Asci and ascospores of Saccharomyces kudriavzevii ZP 591.
© J.P. Sampaio 2008

In spite of the extensive research on Saccharomyces yeasts, little is known about their natural habitats and population genetics. Traditionally S. paradoxus has been regarded as a wild species associated mostly with natural habitats. On the contrary, several authors viewed S. cerevisiae as a domesticated organism chiefly adapted to man-made fermentations and normally absent in natural ecosystems. However, several lines of evidence suggest that S. cerevisiae existed in natural environments long before it was utilized in man-made fermentations. Because ecological data on Saccharomyces is fragmentary and in some cases is based on erroneous species identifications, the habitats of most species remain unknown.

Recently, we found an unprecedented diversity of Saccharomyces sensu stricto species associated with oak trees in Portugal. Among these, a European population of S. kudriavzevii, a species thus far thought to be endemic of Japan, was identified.

Sampling site at Castelo de Vide, Portugal.
© J.P. Sampaio 2008

Isolation of Saccharomyces spp. from oak bark – enrichment step.
© J.P. Sampaio 2008

Temperature adaptation and ecological speciation in Saccharomyces 

Our general goal is to understand how the various Saccharomyces species evolved from their primal Saccharomyces ancestor and what forces drove speciation in this group. We are also interested in understanding the particularities of each present-day species i.e., we would like to know the set of species-specific characteristics - those phenotypic traits that make each species unique. This issue is of particular relevance because (i) the phenotypes of the different Saccharomyces species are very similar, and (ii) according to the principle of niche exclusion, if more than one species exist in the same ecological niche, competition will occur and one species will outcompete and exclude the others. Therefore, if more than one Saccharomyces species are found in the same habitat, what kinds of ecological speciation evens have taken place during the course of evolution?

We hypothesize that temperature adaptation played a crucial role in the phenotypic evolution of sympatric Saccharomyces sensu stricto species. This would explain why S. cerevisiae (thermophilic) and S. kudriavzevii (cryotolerant) are able to apparently evade the competitive exclusion principle and to consistently establish sympatric associations on tree bark.

Galactose utilization in S. kudriavzevii 

Prior to this study, only four S. kudriavzevii strains were known and all exhibited one important trait that distinguished them from other Saccharomyces species: they were unable to grow on galactose. We observed that the Portuguese S. kudriavzevii population exhibits important genetic differences when compared with the strains of this species previously known. The most striking difference is undoubtedly the presence of functional GAL genes in the Portuguese population. It was shown that the inability of the type strain of S. kudriavzevii to grow on galactose was due to extensive and ancient gradual degeneration of the entire set of genes involved in galactose utilization, rather than from a discrete mutation event limited in time that could have taken place very recently (Hittinger et al. 2004). Calculations taking into account the neutral mutation rates in S. kudriavzevii, date GAL pathway degeneration to a period immediately following separation of the lineage leading to S. kudriavzevii. This is difficult to reconcile with the fact that the Portuguese isolates have otherwise only modest sequence divergence with respect to the type strain.

Genomes and the evolution of phenotypes

We are using a genomic approach aiming at identifying genes likely to be involved in phenotypic evolution via disruptive selection and character displacement. The ratios of nonsynonymous versus synonymous nucleotide substitutions were used to perform genome-level pairwise comparisons of genetic differences. We found that while most genes appear to evolve at a similar pace in all species, some exhibit striking differences between the different genomes studied. This analysis allowed us to highlight a set of proteins that is significantly more divergent in S. kudriavzevii and/or S. uvarum than in S. paradoxus, when these genomes are compared to that of S. cerevisiae. Within this set of divergent proteins, some functional classes were found to be particularly well represented.