Robust Domination of Lactobacillus sakei in Microbiota During Traditional Japanese Sake Starter Yamahai-Moto Fermentation and the Accompanying Changes in Metabolites

The successful production of sake (Japanese rice wine) is brought about by drastic changes in microbial flora and chemical components during fermentation. In the traditional manufacturing process of sake starter (yamahai-moto), spontaneous growth of lactic acid bacteria suppresses inappropriate microorganisms and prepares the optimum environment for the alcohol fermentative yeast. In this study, we analyzed the changes in bacterial flora and chemical components of yamahai-moto. High-throughput next-generation sequencing (NGS) of the 16S ribosomal RNA gene V4 region revealed that various kinds of bacteria, including nitrate-reducing bacteria, existed in the early fermentation stage; however, Lactobacillus sakei then increased drastically to become dominant in the middle stage. Interestingly, this result was different from that obtained in the previous year at the same manufacturer; the early-stage major bacterium was Lactobacillus acidipiscis. Lactic acid, glucose, isomaltose, and total free amino acids increased throughout the fermentation process, which was attributable to the metabolism of L. sakei and the koji mold. It is noteworthy that significant ornithine accumulation and arginine consumption were observed from the middle to late stages. Thirty-eight percent of the L. sakei isolates from yamahai-moto exhibited significant ornithine production, indicating that the arginine deiminase pathway of L. sakei was working to survive the extremely low pH environment of the moto after the middle stage. This is the first report that includes concurrent analyses of the NGS-based bacterial flora and chemical components of yamahai-moto, providing further knowledge to help understand and improve the process of sake brewing.