Hybridization and adaptive evolution of diverse species for cellulosic biofuel production.

Background: Lignocellulosic biomass is a common resource across the globe, and its fermentation offers a promising option for generating renewable liquid transportation fuels. The deconstruction of lignocellulosic biomass releases sugars that can be fermented by microbes, but these processes also produce fermentation inhibitors, such as aromatic acids and aldehydes. Several research projects have investigated lignocellulosic biomass fermentation by the baker's yeast . Most projects have taken synthetic biological approaches or have explored naturally occurring diversity in to enhance stress tolerance, xylose consumption, or ethanol production. Despite these efforts, improved strains with new properties are needed. In other industrial processes, such as wine and beer fermentation, interspecies hybrids have combined important traits from multiple species, suggesting that interspecies hybridization may also offer potential for biofuel research.

Results: To investigate the efficacy of this approach for traits relevant to lignocellulosic biofuel production, we generated synthetic hybrids by crossing engineered xylose-fermenting strains of with wild strains from various species. These interspecies hybrids retained important parental traits, such as xylose consumption and stress tolerance, while displaying intermediate kinetic parameters and, in some cases, heterosis (hybrid vigor). Next, we exposed them to adaptive evolution in ammonia fiber expansion-pretreated corn stover hydrolysate and recovered strains with improved fermentative traits. Genome sequencing showed that the genomes of these evolved synthetic hybrids underwent rearrangements, duplications, and deletions. To determine whether the genus contains additional untapped potential, we screened a genetically diverse collection of more than 500 wild, non-engineered isolates and uncovered a wide range of capabilities for traits relevant to cellulosic biofuel production. Notably, strains have high innate tolerance to hydrolysate toxins, while some species have a robust native capacity to consume xylose.

Conclusions: This research demonstrates that hybridization is a viable method to combine industrially relevant traits from diverse yeast species and that members of the genus beyond may offer advantageous genes and traits of interest to the lignocellulosic biofuel industry.