Thermo-adaptive evolution to generate improved Saccharomyces cerevisiae strains for cocoa pulp fermentations.

Abstract Cocoa pulp fermentation is a consequence of the succession of indigenous yeasts, lactic acid bacteria and acetic acid bacteria that not only produce a diversity of metabolites, but also cause the production of flavour precursors. However, as such spontaneous fermentations are less reproducible and contribute to produce variability, interest in a microbial starter culture is growing that could be used to inoculate cocoa pulp fermentations. This study aimed to generate robust S. cerevisiae strains by thermo-adaptive evolution that could be used in cocoa fermentation. We evolved a cocoa strain in a sugary defined medium at high temperature to improve both fermentation and growth capacity. Moreover, adaptive evolution at high temperature (40 °C) also enabled us to unveil the molecular basis underlying the improved phenotype by analysing the whole genome sequence of the evolved strain. Adaptation to high-temperature conditions occurred at different genomic levels, and promoted aneuploidies, segmental duplication, and SNVs in the evolved strain. The lipid profile analysis of the evolved strain also evidenced changes in the membrane composition that contribute to maintain an appropriate cell membrane state at high temperature. Our work demonstrates that experimental evolution is an effective approach to generate better-adapted yeast strains at high temperature for industrial processes.