Rapid selection response to ethanol in Saccharomyces eubayanus emulates the domestication process under brewing conditions

Abstract

Although the typical genomic and phenotypic changes that characterize the evolution of organisms under the human domestication syndrome represent textbook examples of rapid evolution, the molecular processes that underpin such changes are still poorly understood. Domesticated yeasts for brewing, where short generation times and large phenotypic and genomic plasticity were attained in a few generations under selection, are prime examples. To experimentally emulate the lager yeast domestication process, we created a genetically complex (panmictic) artificial population of multiple Saccharomyces eubayanus genotypes, one of the parents of lager yeast. Then, we imposed a constant selection regime under a high ethanol concentration in 10 replicated populations during 260 generations (6 months) and compared them with propagated controls exposed solely to glucose. Propagated populations exhibited a selection differential of 60% in growth rate in ethanol, mostly explained by the proliferation of a single lineage (CL248.1) that competitively displaced all other clones. Interestingly, the outcome does not require the entire time-course of adaptation, as four lineages monopolized the culture at generation 120. Sequencing demonstrated that de novo genetic variants were produced in all propagated lines, including SNPs, aneuploidies, INDELs and translocations. In addition, the different propagated populations showed correlated responses resembling the domestication syndrome: genomic rearrangements, faster fermentation rates, lower production of phenolic offflavours and lower volatile compound complexity. Expression profiling in beer wort revealed altered expression levels of genes related to methionine metabolism, flocculation, stress tolerance and diauxic shift, likely contributing to higher ethanol and fermentation stress tolerance in the evolved populations. Our study shows that experimental evolution can rebuild the brewing domestication process in 'fast motion' in wild yeast, and also provides a powerful tool for studying the genetics of the adaptation process in complex populations.

A Saccharomyces eubayanus haploid resource for research studies

“…Also, we used four different S. cerevisiae strains that represent the main lineages described in the species: YPS128 (North American, NA), DBVPG6044 (West African, WA), Y12 (Sake, SA) and DBVPG6765 (Wine/European, WE) (Supplementary Table S10 ) 8 . The number of generations was determined using the formula log 2 (final cells/1.5 × 10 6 ) 102 . Finally, after each fermentation, the proportions of diploid and haploid cells were determined by colony PCR of colonies isolated for each replicate on solid YPD medium, as described above.…”

Microbial Adaptation to Enhance Stress Tolerance

“…In addition, adaptation can rapidly improve tolerance to stress. adaption of Saccharomyces eubayanus in the culture with ethanol boosted the strain growth under ethanol condition ( Mardones et al, 2021 ). Adaptation exhibits a great ability to improve microbial performance in a short time.…”

Natural Variation in Diauxic Shift between Patagonian Saccharomyces eubayanus Strains

“…Fermentations were carried out in three biological replicates as previously described (30). Briefly, 12 °Plato (°P) beer wort was oxygenated (15 mg L -1 ) and supplemented with 0.3 ppm Zn 2+ (as ZnCl 2 ).…”

“…RNA integrity was confirmed using a Fragment Analyzer (Agilent). RNA library preparation and Illumina sequencing were performed in the BGI facilities (Hong Kong, China) as previously described (30).…”

“…This sugar represses the expression of genes involved in the consumption of more complex sugars such as maltose, as well as genes involved in respiration, a phenomenon known as ‘glucose repression’ (29). Glucose depletion activates a metabolic rewiring triggering the metabolism of other sugars, a process called diauxic shift, which may explain differences in the adaptation time between different S. eubayanus strains (30, 31). Then, a detailed molecular analysis of the diauxic shift in S. eubayanus strains is fundamental for the potential use of this wild yeast in the industry.…”