Molecular study of co-cultures and hybrids of Saccharomyces to address challenges in the wine industry

The increase in the average global temperature due to climate change has also had an effect on the composition of the grape. The higher temperatures have caused a decompensation in the maturation of sugars and phenolic maturity in the berries, with the sugars ripening faster. This decompensation means that the time of harvesting is even more critical, given that if the grapes are harvested when the grape berry has an adequate sugar concentration, the phenolic maturation is not yet optimal and the fermentation of these musts would result in astringent and not very aromatic wines. On the other hand, if it is harvested when phenolic maturity is complete, the concentration of sugars on the grapes will be very high and they would be partially dehydrated, resulting in a low quantity of must and a high concentration of sugar which, when fermented, would give rise to wines with a high alcoholic content. On the other hand, the industry must bear in mind that consumers' tastes are changing, so they will have to adapt to new demands. Nowadays, the consumer trend is to look for wines with low alcohol content, so that they are ‘healthier’ and also have richer aromatic profiles. Therefore, if we put both facts together, the wine industry has a challenge to face from a vineyard point of view with the study of viticulture, but also from a microbiological point of view, with the use of different fermentation strategies. The use of alternative species to Saccharomyces cerevisiae, the yeast par excellence used for alcoholic fermentation, has been a starting point since other species such as S. uvarum or S. kudriavzevii would have a lower ethanol yield and would produce more concentration of interesting compounds such as glycerol or aromatic compounds. In this case, the solution we propose is the use of co-cultures, carrying out mixed S. cerevisiae and S. kudriavzevii fermentations as well as the use of hybrid strains. Hybrid strains would be strains containing S. cerevisiae and S. kudriavzevii genetic material on the one hand and S. cerevisiae and S. uvarum on the other hand. With regard to cocultures, and to study the interaction that may be occurring between both strains when they are in the same medium, objective 1 of the thesis was set out. This objective was to study the competitive ability of numerous S. cerevisiae strains from a wide variety of origins (wine strains, natural strains, traditional fermented beverages, cocoa, etc.) against CR85, a cryotolerant strain of the S. kudriavzevii species. We measured competitive ability as a function of the proportion of each species achieved during fermentation and a wide range of competitive abilities was observed. Interestingly, although the strains of wine origin included the most competitive strains of all, some strains also had a low competitive ability. Therefore, wine strains were selected as representatives of the different fermentative capacities and studied in depth. This led us to a second objective, which was to find out what characteristics the good competing strains have that the bad strains do not have and that are giving them this capacity. We studied their nutrient consumption profiles, observing different strategies, but we also studied at the transcriptomic level, where we observed a high activation of glycolytic, ribosomal and nucleotide synthesis pathways, as well as a faster response from the better competitor strains. On the other hand, with regard to the study of hybrid strains, in order to be able to implement them in the industry, it is necessary to know how they behave metabolically. For this purpose, the metabolome (more than 500 intracellular metabolites) and the production of fermentative compounds such as ethanol or glycerol, among others, of three hybrid strains, two natural strains and one synthetic strain, were analysed. The hybrid strains showed different profiles in the consumption of nitrogen and carbohydrate sources and in the production of volatile compounds and fermentative by-products such as ethanol and glycerol. Significant differences in central carbon metabolism, amino acid synthesis and glutathione metabolism were also revealed between the different strains, highlighting the complexity of the genomic impact on the metabolic behaviour of the hybrids and their phenotypic variability.