This study investigated the effects of different methods of Bacillus inoculation (separate cultures, co-culture, and no inoculation) on the sensory characteristics and flavor pyrazine compounds of high-temperature Daqu. During the fermentation process, total acidity and moisture content decreased progressively, and sensory attributes of Bacillus-fortified Daqu were superior to those of the control group. For finished Daqu, the total content of pyrazine compounds was significantly higher in the experimental groups (3376.89 μg/g for fortified Daqu Ⅱ, 2842.47 μg/g for fortified Daqu Ⅰ) than in the control group (1868.78 μg/g). A total of 24 prokaryotic and 16 eukaryotic genera were identified as dominant microbiota. The abundance of Bacillus, Thermomyces, and Thermoascus in fortified Daqu was higher than that in the control group. A total of 21 prokaryotic and 19 eukaryotic differentially abundant microorganisms were identified. In summary, the inoculation of Bacillus seed culture contributed to improved sensory attributes, enhanced microbial diversity to some extent, and increased the concentration of pyrazine compounds, thereby improving the overall quality of high-temperature Daqu.

This study explored the effect of Bacillus subtilis JP1, isolated from the fermentation process of Nongxiangxing Baijiu, on the assembly and succession of microbial communities in fermented grains (Jiupei). A mixed-culture fermentation system was constructed using Jiupei and yellow water from Nongxiangxing Baijiu production. A bio-perturbation experiment was conducted by adding B. subtilis JP1 to the system. Amplicon sequencing combined with null model analysis was used to investigate the structure, composition and assembly process of microbial communities. R language was employed to calculate the environmental niche breadth of microorganisms to analyze potential reasons for changes in microbial community assembly. The results showed that compared with the control group, B. subtilis JP1 reduced the microbial diversity during fermentation while increasing the relative abundance of ethanol-producing microorganisms. Additionally, B. subtilis JP1 decreased the environmental niche breadth of the microbial community, leading to a significant increase in the number of rare and conditionally rare taxa. The interactions among microorganisms were enhanced, and the complexity of the microbial network was increased. These changes collectively influenced the assembly process of the microbial community during fermentation. The addition of B. subtilis JP1 increased the contribution of heterogeneous selection for bacteria and homogeneous selection for fungi to the microbial community assembly process. Furthermore, it accelerated bacterial succession while slowing down fungal succession. This study elucidates the mechanism of the effect of B. subtilis on the establishment of microbial communities in fermented grains for Nongxiangxing Baijiu, providing a theoretical basis for the targeted regulation of Bacillus during the brewing process of Nongxiangxing Baijiu.