Network-based insights into the temperature-driven mechanisms of bacterial self-assembly in the solid-state fermented vinegar seed Pei

Seed Pei, the fermentation starter used in the traditional solid-state acetic acid fermentation of vinegar, is produced via cyclic inoculation. Although environmental temperature is a known driver of microbial community dynamics in fermented foods, its role in shaping the assembly of seed Pei microbiota remains unclear. In this study, we simulated the assembly process of bacterial communities in seed Pei from the same source over five successive inoculation rounds under three temperatures (25 °C, 30 °C, and 35 °C). Results showed no significant differences in total acid content at the fermentation endpoint, though minor differences in non-volatile acid content were observed. Bacterial communities stabilized by the third round, displaying similar taxa but with significant structural divergence across temperatures. Distinct lactic acid and acetic acid bacteria dominated under each temperature, consistent with their thermal adaptation profiles. Molecular ecological network analysis revealed that self-assembled communities under different temperatures exhibited comparable stability but varied in complexity. Lower temperature (25 °C) generated the most complex ecological networks with the lowest maximum vulnerability, while networks at 30 °C and 35 °C exhibited reduced complexity but higher connectivity. These findings suggest that during microbial assembly in seed Pei, environmental temperature acts as a thermal filter that preferentially replaces original taxa with metabolically similar acid-producing species. Additionally, keystone species contributed to community structuring by mediating interspecies interactions. This study provides new insights into temperature-driven self-assembly of seed Pei bacterial communities and informs the rational design of functional fermentation starters.