Development of a new brewing strain with improved aroma through hybridization and UV mutagenesis

Yeasts are fundamental in beer production, influencing fermentation, flavor, and stability. To meet consumer demand, we developed a novel, GMO-free yeast hybrid between a sourdough strain of Saccharomyces cerevisiae and a cryotolerant S. eubayanus x S. bayanus via spore-to-spore hybridization. Microfermentations showed improved brewing traits in one hybrid but also the undesired Phenolic Off-Flavor (POF+) phenotype, caused by the conversion of ferulic acid into 4-vinyl guaiacol (4VG). To eliminate this trait, we applied UV mutagenesis, optimizing irradiation conditions (50 cm distance, 30 s exposure) to achieve 8-22% mortality. Nystatin treatment, which selects slow-growing mutants, reduced the mutation success rate (13.3% without vs. 0.4% with nystatin). Mutants were screened on YPDA with 4 mM ferulic acid and analyzed via Ferulic Acid Conversion Index (FACI) and Cinnamic Acid Sensitivity Index (CASI). POF- candidates exhibited low FACI and high CASI values, confirming reduced 4VG production. Further microfermentations confirmed that selected mutants retained improved brewing traits without the POF+ phenotype. This study demonstrates that hybridization combined with UV mutagenesis is an effective, GMO-free strategy for developing superior brewing yeasts.