Aspergillus-Enterobacteriaceae dynamics and branched-chain amino acid metabolism in fermented fish sauce

Fish sauce is a traditional fermented aquatic condiment valued for its unique umami and complex flavor. Its flavor formation involves intricate microbial interactions and metabolic pathways that remain not fully clarified. This study aimed to unravel the core mechanism by focusing on Aspergillus-Enterobacteriaceae synergy and branched-chain amino acid (BCAA) catabolism. A 36-day fermentation was monitored via physicochemical analysis, microbial profiling, untargeted metabolomics, volatile compound detection, and proteomics. Fungal ITS analysis identified Aspergillus as the dominant genus, whereas Enterobacteriaceae formed the bacterial core. Their co-occurrence was associated with complementary metabolic functions. Aspergillus abundance was correlated with enhanced proteolytic and lipolytic activity, releasing BCAAs and fatty acids. Enterobacteriaceae dominance coincided with activation of BCAA catabolism via the Ehrlich pathway, converting leucine, isoleucine, and valine into key flavor precursors. Proteomics confirmed upregulation of exopeptidase, BCAA degradation, and ester metabolism enzymes. Volatile compounds showed an orderly transition, with aldehydes in the initial stage, alcohols and ketones in the mid-stage, and esters in the late stage. Key flavor markers, including 3-methyl-1-butanol and ethyl heptanoate, are linked closely to BCAA catabolism and microbial synergy. This study demonstrates that Aspergillus-Enterobacteriaceae co-occurrence is associated with BCAA catabolism and flavor compound profiles in fish sauce, providing molecular-level insights for quality optimization of traditional fermented fish products.