Isolation and characterization of Zygosaccharomyces sp. yeast strains from miso

There is currently great interest in the salt-tolerant yeast strains used to produce miso and soy sauce. Since the isolation of Zygosaccharomyces sp. strain from Japanese miso more than 60 years, several hybrid strains have been identified in fermented foods. Studies have shown that the active mating-type locus of the original Zygosaccharomyces sp. yeast strain is located between the T-subgenome sequence and the P-subgenome sequence. In this study, 32 salt-tolerant Zygosaccharomyces sp. yeast strains were isolated from five miso factories in Hiroshima Prefecture, Japan. Analysis by flow cytometry revealed that 27 strains were diploid and five strains were haploid. PCR analysis indicated that the 27 diploid strains had the same chromosomal structure of the active mating-type (MAT) locus as the original yeast strain isolated from miso 60 years ago. In addition, the 27 diploid strains were allodiploid, namely, natural hybrids of Z. rouxii and a related species, while the five haploid strains were all Z. rouxii. We found that cells of yeast strains isolated from miso changed morphologically when co-cultured with a yeast strain of opposite mating-type under nitrogen starvation conditions. The DNA sequence of the active mating-type locus and the results of cell morphology changes by co-culture were consistent with the mating type of each strain shown in the mating experiments. These findings will be useful for the future production of miso and soy sauce.