Soaking reshapes the structure, function, and co-occurrence patterns of barley surface bacterial communities.

Surface microbiota on cereal grains can influence fermentation outcomes and product quality in food processing. To better understand microbial responses during pre-processing, we analyzed the succession of bacterial communities on barley seeds from four cultivars over five soaking stages (0, 1, 2, 4, and 8 days). Using 16S rRNA gene sequencing, functional prediction, and ecological modeling, we observed a sharp decline in bacterial diversity and a compositional shift toward fermentative Firmicutes, particularly Lactobacillales. Functional profiles indicated reduced metabolic breadth and enhanced expression of fermentation- and transport-related pathways over time. Community assembly analysis indicated increasing dominance of stochastic factors-primarily limited dispersal-over time. Co-occurrence networks became simpler and shifted toward more negative associations over time. Our findings suggest that soaking not only reshapes microbial structure and function but also alters ecological stability, providing a scientific foundation for microbial management strategies in malting, germination, and cereal-based fermentation processes.