Proso millet, a high-quality fermentation material used for Chinese yellow wine production, can produce special flavored substances; however, its role in improving the flavor and altering microbial communities of light-flavored Baijiu during fermentation remain unknown. Thus, we aimed to investigate the effect of proso millet on improving the flavor of light-flavored Baijiu and altering microbial communities during different fermentation stages. The dynamic changes in the microbial communities and flavor of proso millet (50%) + sorghum (50%) mixed fermentation samples were analyzed through intermittent sampling on days 7, 14, 21, and 28 of the fermentation process. Microbial high-throughput sequencing and the analysis of flavor characteristics were conducted through 16S DNA/ ITS amplicon sequencing and gas chromatography (multi-capillary column)-ion mobility spectrometry, respectively. Proso millet significantly changed the core flavor compound composition of traditional light-flavored Baijiu from ethyl acetate, ethyl hexanoate, ethyl hexanoate dimer, ethyl butanoate, ethyl lactate, and butyl acetate to oct-2-ene, 2-butanol, propyl propanoate, 2-pentenal, and 4-methylpentanal. The amplicon sequencing analysis revealed that the alpha diversity parameters of bacterial and fungal communities, including the Chao1, Pielou_e, Shannon, and Simpson indices, for proso millet-sorghum mixed fermentation samples were significantly higher than those for sorghum fermentation samples (p < 0.05). Of the 40 most significant microbial genera in two treatments, proso millet significantly increased the abundance of 12 bacterial and 18 fungal genera. Among the 40 most significant bacterial and fungal species, 23 bacterial species belonged to the Lactobacillus genus, whereas the 30 primary fungal species belonged to 28 different genera. The analysis of the relationship between microbial changes and the main flavor compounds of light-flavored Baijiu showed that bacteria from the Weissella, Acinetobacter, Bacteroides, Psychrobacter, Pseudarthrobacter, Lactococcus, Chloroplast, Saccharopolyspora, Psychrobacter, Saccharopolyspora, Pseudonocardiaceae, Bacteroides genera and fungi from the Thermoascus, Aspergillus, Pichia, Rhizomucor, Papiliotrema, Hyphopichia, and Mucor genera significantly inhibited the synthesis of ethyl hexanoate, ethyl butanoate, ethyl lactate ethyl lactate, and butyl acetate but increased the synthesis of ethyl acetate (p < 0.05). Moreover, these microbes exhibited a significantly greater abundance in proso millet-sorghum mixed fermentation samples than in sorghum samples. The synthesis of special flavored compounds in proso millet Baijiu was significantly positively correlated with the presence of fungi from the Rhizopus, Papiliotrema, Wickerhamomyces, Aspergillus, and Thermoascus genera but negative correlated with the presence of bacteria from the Weissella, Acinetobacter, Psychrobacter, Pseudarthrobacter, Bacteroides, and Saccharopolyspora genera. Regarding ethanol content, the low alcohol content of Fenjiu may be due to the significantly high abundance of fungi from the Psathyrella genus and bacteria from the Staphylococcus, Kroppenstedtia, Brevibacterium, and Acetobacter genera during fermentation. In summary, proso millet significantly altered the flavor of light-flavored Baijiu by inducing the formation of a special microbial community; however, it did not increase alcohol concentration. This study lays the foundation for future research on Baijiu fermentation. Additionally, the study findings may help improve the production efficiency and elevate the quality and flavor of the final product.
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