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The wild, cold-adapted parent of hybrid lager-brewing yeasts, Saccharomyces eubayanus, has a complex and understudied natural history. The exploration of this diversity can be used both to develop new brewing applications and to enlighten our understanding of the dynamics of yeast evolution in the wild. Here, we integrate whole genome sequence and phenotypic data of 200 S. eubayanus strains, the largest collection known to date. S. eubayanus has a multilayered population structure, consisting of two major populations that are further structured into six subpopulations. Four of these subpopulations are found exclusively in the Patagonian region of South America; one is found predominantly in Patagonia and sparsely in Oceania and North America; and one is specific to the Holarctic ecozone. Plant host associations differed between subpopulations and between S. eubayanus and its sister species, Saccharomyces uvarum. S. eubayanus is most abundant and genetically diverse in northern Patagonia, where some locations harbor more genetic diversity than is found outside of South America, suggesting that northern Patagonia east of the Andes was a glacial refugium for this species. All but one subpopulation shows isolation-by-distance, and gene flow between subpopulations is low. However, there are strong signals of ancient and recent outcrossing, including two admixed lineages, one that is sympatric with and one that is mostly isolated from its parental populations. Using our extensive biogeographical data, we build a robust model that predicts all known and a handful of additional regions of the globe that are climatically suitable for S. eubayanus, including Europe where host accessibility and competitive exclusion by other Saccharomyces species may explain its continued elusiveness. We conclude that this industrially relevant species has rich natural diversity with many factors contributing to its complex distribution and natural history., This material is based upon work supported by the National Science Foundation (https://www.nsf.gov) under Grant Nos. DEB-1253634 (to CTH) and DGE-1256259 (Graduate Research Fellowship to QKL), the USDA National Institute of Food and Agriculture (https://www.nifa.usda.gov) Hatch Project Nos. 1003258 and 1020204 to CTH, and in part by the DOE Great Lakes Bioenergy Research Center (DOE BER Office of Science (https://www.energy.gov/science/office-science) Nos. DE-SC0018409 and DE-FC02-07ER64494 to Timothy J. Donohue). QKL was also supported by the Predoctoral Training Program in Genetics, funded by the National Institutes of Health (5T32GM007133, https://www.nih.gov). DP is a Marie Sklodowska-Curie fellow of the European Union’s Horizon 2020 (https://ec.europa.eu/programmes/horizon2020/en) research and innovation program (Grant Agreement No. 747775). DL was supported by CONICET (PIP11220130100392CO, https://www.conicet.gov.ar), FONCyT (PICT 3677, PICT 2542, https://www.argentina.gob.ar/ciencia/agencia/fondo-para-la-investigacion-cientifica-y-tecnologica-foncyt), Universidad Nacional del Comahue (B199, https://www.uncoma.edu.ar), and NSF-CONICET grant (https://www.conicet.gov.ar). CTH is a Pew Scholar in the Biomedical Sciences and H. I. Romnes Faculty Fellow, supported by the Pew Charitable Trusts (https://www.pewtrusts.org/en) and Office of the Vice Chancellor for Research and Graduate Education (https://research.wisc.edu) with funding from the Wisconsin Alumni Research Foundation (WARF, https://www.warf.org), respectively.

Cell-cycle checkpoints and DNA repair processes protect organisms from potentially lethal mutational damage. Compared to other budding yeasts in the subphylum Saccharomycotina, we noticed that a lineage in the genus Hanseniaspora exhibited very high evolutionary rates, low Guanine–Cytosine (GC) content, small genome sizes, and lower gene numbers. To better understand Hanseniaspora evolution, we analyzed 25 genomes, including 11 newly sequenced, representing 18/21 known species in the genus. Our phylogenomic analyses identify two Hanseniaspora lineages, a faster-evolving lineage (FEL), which began diversifying approximately 87 million years ago (mya), and a slower-evolving lineage (SEL), which began diversifying approximately 54 mya. Remarkably, both lineages lost genes associated with the cell cycle and genome integrity, but these losses were greater in the FEL. E.g., all species lost the cell-cycle regulator WHIskey 5 (WHI5), and the FEL lost components of the spindle checkpoint pathway (e.g., Mitotic Arrest-Deficient 1 [MAD1], Mitotic Arrest-Deficient 2 [MAD2]) and DNA-damage–checkpoint pathway (e.g., Mitosis Entry Checkpoint 3 [MEC3], RADiation sensitive 9 [RAD9]). Similarly, both lineages lost genes involved in DNA repair pathways, including the DNA glycosylase gene 3-MethylAdenine DNA Glycosylase 1 (MAG1), which is part of the base-excision repair pathway, and the DNA photolyase gene PHotoreactivation Repair deficient 1 (PHR1), which is involved in pyrimidine dimer repair. Strikingly, the FEL lost 33 additional genes, including polymerases (i.e., POLymerase 4 [POL4] and POL32) and telomere-associated genes (e.g., Repressor/activator site binding protein-Interacting Factor 1 [RIF1], Replication Factor A 3 [RFA3], Cell Division Cycle 13 [CDC13], Pbp1p Binding Protein [PBP2]). Echoing these losses, molecular evolutionary analyses reveal that, compared to the SEL, the FEL stem lineage underwent a burst of accelerated evolution, which resulted in greater mutational loads, homopolymer instabilities, and higher fractions of mutations associated with the common endogenously damaged base, 8-oxoguanine. We conclude that Hanseniaspora is an ancient lineage that has diversified and thrived, despite lacking many otherwise highly conserved cell-cycle and genome integrity genes and pathways, and may represent a novel, to our knowledge, system for studying cellular life without them., A lineage in the budding yeast genus Hanseniaspora has remarkably high evolutionary rates, low GC content, small genome sizes, and lower gene numbers than the rest of the subphylum. Analysis of 25 genomes reveals that this lineage has survived and diversified for tens of millions of years despite lacking dozens of cell cycle and DNA repair genes normally thought to be essential.

Lager-style beers constitute the vast majority of the beer market, and yet, the genetic origin of the yeast strains that brew them has been shrouded in mystery and controversy. Unlike ale-style beers, which are generally brewed with Saccharomyces cerevisiae, lagers are brewed at colder temperatures with allopolyploid hybrids of Saccharomyces eubayanus x S. cerevisiae. Since the discovery of S. eubayanus in 2011, additional strains have been isolated from South America, North America, Australasia, and Asia, but only interspecies hybrids have been isolated in Europe. Here, using genome sequence data, we examine the relationships of these wild S. eubayanus strains to each other and to domesticated lager strains. Our results support the existence of a relatively low-diversity (π = 0.00197) lineage of S. eubayanus whose distribution stretches across the Holarctic ecozone and includes wild isolates from Tibet, new wild isolates from North America, and the S. eubayanus parents of lager yeasts. This Holarctic lineage is closely related to a population with higher diversity (π = 0.00275) that has been found primarily in South America but includes some widely distributed isolates. A second diverse South American population (π = 0.00354) and two early-diverging Asian subspecies are more distantly related. We further show that no single wild strain from the Holarctic lineage is the sole closest relative of lager yeasts. Instead, different parts of the genome portray different phylogenetic signals and ancestry, likely due to outcrossing and incomplete lineage sorting. Indeed, standing genetic variation within this wild Holarctic lineage of S. eubayanus is responsible for genetic variation still segregating among modern lager-brewing hybrids. We conclude that the relationships among wild strains of S. eubayanus and their domesticated hybrids reflect complex biogeographical and genetic processes., Author Summary Yeasts are key industrial microbes, most notably Saccharomyces cerevisiae, which is used to make a variety of products, including bread, wine, and ale-style beers. However, lager-style beers are brewed with interspecies hybrids of S. cerevisiae x Saccharomyces eubayanus. After its discovery in South America in 2011, rare strains of S. eubayanus have also been isolated outside of South America. Here we compare the genome sequences of several new and recent isolates of S. eubayanus from South America, North America, Australasia, and Asia to unravel the relationships of these wild isolates and their domesticated European hybrids. Two South American populations have the highest genetic diversity. One of these populations is closely related to a relatively low-diversity lineage that is spread across the Northern Hemisphere and includes the S. eubayanus parents of lager yeasts. Interestingly, we find that none of the wild isolates of S. eubayanus is the sole closest relative of lager-brewing hybrids. Instead, we show that standing variation among wild S. eubayanus strains contributed to the genetic makeup of lager yeasts. Our findings highlight the complex ancestries of lager yeasts and the importance of broader sampling of wild yeasts to illuminate our understanding of the sources of genetic variation among industrial hybrids.