Integrated microbiome and metaproteome analyses reveal microbial functions underlying traditional fermented amaranth stem fermentation

Fermented amaranth stems are a traditional Chinese fermented vegetable characterized by a distinctive pungent flavor and complex microbial ecology. This study compared samples from Anhui (AH), Jiangsu (JS), and Zhejiang (ZJ) using physicochemical analysis, volatile profiling, free amino acid (FAA) determination, 16S rRNA sequencing, and metaproteomics. Across the three regions, pH ranged from 4.08 to 4.39, total titratable acidity from 2.7 to 3.5 mL, viable counts from 7.7 to 8.6 log CFU/g, and total volatile concentrations from 99,021 to 332,864 μg/L. After removing chloroplast- and mitochondrial-derived reads, Firmicutes (approximately 56–77%) and Proteobacteria (approximately 23–40%) dominated the bacterial communities, with Lactobacillus, Lactococcus, and Pseudomonas as the major genera. Total FAA concentrations were 888.5, 778.0, and 582.0 μg/mL in AH, JS, and ZJ, respectively. Metaproteomics identified 7,158 unique protein accession records, of which 3,926 received taxonomic assignments. Functional annotation highlighted carbohydrate metabolism, energy metabolism, stress-response functions, and carbohydrate-active enzymes. These results demonstrate region-associated differences in bacterial composition, expressed functions, and flavor-related traits and provide candidate microbial–functional links for future targeted validation.