Microbial biogeography of pit mud from an artificial brewing ecosystem on a large time scale: all roads lead to Rome

The fermentation process of strong-flavor Baijiu occurs in an underground cellar surrounded by mud, where grains are decomposed and generate new flavor substances. Pit mud microbiome plays a pivotal role in the formation of Baijiu flavor, which is positively correlated with cellar age. Previous research on the pit mud microbiome was often conducted for a specific distillery, making it hard to draw a general conclusion to decipher the pit mud microecosystem in such a unique brewing environment. Hence, in this study, we attempt to unravel community formation mechanisms through research in pit mud microbial biogeography based on 12 independent bacterial studies comprising qualified data sets of 302 samples clearly marked with cellar age from five major producing provinces. We found cellar age explained more variations in the bacterial community than geographical region. The bacterial community displayed evident dominant species alternations on a large time scale. Lactobacillus was found to be the dominant species in young pit mud (<10 years), while Caproiciproducens dominated in aged pit mud. Null model analysis revealed that as pit mud aged, the influence of stochastic and deterministic processes in the bacterial community shifted, and the relative contribution of homogeneous selection increased. Positive correlations dominated the bacterial co-occurrence networks, and the network complexity decreased and stability increased with cellar age. These findings shed new light on the formation of pit mud microbiomes under the impact of long-term batch-to-batch continuous brewing, providing a basis for the evaluation and production of high-quality pit mud. IMPORTANCE: Baijiu is a typical example of how humans employ microorganisms to convert grains into new flavors. Mud cellars are used as the fermentation vessel for strong-flavor Baijiu (SFB) to complete the decomposition process of grains. The typical flavor of SFB is mainly attributed to the metabolites of the pit mud microbiome. China has a large number of SFB-producing regions. Previous research revealed the temporal profiles of the pit mud microbiome in different geographical regions. However, each single independent study rarely yields a thorough understanding of the pit mud ecosystem. Will the pit mud microbial communities in different production regions exhibit similar succession patterns and structures under the impact of the brewing environment? Hence, we conducted research in pit mud microbial biogeography to uncover the impact of specific environment on the microbial community over a long time scale.