In this study, we presented the first metabolome time course analysis performed among a set of S. uvarum, S. kudriavzevii and S. cerevisiae strains under winemaking conditions. Extracellular and intracellular metabolites, as well as physiological parameters of yeast cells, were monitored along the process to find evidence of different metabolic strategies among species to perform alcoholic fermentation. A thorough inspection of time trends revealed several differences in utilization or accumulation of fermentation by-products. We confirmed the ability of S. uvarum and S. kudriavzevii

Nonconventional yeast species hold the promise to provide novel metabolic routes to produce industrially relevant compounds and tolerate specific stressors, such as cold temperatures. This work validated the first multiphase multiobjective genome-scale dynamic model to describe carbon and nitrogen metabolism throughout batch fermentation.

The utilization of S. eubayanus has recently become a topic of interest due to the novel organoleptic properties imparted to beer. However, the utilization of S. eubayanus in brewing requires the comprehension of the mechanisms that underlie fermentative differences generated from its natural genetic variability. Here, we evaluated fermentation performance and volatile compound production in ten genetically distinct S. eubayanus strains in a brewing fermentative context. The evaluated strains showed a broad phenotypic spectrum, some of them exhibiting a high fermentation capacity and high levels of volatile esters and/or higher alcohols. Subsequently, we obtained molecular profiles by generating 'end-toend' genome assemblies, as well as metabolome and transcriptome profiling of two Patagonian isolates exhibiting significant differences in beer aroma profiles. These strains showed clear differences in concentrations of intracellular metabolites, including amino acids, such as valine, leucine and isoleucine, likely impacting the production of 2-methylpropanol and 3-methylbutanol. These differences in the production of volatile compounds are attributed to gene expression variation, where the most profound differentiation is attributed to genes involved in assimilatory sulfate reduction, which in turn validates phenotypic differences in H 2 S production. This study lays a solid foundation for future research to improve fermentation performance and select strains for new lager styles based on aroma and metabolic profiles.

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The monitoring of fermentation at low temperatures (12-15 C) is a current practice in the winery for retention and enhancement of the flavour volatile content of wines. Among Saccharomyces species, Saccharomyces uvarum and Saccharomyces kudriavzevii have revealed interesting industrial properties, including better adaptation at low temperatures. To gather deeper knowledge of the fermentative metabolism at a low temperature of these species together with S. cerevisiae, we performed a comparative metabolomic analysis using four representative strains. We used batch cultures to obtain an exhaustive and dynamic image of the metabolome of strains passing through the sequential stresses related to the winemaking environment. A great variety of intra-and extracellular metabolites (>500 compounds) were quantified across fermentation using distinct chromatographic methods. Besides a global decrease in the lipid composition of the four strains when they entered into the stationary phase, we reported some strain-specific high magnitude changes. Examples of these differences included divergent patterns of production of short-chain fatty acids and erythritol in the S. uvarum strain. Strains also differed in expression for aromatic amino acid biosynthesis and sulphur metabolism, including the glutathione pathway. These data will allow us to refine and obtain the most value of fermentations with this alternative Saccharomyces species.

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Yeasts constitute over 1500 species with great potential for biotechnology. Still, the yeast Saccharomyces cerevisiae dominates industrial applications and many alternative physiological capabilities of lesser-known yeasts arenot being fully exploited. While comparative genomics receives substantial attention, little is known about yeasts' metabolic specificity in batch cultures. Here we propose a multi-phase multi-objective dynamic genome-scale model of yeast batch cultures that describes the uptake of carbon and nitrogen sources and the production of primary and secondary metabolites. The model integrates a specific metabolic reconstruction, based on the consensus Yeast8, and a kinetic model describing the time-varying culture environment. Besides, we proposed a multi-phase multi-objective flux balance analysis to compute the dynamics of intracellular fluxes. We then compared the metabolism of S. cerevisiae and S. uvarum strains in a rich medium fermentation. The model successfully explained the experimental data and brought novel insights into how cryotolerant strains achieve redox balance. The proposed model (along with the corresponding code) provides a comprehensive picture of the main steps occurring inside the cell during batch cultures and offers a systematic approach to prospect or metabolically engineering novel yeast cell factories.

This work aims to describe the wine fermentation characteristics of 23 natural S. cerevisiae x S. kudriavzevii hybrid yeasts related to fermentative environments isolated from different regions and their significance for the aroma spectra of the produced wines. Fermentations were performed at 12 °C in artificial must, and S. cerevisiae and S. kudriavzevii pure species strains were used for comparison purposes. We determined the relevant kinetic parameters of fermentation, the concentration of the main metabolites and the main aromarelated compounds produced after fermentation. The results revealed that some strains that show well-rounded characteristics could be profitable yeast starters for low-temperature fermentation in winemaking, such as wine hybrid SPG172 but, surprisingly, also beer hybrid CECT11002, adding the efficient fermentative kinetics to the high production of aroma-related compounds. In addition, a novel metabolic correlation between fermentation performance and aroma production is described.