts present in the must, which can respect the typicality of the resulting wine. However, the real impact of this practice on the yeast population has not been properly evaluated. In this study, we examined the effects of sulphur dioxide (SO2), temperature, ethanol supplementation, and time on the dynamics and selection of yeasts during spontaneous fermentation to be used as PdC. The experimentation was conducted in a synthetic medium and sterile must using a multi-species yeast consortium and in un-inoculated natural grape must. Saccharomyces cerevisiae dominated both the PdC and fermentations inoculated with commercial wine yeast, displaying similar population growth regardless of the tested conditions. However, using 40 mg/L of SO2 and 1% (v/v) ethanol during spontaneous fermentation of Muscat of Alexandria must allowed the non-Saccharomyces to be dominant during the first stages, regardless of the temperature tested. These findings suggest that it is possible to apply the studied parameters to modulate the yeast population during spontaneous fermentation while confirming the effectiveness of the PdC methodology in controlling alcoholic fermentation.

Catatonic postpartum paternal depression as a first debut of a bipolar disorder: a case report.

Paternal postpartum depression (PD) is considered an affective disorder that affects fathers during the months following childbirth. Interestingly, it has been observed that during these months the chances of a male parent suffering from depression are double that for a non-parent male counterpart. We present the case of a 34-year-old man with no relevant medical history in who, overlapping her daughter's birth, several depressive symptoms emerged, such as fatigue, lack of concentration, sleeping disturbances and abandonment of care of the newborn. Prior to consultation, patient refused to eat and open his eyes, and his speech became progressively more parsimonious until reaching mutism. The patient was diagnosed with a severe depressive disorder with catatonia. Given the lack of improvement with pharmacological treatment and due to the evidence of electroconvulsive therapy (ECT)'s effectiveness on patients with catatonia, acute ECT treatment was indicated and started. It should be noted that PD is an important entity to consider in our differential diagnosis of young parents who present a depressive episode. Few cases of relatively young patients presenting with such clinical presentation have been described and, although this case presents some of the characteristics described in the epidemiology of PD, other clinical aspects are not typical of this entity. Informed consent was obtained from the patient for the purpose of publication.

Medical Students' Opinion of Their Learning Process.

Introduction: The opinion of students is of utmost importance to identify areas of improvement in undergraduate studies. Medical schools would use this information to plan actions to ensure that the students achieve the necessary medical knowledge. The aim of this study was to analyse the opinion of medical students about their learning process and to analyse the influence of their experience according to their year of medical degree. Methods: A questionnaire including 21 items, divided into four sections (motivation, theory lectures, hospital internships, and research) and two overall questions, was distributed among eligible 246 students. Each item was scored from 1 (strongly disagree) to 5 (strongly agree). The opinions of intermediate-year students of medical degree (3rd and 4th) were compared to late-year students (5th and 6th). Results: A total of 148 students answered the questionnaire (60.2% response rate). The mean scores for overall student motivation and teaching quality were 6.15 and 7.10, respectively. The student-teacher interaction and new learning technological tools were considered important for student motivation. The only differences found between the two groups of students were that late-year students wished to become part of a medical team and to learn writing scientific papers more than the intermediate-year students. Conclusions: This questionnaire revealed that the year of career had little influence on the medical students' opinion on their learning process during their undergraduate studies. Late-year students rated highest on being more interested in being part of a medical team and their knowledge on writing scientific articles. The use of new technologies and the student-teacher interaction is key to motivate students.

Pointwise monotonicity of heat kernels.

In this paper we present an elementary proof of a pointwise radial monotonicity property of heat kernels that is shared by the Euclidean spaces, spheres and hyperbolic spaces. The main result was discovered by Cheeger and Yau in 1981 and rediscovered in special cases during the last few years. It deals with the monotonicity of the heat kernel from special points on revolution hypersurfaces. Our proof hinges on a non straightforward but elementary application of the parabolic maximum principle. As a consequence of the monotonicity property, we derive new inequalities involving classical special functions.

Predictive evolution of metabolic phenotypes using model-designed environments.

Adaptive evolution under controlled laboratory conditions has been highly effective in selecting organisms with beneficial phenotypes such as stress tolerance. The evolution route is particularly attractive when the organisms are either difficult to engineer or the genetic basis of the phenotype is complex. However, many desired traits, like metabolite secretion, have been inaccessible to adaptive selection due to their trade-off with cell growth. Here, we utilize genome-scale metabolic models to design nutrient environments for selecting lineages with enhanced metabolite secretion. To overcome the growth-secretion trade-off, we identify environments wherein growth becomes correlated with a secondary trait termed tacking trait. The latter is selected to be coupled with the desired trait in the application environment where the trait manifestation is required. Thus, adaptive evolution in the model-designed selection environment and subsequent return to the application environment is predicted to enhance the desired trait. We experimentally validate this strategy by evolving Saccharomyces cerevisiae for increased secretion of aroma compounds, and confirm the predicted flux-rerouting using genomic, transcriptomic, and proteomic analyses. Overall, model-designed selection environments open new opportunities for predictive evolution.

Importance of micronutrients and organic nitrogen in fermentations with Torulaspora delbrueckii and Saccharomyces cerevisiae.

The current use of non-Saccharomyces yeasts in mixed fermentations increases the relevance of the interactions between yeast species. In this work, the interactions between Saccharomyces cerevisiae and Torulaspora delbrueckii were analyzed. For this purpose, fermentations with and without contact between strains of those yeast species were performed in synthetic must. Fermentation kinetics, yeast growth and dynamics were measured over time. Additionally, the effects of nitrogen and other nutrient supplementations on the mixed fermentations were determined. Our results showed that S. cerevisiae did not always dominate the sequential fermentations, and experiments without yeast contact (in which T. delbrueckii cells were removed from the medium before inoculating S. cerevisiae at 48 h) resulted in stuck fermentations except when the inoculum size was increased (from 2 × 106 to 108 cells/mL) or there was a supplementation of thiamine, zinc and amino acids at the same concentration as initially found in the synthetic must. Our findings highlight the importance of inoculum size and ensuring the availability of enough micronutrients for all yeast species, especially in sequential fermentations.

Strategies for microbiological control of the alcoholic fermentation in wines by exploiting the microbial terroir complexity: A mini-review.

A complex consortium of yeasts is the principal responsible of wine fermentation, being Saccharomyces cerevisiae the main driver. The use of selected yeast, beginning with Saccharomyces strains, is one of the main achievements in microbiological control in the wine industry. However, the use of single strain starters of S. cerevisiae and the limited variability of strains has increased the objections to its use due to the production of wines with a homogeneous profile. New tendencies in winemaking have emphasized the microbiological terroir and challenged the use of selected starters from different areas, including Non-Saccharomyces yeast or multi-strain starters in order to add complexity to the biochemical composition of wines. Nevertheless, these strategies also harbor their own challenges. In the present mini-review, we focus on the alternatives to current commercial yeast starters mainly based on the local multispecies starters or controlled spontaneous fermentations, considering the advantages and limitations of each strategy. Also, we present an ancestral technique based on the use of pre-fermented must (Pied-de-Cuve) because it provides certain microbial control to the alcoholic fermentation while allows the development of autochthonous microorganisms that might confer microbial typicity to the produced wines. Nevertheless, the latest strategy needs further research to establish a scientific background for the selection of the best Pied-de-Cuve development. Finally, the tendencies in winemaking should find a commitment between microbial control to guarantee alcoholic fermentation fulfillment and the production of wines reflecting microbial typicity to respond to consumer demands, without forgetting the scaling up in the cellars.

Protective Effects of Melatonin on Saccharomyces cerevisiae under Ethanol Stress.

During alcoholic fermentation, Saccharomyces cerevisiae is subjected to several stresses, among which ethanol is of capital importance. Melatonin, a bioactive molecule synthesized by yeast during alcoholic fermentation, has an antioxidant role and is proposed to contribute to counteracting fermentation-associated stresses. The aim of this study was to unravel the protective effect of melatonin on yeast cells subjected to ethanol stress. For that purpose, the effect of ethanol concentrations (6 to 12%) on a wine strain and a lab strain of S. cerevisiae was evaluated, monitoring the viability, growth capacity, mortality, and several indicators of oxidative stress over time, such as reactive oxygen species (ROS) accumulation, lipid peroxidation, and the activity of catalase and superoxide dismutase enzymes. In general, ethanol exposure reduced the cell growth of S. cerevisiae and increased mortality, ROS accumulation, lipid peroxidation and antioxidant enzyme activity. Melatonin supplementation softened the effect of ethanol, enhancing cell growth and decreasing oxidative damage by lowering ROS accumulation, lipid peroxidation, and antioxidant enzyme activities. However, the effects of melatonin were dependent on strain, melatonin concentration, and growth phase. The results of this study indicate that melatonin has a protective role against mild ethanol stress, mainly by reducing the oxidative stress triggered by this alcohol.

Effect of a Multistarter Yeast Inoculum on Ethanol Reduction and Population Dynamics in Wine Fermentation.

Microbiological strategies are currently being considered as methods for reducing the ethanol content of wine. Fermentations started with a multistarter of three non-Saccharomyces yeasts (Metschnikowia pulcherrima (Mp), Torulaspora delbrueckii (Td) and Zygosaccharomyces bailii (Zb)) at different inoculum concentrations. S. cerevisiae (Sc) was inoculated into fermentations at 0 h (coinoculation), 48 h or 72 h (sequential fermentations). The microbial populations were analyzed by a culture-dependent approach (Wallerstein Laboratory Nutrient (WLN) culture medium) and a culture-independent method (PMA-qPCR). The results showed that among these three non-Saccharomyces yeasts, Td became the dominant non-Saccharomyces yeast in all fermentations, and Mp was the minority yeast. Sc was able to grow in all fermentations where it was involved, being the dominant yeast at the end of fermentation. We obtained a significant ethanol reduction of 0.48 to 0.77% (v/v) in sequential fermentations, with increased concentrations of lactic and acetic acids. The highest reduction was achieved when the inoculum concentration of non-Saccharomyces yeast was 10 times higher (107 cells/mL) than that of S. cerevisiae. However, this reduction was lower than that obtained when these strains were used as single non-Saccharomyces species in the starter, indicating that interactions between them affected their performance. Therefore, more combinations of yeast species should be tested to achieve greater ethanol reductions.

High-Throughput Sequencing Approach to Analyze the Effect of Aging Time and Barrel Usage on the Microbial Community Composition of Red Wines.

Wine aged in barrels or bottles is susceptible to alteration by microorganisms that affect the final product quality. However, our knowledge of the microbiota during aging and the factors modulating the microbial communities is still quite limited. The present work uses high-throughput sequencing (HTS) techniques to deal with the meta-taxonomic characterization of microbial consortia present in red wines along 12 months aging. The wines obtained from two different grape varieties were aged at two different cellars and compared based on time of wine aging in the barrels, previous usage of the barrels, and differences between wine aging in oak barrels or glass bottles. The aging in barrels did not significantly affect the microbial diversity but changed the structure and composition of fungal and bacterial populations. The main microorganisms driving these changes were the bacterial genera Acetobacter, Oenococcus, Lactobacillus, Gluconobacter, Lactococcus, and Komagataeibacter and the fungal genera Malassezia, Hanseniaspora, and Torulaspora. Our results showed that the oak barrels increased effect on the microbial diversity in comparison with the glass bottles, in which the microbial community was very similar to that of the wine introduced in the barrels at the beginning of the aging. Furthermore, wine in the bottles harbored higher proportion of Lactobacillus but lower proportion of Acetobacter. Finally, it seems that 1 year of previous usage of the barrels was not enough to induce significant changes in the diversity or composition of microbiota through aging compared with new barrels. This is the first meta-taxonomic study on microbial communities during wine aging and shows that the microorganism composition of barrel-aged wines was similar at both cellars. These results hint at the possibility of a common and stable microbiota after aging in the absence of exogenous alterations. Further corroborations on the current outcome would be valuable for the comparison and detection of microbial alterations during aging that could potentially prevent economic losses in the wine industry.

Transcriptomic Insights into the Effect of Melatonin in Saccharomyces cerevisiae in the Presence and Absence of Oxidative Stress.

Melatonin is a ubiquitous indolamine that plays important roles in various aspects of biological processes in mammals. In Saccharomyces cerevisiae, melatonin has been reported to exhibit antioxidant properties and to modulate the expression of some genes involved in endogenous defense systems. The aim of this study was to elucidate the role of supplemented melatonin at the transcriptional level in S. cerevisiae in the presence and absence of oxidative stress. This was achieved by exposing yeast cells pretreated with different melatonin concentrations to hydrogen peroxide and assessing the entry of melatonin into the cell and the yeast response at the transcriptional level (by microarray and qPCR analyses) and the physiological level (by analyzing changes in the lipid composition and mitochondrial activity). We found that exogenous melatonin crossed cellular membranes at nanomolar concentrations and modulated the expression of many genes, mainly downregulating the expression of mitochondrial genes in the absence of oxidative stress, triggering a hypoxia-like response, and upregulating them under stress, mainly the cytochrome complex and electron transport chain. Other categories that were enriched by the effect of melatonin were related to transport, antioxidant activity, signaling, and carbohydrate and lipid metabolism. The overall results suggest that melatonin is able to reprogram the cellular machinery to achieve tolerance to oxidative stress.

Qualitative Factor-Based Comparison of NMR, Targeted and Untargeted GC-MS and LC-MS on the Metabolomic Profiles of Rioja and Priorat Red Wines.

Wine origin and ageing are two factors related to wine quality which in turn is associated to wine metabolome. Currently, new metabolomic techniques and proper statistics procedures allow accurate profiling of wine metabolome. Thus, the main goal was to evaluate different metabolomic methodologies on their ability to provide patterns on the wine metabolome based on selected factors, such as ageing of barrel-aged wine (factor time), prior usage of the barrels (factor barrel-type), and differences between wine ageing in barrels or glass bottles (factor bottled-wine). In the current study, we implement NMR, targeted and untargeted GC-MS and LC-MS metabolomic analytical techniques so as to gain insights into the volatile and nonvolatile wine metabolome composition of red wines from two cellars located in the only two Spanish Qualified Appellations of Origin; DOQ Priorat and DOCa Rioja regions. Overall, 95 differentially significant metabolites were identified facilitating the evaluation of the analytical methodologies performance and finding common trends of those metabolites depending on the considered factor. The results did not favor NMR as an effective technique on the current dataset whereas suggested LC-MS as an adequate technique for revealing differences based on the factor time, targeted GC-MS on the factor barrel-type, and untargeted GC-MS on the factor bottled-wine. Thus, a combination of different metabolomic techniques is necessary for a complete overview of the metabolome changes. These results ease the selection of the correct methodology depending on the specific factor investigated.

Viability-PCR Allows Monitoring Yeast Population Dynamics in Mixed Fermentations Including Viable but Non-Culturable Yeasts.

The use of controlled mixed inocula of Saccharomyces cerevisiae and non-Saccharomyces yeasts is a common practice in winemaking, with Torulaspora delbrueckii, Lachancea thermotolerans and Metschnikowia pulcherrima being the most commonly used non-Saccharomyces species. Although S. cerevisiae is usually the dominant yeast at the end of mixed fermentations, some non-Saccharomyces species are also able to reach the late stages; such species may not grow in culture media, which is a status known as viable but non-culturable (VBNC). Thus, an accurate methodology to properly monitor viable yeast population dynamics during alcoholic fermentation is required to understand microbial interactions and the contribution of each species to the final product. Quantitative PCR (qPCR) has been found to be a good and sensitive method for determining the identity of the cell population, but it cannot distinguish the DNA from living and dead cells, which can overestimate the final population results. To address this shortcoming, viability dyes can be used to avoid the amplification and, therefore, the quantification of DNA from non-viable cells. In this study, we validated the use of PMAxx dye (an optimized version of propidium monoazide (PMA) dye) coupled with qPCR (PMAxx-qPCR), as a tool to monitor the viable population dynamics of the most common yeast species used in wine mixed fermentations (S. cerevisiae, T. delbrueckii, L. thermotolerans and M. pulcherrima), comparing the results with non-dyed qPCR and colony counting on differential medium. Our results showed that the PMAxx-qPCR assay used in this study is a reliable, specific and fast method for quantifying these four yeast species during the alcoholic fermentation process, being able to distinguish between living and dead yeast populations. Moreover, the entry into VBNC status was observed for the first time in L. thermotolerans and S. cerevisiae during alcoholic fermentation. Further studies are needed to unravel which compounds trigger this VBNC state during alcoholic fermentation in these species, which would help to better understand yeast interactions.

Self-Adjoint Dirac Operators on Domains in R 3.

In this paper, the spectral and scattering properties of a family of self-adjoint Dirac operators in L 2 ( Ω ; C 4 ) , where Ω âŠ‚ R 3 is either a bounded or an unbounded domain with a compact C 2 -smooth boundary, are studied in a systematic way. These operators can be viewed as the natural relativistic counterpart of Laplacians with boundary conditions as of Robin type. Our approach is based on abstract boundary triple techniques from extension theory of symmetric operators and a thorough study of certain classes of (boundary) integral operators, that appear in a Krein-type resolvent formula. The analysis of the perturbation term in this formula leads to a description of the spectrum and a Birman-Schwinger principle, a qualitative understanding of the scattering properties in the case that Ω is an exterior domain, and corresponding trace formulas.

Genetic and transcriptomic evidences suggest ARO10 genes are involved in benzenoid biosynthesis by yeast.

Benzenoids are compounds associated with floral and fruity flavours in flowers, fruits and leaves and present a role in hormonal signalling in plants. These molecules are produced by the phenyl ammonia lyase pathway. However, some yeasts can also synthesize them from aromatic amino acids using an alternative pathway that remains unknown. Hanseniaspora vineae can produce benzenoids at levels up to two orders of magnitude higher than Saccharomyces species, so it is a model microorganism for studying benzenoid biosynthesis pathways in yeast. According to their genomes, several enzymes have been proposed to be involved in a mandelate pathway similar to that described for some prokaryotic cells. Among them, the ARO10 gene product could present benzoylformate decarboxylase activity. This enzyme catalyses the decarboxylation of benzoylformate into benzaldehyde at the end of the mandelate pathway in benzyl alcohol formation. Two homologous genes of ARO10 were found in the two sequenced H. vineae strains. In this study, nine other H. vineae strains were analysed to detect the presence and per cent homology of ARO10 sequences by PCR using specific primers designed for this species. Also, the copy number of the genes was estimated by quantitative PCR. To verify the relation of ARO10 with the production of benzyl alcohol during fermentation, a deletion mutant in the ARO10 gene of Saccharomyces cerevisiae was used. The two HvARO10 paralogues were analysed and compared with other α-ketoacid decarboxylases at the sequence and structural level.

Effect of Several Nutrients and Environmental Conditions on Intracellular Melatonin Synthesis in Saccharomyces cerevisiae.

Melatonin is a bioactive compound that is present in fermented beverages and has been described to be synthesized by yeast during alcoholic fermentation. The aim of this study was to assess the capacity of intracellular and extracellular melatonin production by different Saccharomyces strains from diverse food origin and to study the effects of different fermentation parameters, such as sugar and nitrogen concentration, temperature or initial population, on melatonin production using a synthetic grape must medium. Melatonin from fermentation samples was analyzed by liquid chromatography mass spectrometry. Intracellular melatonin synthesis profile did not present differences between yeast strains. However, extracellular melatonin production depended on the yeast origin. Thus, we suggest that melatonin production and secretion during the different yeast growth phases follows a species-specific pattern. Other parameters that affected the fermentation process such as sugar content and low temperature had an impact on intracellular melatonin production profile, as well as the melatonin content within the cell. This study reports the effect of several conditions on the melatonin synthesis profile, highlighting its possible role as a signal molecule.

A Rapid Method for Selecting Non-Saccharomyces Strains with a Low Ethanol Yield.

The alcohol content in wine has increased due to external factors in recent decades. In recent reports, some non-Saccharomyces yeast species have been confirmed to reduce ethanol during the alcoholic fermentation process. Thus, an efficient screening of non-Saccharomyces yeasts with low ethanol yield is required due to the broad diversity of these yeasts. In this study, we proposed a rapid method for selecting strains with a low ethanol yield from forty-five non-Saccharomyces yeasts belonging to eighteen species. Single fermentations were carried out for this rapid selection. Then, sequential fermentations in synthetic and natural must were conducted with the selected strains to confirm their capacity to reduce ethanol compared with that of Saccharomyces cerevisiae. The results showed that ten non-Saccharomyces strains were able to reduce the ethanol content, namely, Hanseniaspora uvarum (2), Issatchenkia terricola (1), Metschnikowia pulcherrima (2), Lachancea thermotolerans (1), Saccharomycodes ludwigii (1), Torulaspora delbrueckii (2), and Zygosaccharomyces bailii (1). Compared with S. cerevisiae, the ethanol reduction of the selected strains ranged from 0.29 to 1.39% (v/v). Sequential inoculations of M. pulcherrima (Mp51 and Mp FA) and S. cerevisiae reduced the highest concentration of ethanol by 1.17 to 1.39% (v/v) in synthetic or natural must. Second, sequential fermentations with Z. bailii (Zb43) and T. delbrueckii (Td Pt) performed in natural must yielded ethanol reductions of 1.02 and 0.84% (v/v), respectively.

Nitrogen Preferences during Alcoholic Fermentation of Different Non-Saccharomyces Yeasts of Oenological Interest.

Non-Saccharomyces yeasts have long been considered spoilage microorganisms. Currently, oenological interest in those species is increasing, mostly due to their positive contribution to wine quality. In this work, the fermentative capacity and nitrogen consumption of several non-Saccharomyces wine yeast (Torulaspora delbrueckii, Lachancea thermotolerans, Starmerella bacillaris, Hanseniaspora uvarum, and Metschnikowia pulcherrima) were analyzed. For this purpose, synthetic must with three different nitrogen compositions was used: a mixture of amino acids and ammonium, only organic or inorganic nitrogen. The fermentation kinetics, nitrogen consumption, and yeast growth were measured over time. Our results showed that the good fermentative strains, T. delbrueckii and L. thermotolerans, had high similarities with Saccharomyces cerevisiae in terms of growth, fermentation profile, and nitrogen assimilation preferences, although L. thermotolerans presented an impaired behavior when only amino acids or ammonia were used, being strain-specific. M. pulcherrima was the non-Saccharomyces strain least affected by the nitrogen composition of the medium. The other two poor fermentative strains, H. uvarum and S. bacillaris, behaved similarly regarding amino acid uptake, which occurred earlier than that of the good fermentative species in the absence of ammonia. The results obtained in single non-Saccharomyces fermentations highlighted the importance of controlling nitrogen requirements of the wine yeasts, mainly in sequential fermentations, in order to manage a proper nitrogen supplementation, when needed.

Melatonin and glycolytic protein interactions are related to yeast fermentative capacity.

Melatonin is an indole amine that interacts with some proteins in mammals, such as calreticulin, calmodulin or sirtuins. In yeast, melatonin is synthetized and interacts with glycolytic proteins during alcoholic fermentation in Saccharomyces cerevisiae. Due to its importance as an antioxidant molecule in both Saccharomyces and non-Saccharomyces yeasts, the aim of this study was to determine the intracellular and extracellular synthesis profiles of melatonin in four non-Saccharomyces strains (Torulaspora delbrueckii, Hanseniaspora uvarum, Starmeralla bacillaris and Metschnikowia pulcherrima) and to confirm whether glycolytic enzymes can also interact with this molecule in non-conventional yeast cells. Melatonin from fermentation samples was analyzed by liquid chromatography mass spectrometry, and proteins bound to melatonin were immunopurified by melatonin-IgG-Dynabeads. Melatonin was produced in a similar pattern in all non-Saccharomyces yeast, with M. pulcherrima and S. bacillaris being the highest producers. However, melatonin only bound to proteins in two non-conventional yeasts, S. bacillaris and T. delbrueckii, which specifically had higher fermentative capacities. Sequence analysis showed that most proteins shared high levels of homology with glycolytic enzymes, but an RNA-binding protein, the elongation alpha factor, which is related to mitochondria, was also identified. This study reports for the first time the interaction of melatonin with proteins inside non-Saccharomyces yeast cells. These results reinforce the possible role of melatonin as a signal molecule, likely related to fermentation metabolism and provide a new perspective for understanding its role in yeast.

Brettanomyces bruxellensis wine isolates show high geographical dispersal and long persistence in cellars.

Brettanomyces bruxellensis is the main wine spoiler yeast all over the world, yet the structure of the populations associated with winemaking remains elusive. In this work, we considered 1411 wine isolates from 21 countries that were genotyped using twelve microsatellite markers. We confirmed that B. bruxellensis isolates from wine environments show high genetic diversity, with 58 and 42% of putative triploid and diploid individuals respectively distributed in 5 main genetic groups. The distribution in the genetic groups varied greatly depending on the country and/or the wine-producing region. However, the two possible triploid wine groups showing sulfite resistance/tolerance were identified in almost all regions/countries. Genetically identical isolates were also identified. The analysis of these clone groups revealed that a given genotype could be isolated repeatedly in the same winery over decades, demonstrating unsuspected persistence ability. Besides cellar residency, a great geographic dispersal was also evidenced, with some genotypes isolated in wines from different continents. Finally, the study of old isolates and/or isolates from old vintages revealed that only the diploid groups were identified prior 1990 vintages. The putative triploid groups were identified in subsequent vintages, and their proportion has increased steadily these last decades, suggesting adaptation to winemaking practices such as sulfite use. A possible evolutionary scenario explaining these results is discussed.