SSU1 Checkup, a Rapid Tool for Detecting Chromosomal Rearrangements Related to the SSU1 Promoter in Saccharomyces cerevisiae: An Ecological and Technological Study on Wine Yeast

Chromosomal rearrangements (CR) such as translocations, duplications and inversions play a decisive role in the adaptation of microorganisms to specific environments. In enological Saccharomyces cerevisiae strains, CR involving the promoter region of the gene SSU1 lead to a higher sulfite tolerance by enhancing the SO2 efflux. To date, three different SSU1 associated CR events have been described, including translocations XV-t-XVI and VIII-t-XVI and inversion inv-XVI. In the present study, we developed a multiplex PCR method (SSU1 checkup) that allows a rapid characterization of these three chromosomal configurations in a single experiment. Nearly 600 S. cerevisiae strains collected from fermented grape juice were genotyped by microsatellite markers. We demonstrated that alleles of the SSU1 promoter are differently distributed according to the wine environment (cellar versus vineyard) and the nature of the grape juice. Moreover, rearranged SSU1 promoters are significantly enriched among commercial starters. In addition, the analysis of nearly isogenic strains collected in wine related environments demonstrated that the inheritance of these CR shapes the genetic diversity of clonal populations. Finally, the link between the nature of SSU1 promoter and the tolerance to sulfite was statistically validated in natural grape juice containing various SO2 concentrations. The SSU1 checkup is therefore a convenient new tool for addressing population genetics questions and for selecting yeast strains by using molecular markers.Fil: Marullo, Philippe. Universite de Bordeaux; FranciaFil: Claisse, Olivier. Universite de Bordeaux; FranciaFil: Raymond Eder, María Laura. Universidad Católica de Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J.; ArgentinaFil: Börlin, Marine. Universite de Bordeaux; FranciaFil: Feghali, Nadine. Lebanese University; LíbanoFil: Bernard, Margaux. Universite de Bordeaux; FranciaFil: Legras, Jean Luc. Université Montpellier II; FranciaFil: Albertin, Warren. Universite de Bordeaux; FranciaFil: Rosa, Alberto Luis. Universidad Católica de Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Recursos Naturales y Sustentabilidad José Sanchez Labrador S. J.; ArgentinaFil: Masneuf Pomarede, Isabelle. Universite de Bordeaux; Franci

Front Microbiol

Chromosomal rearrangements (CR) such as translocations, duplications and inversions play a decisive role in the adaptation of microorganisms to specific environments. In enological strains, CR involving the promoter region of the gene lead to a higher sulfite tolerance by enhancing the SO efflux. To date, three different associated CR events have been described, including translocations XV-t-XVI and VIII-t-XVI and inversion inv-XVI. In the present study, we developed a multiplex PCR method ( checkup) that allows a rapid characterization of these three chromosomal configurations in a single experiment. Nearly 600 strains collected from fermented grape juice were genotyped by microsatellite markers. We demonstrated that alleles of the promoter are differently distributed according to the wine environment (cellar versus vineyard) and the nature of the grape juice. Moreover, rearranged promoters are significantly enriched among commercial starters. In addition, the analysis of nearly isogenic strains collected in wine related environments demonstrated that the inheritance of these CR shapes the genetic diversity of clonal populations. Finally, the link between the nature of promoter and the tolerance to sulfite was statistically validated in natural grape juice containing various SO concentrations. The checkup is therefore a convenient new tool for addressing population genetics questions and for selecting yeast strains by using molecular markers

Cellars and vineyards Saccharomyces cerevisiae populations are connected by assyletric bidirectional gene flow.

Vineyards and wineries are ecological habitats that house a diverse community of yeast and bacteria. Grapes and cellars are two sources of S. cerevisiae strains involved in the winemaking process, but the relationship between both S. cerevisiae populations is still poorly understood. Indeed strains in vineyards samples are rarely the same as those isolated from vats. In order to better apprehend this issue, 1374 S. cerevisiae isolates were collected from 193 samples of Merlot grapes obtained across 5 Bordeaux regions and from 11 spontaneously fermenting must of 7 cellars. We obtained 402 different genotypes using 17 microsatellite markers. A first analysis of genotypes indicated that approximately ¼thof isolates presented more than 75% of similarity with commercial yeast starters, suggesting that they escaped from the cellar environment, but presented variations higher that could be detected from the analysis of their industrial batch production. The resulting S. cerevisiae populations of 302 grapes-associated and 225 cellar-associated unique profiles revealed a global low differentiation (Fst=0.036), but with differences from sites to sites. To limit potential geographic or sampling effects, a subset of 72 individuals were selected among the vineyard and cellar isolates, representing 5 cellars and the vines located in their immediate environment. Again a low differentiation was noticed (Fst= 0.03 +/- 0.001). The geneflow between vines and cellars was inferred, using a likelihood approach implemented in the software MIGRATE. Inferences showed that cellar and grape metapopulations present similar theoretical sizes and are connected by asymmetric geneflow: almost 4 times higher in the direction "grapes-to-cellar" than for "cellar-to-grapes" (number of migrants per generation 191 [166 -226] versus 55 [25 - 83], respectively). This reveals that vines and cellars are two compartments of the same ecosystem, which has deep ecological significance

Cellars and vineyards Saccharomyces cerevisiae populations are connected by assyletric bidirectional gene flow.

Vineyards and wineries are ecological habitats that house a diverse community of yeast and bacteria. Grapes and cellars are two sources of S. cerevisiae strains involved in the winemaking process, but the relationship between both S. cerevisiae populations is still poorly understood. Indeed strains in vineyards samples are rarely the same as those isolated from vats. In order to better apprehend this issue, 1374 S. cerevisiae isolates were collected from 193 samples of Merlot grapes obtained across 5 Bordeaux regions and from 11 spontaneously fermenting must of 7 cellars. We obtained 402 different genotypes using 17 microsatellite markers. A first analysis of genotypes indicated that approximately ¼thof isolates presented more than 75% of similarity with commercial yeast starters, suggesting that they escaped from the cellar environment, but presented variations higher that could be detected from the analysis of their industrial batch production. The resulting S. cerevisiae populations of 302 grapes-associated and 225 cellar-associated unique profiles revealed a global low differentiation (Fst=0.036), but with differences from sites to sites. To limit potential geographic or sampling effects, a subset of 72 individuals were selected among the vineyard and cellar isolates, representing 5 cellars and the vines located in their immediate environment. Again a low differentiation was noticed (Fst= 0.03 +/- 0.001). The geneflow between vines and cellars was inferred, using a likelihood approach implemented in the software MIGRATE. Inferences showed that cellar and grape metapopulations present similar theoretical sizes and are connected by asymmetric geneflow: almost 4 times higher in the direction "grapes-to-cellar" than for "cellar-to-grapes" (number of migrants per generation 191 [166 -226] versus 55 [25 - 83], respectively). This reveals that vines and cellars are two compartments of the same ecosystem, which has deep ecological significance

Quantifying the effect of human practices on Saccharomyces cerevisiae vineyardmetapopulation diversity

Saccharomyces cerevisiae is the main actor of wine fermentation but at present, still little is known about the factors impacting its distribution in the vineyards. In this study, 23 vineyards and 7 cellars were sampled over 2 consecutive years in the Bordeaux and Bergerac regions. The impact of geography and farming system and the relation between grape and vat populations were evaluated using a collection of 1374 S. cerevisiae merlot grape isolates and 289 vat isolates analyzed at 17 microsatellites loci. A very high genetic diversity of S. cerevisiae strains was obtained from grape samples, higher in conventional farming system than in organic one. The geographic appellation and the wine estate significantly impact the S. cerevisiae population structure, whereas the type of farming system has a weak global effect. When comparing cellar and vineyard populations, we evidenced the tight connection between the two compartments, based on the high proportion of grape isolates (25%) related to the commercial starters used in the cellar and on the estimation of bidirectional geneflows between the vineyard and the cellar compartments

Appl Environ Microbiol

Three wine estates (designated A, B, and C) were sampled in Sauternes, a typical appellation of the Bordeaux wine area producing sweet white wine. From those wine estates, 551 yeast strains were collected between 2012 and 2014, added to 102 older strains from 1992 to 2011 from wine estate C. All the strains were analyzed through 15 microsatellite markers, resulting in 503 unique Saccharomyces cerevisiae genotypes, revealing high genetic diversity and a low presence of commercial yeast starters. Population analysis performed using Fst genetic distance or ancestry profiles revealed that the two closest wine estates, B and C, which have juxtaposed vineyard plots and common seasonal staff, share more related isolates with each other than with wine estate A, indicating exchange between estates. The characterization of isolates collected 23 years ago at wine estate C in relation to recent isolates obtained at wine estate B revealed the long-term persistence of isolates. Last, during the 2014 harvest period, a temporal succession of ancestral subpopulations related to the different batches associated with the selective picking of noble rotted grapes was highlighted. High genetic diversity of S. cerevisiae isolates from spontaneous fermentation on wine estates in the Sauternes appellation of Bordeaux was revealed. Only 7% of all Sauternes strains were considered genetically related to specific commercial strains. The long-term persistence (over 20 years) of S. cerevisiae profiles on a given wine estate is highlighted

Des outils pour fiabiliser les fermentations des vins et cidres biologiques en utilisant les levures et bactéries indigènes

Ce numéro comprend les articles correspondant aux présentations du Colloque Casdar 2018.With the development of organic wines and ciders, there is a real tendency to carry out spontaneousfermentations, which involve the development of indigenous yeasts and bacteria. Indeed, thesemicroorganisms are sometimes considered as components of the terroir that participate in the typicity ofwines and ciders. However, no scientific knowledge allows us to assert such a specificity, while the lackof control of these microorganisms can lead to difficulties of fermentation, aromatic deviations oralterations. The project CASDAR Levains Bio relied on a network of laboratories, technical institutes andassociations of organic producers to provide the necessary knowledge and practical solutions forcarrying out indigenous fermentations with a good level of control. It has been shown that there is awide diversity of strains of the yeast Saccharomyces cerevisiae and the lactic acid bacteriumOenococcus oeni, that strains are genetically adapted to certain products, but not to regions orproduction sites. Protocols have been developed to allow for the selection of strains from farms or forthe production of "pieds de cuve". Some of the solutions have been successfully transferred toproducers.Avec le développement des vins et cidres bio, on observe une vraie tendance à la réalisation defermentations spontanées, en laissant se développer les levures et bactéries indigènes. En effet, cesmicroorganismes sont parfois considérés comme des éléments du terroir qui participent à la typicité desvins et des cidres. Pourtant, aucune connaissance scientifique ne permet d’affirmer une telle spécificité,alors que la non-maîtrise de ces microorganismes peut conduire à des difficultés de fermentation, desdéviations aromatiques ou des altérations. Le projet Casdar Levains Bio s’est appuyé sur un réseau delaboratoires, instituts techniques et associations de producteurs bio pour apporter les connaissancesnécessaires et des solutions pratiques pour réaliser des fermentations indigènes avec un bon niveau demaîtrise. Il a été montré qu’il existe une grande diversité de souches de la levure Saccharomycescerevisiae et de la bactérie lactique Oenococcus oeni, que des souches sont génétiquement adaptées àcertains produits, mais pas à des régions ou à des sites de production. Des protocoles ont été mis aupoint pour permettre de sélectionner des souches issues des exploitations ou pour réaliser des pieds decuve de microorganismes indigènes. Certaines des solutions ont été transférées avec succès auprèsdes producteurs

Innov. agron.

Avec le développement des vins et cidres bio, on observe une vraie tendance à la réalisation de fermentations spontanées, en laissant se développer les levures et bactéries indigènes. En effet, ces microorganismes sont parfois considérés comme des éléments du terroir qui participent à la typicité des vins et des cidres. Pourtant, aucune connaissance scientifique ne permet d’affirmer une telle spécificité, alors que la non-maîtrise de ces microorganismes peut conduire à des difficultés de fermentation, des déviations aromatiques ou des altérations. Le projet Casdar Levains Bio s’est appuyé sur un réseau de laboratoires, instituts techniques et associations de producteurs bio pour apporter les connaissances nécessaires et des solutions pratiques pour réaliser des fermentations indigènes avec un bon niveau de maîtrise. Il a été montré qu’il existe une grande diversité de souches de la levure Saccharomyces cerevisiae et de la bactérie lactique Oenococcus oeni, que des souches sont génétiquement adaptées à certains produits, mais pas à des régions ou à des sites de production. Des protocoles ont été mis au point pour permettre de sélectionner des souches issues des exploitations ou pour réaliser des pieds de cuve de microorganismes indigènes. Certaines des solutions ont été transférées avec succès auprès des producteurs. | With the development of organic wines and ciders, there is a real tendency to carry out spontaneous fermentations, which involve the development of indigenous yeasts and bacteria. Indeed, these microorganisms are sometimes considered as components of the terroir that participate in the typicity of wines and ciders. However, no scientific knowledge allows us to assert such a specificity, while the lack[br/] of control of these microorganisms can lead to difficulties of fermentation, aromatic deviations or alterations. The project CASDAR Levains Bio relied on a network of laboratories, technical institutes and associations of organic producers to provide the necessary knowledge and practical solutions for carrying out indigenous fermentations with a good level of control. It has been shown that there is a wide diversity of strains of the yeast Saccharomyces cerevisiae and the lactic acid bacterium Oenococcus oeni, that strains are genetically adapted to certain products, but not to regions or production sites. Protocols have been developed to allow for the selection of strains from farms or for the production of "pieds de cuve". Some of the solutions have been successfully transferred to producers