Simultaneous improvement of fructophilicity and ethanol tolerance of Saccharomyces cerevisiae strains through a single Adaptive Laboratory Evolution Strategy

Saccharomyces cerevisiae is the main yeast used in the winemaking industry. Its innate glucophilicity provokes a discrepancy in glucose and fructose consumption during alcoholic fermentation of grape must, which, combined with the inhibitory effect of ethanol accumulated in the fermentation broth, might lead to stuck or sluggish fermentations. In the present study, we realized an Adaptive Laboratory Evolution strategy, where an alcoholic fermentation of a 20 g/L fructose broth was followed by cell selection in a high ethanol concentration environment, employed in two different S. cerevisiae strains, named CFB and BLR. The evolved populations originated from each strain after 100 generations of evolution exhibited diverse fermentative abilities. One evolved population, originated from CFB strain, fermented a synthetic broth of 100 g/L glucose and 100 g/L fructose to dryness in 170 h, whereas the parental strain did not complete the fermentation even after 1000 h of incubation. The parameters of growth of the parental and evolved populations of the present study, as well as of the ethanol tolerant populations acquired in a previous study, when grown in a synthetic broth of 100 g/L glucose and 100 g/L fructose, were calculated through a kinetic model, and were compared to each other in order to identify the effect of evolution on the biochemical behavior of the strains. Finally, in a 200 g/L fructose synthetic broth fermentation, only the evolved population derived from CFB strain showed improved fermentative behavior than its parental strain.