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Rice  koji  is a raw material used in the production of  sake ; however, details regarding the microbial flora and their dynamics in rice  koji  during the production process are poorly understood. Clarifying these issues can contribute to proposing a method and evaluation that will improve the quality of rice  koji  and  sake . The aim of this study was to determine the microflora in rice  koji  and the effectiveness of the traditional microbial control techniques used in the  sake  production process. We analyzed the diversity and changes in bacterial flora during rice  koji  production by amplicon sequencing of the 16S rRNA gene. The predominant taxon in all rice  koji  samples was family  Staphylococcaceae . The microbial population and the changes in its distribution for five consecutive stages in rice  koji  production were examined by direct colony counting. Bacteria counts in all samples were below the limit of detection initially, then increased rapidly toward the final stage. The predominant bacterial colonies from all samples were yellow and were identified as  Staphylococcus gallinarum  through 16S rRNA gene sequence similarity. The  S. gallinarum  isolates exhibited faster growth in pregelatinized rice medium. Interestingly, the growth of  S. gallinarum  isolates was suppressed by low temperature (12 °C), ethanol concentration (≥6%) and the addition of lactic acid, which are traditional microbial control methods used during  sake  fermentation. Therefore, proper control of the traditional  sake  production process can effectively inhibit the growth of undesirable bacteria such as  S. gallinarum  that emerge during the production of rice  koji .

Traditional microbial control methods used in sake brewing effectively suppress predominant bacteria emerging during production of rice koji