Additive-induced pH determines bacterial community composition and metabolome in traditional mustard seed fermented products

IntroductionKahudi and Kharoli are unique naturally fermented mustard seed products prepared and consumed in the northeastern region of India. The pre-fermentation processing of mustard seeds (soaking, pan-frying, mixing with alkaline or acidic additives, airtight packaging) renders a stringent fermentation environment. The metabolic activities of fermenting bacterial populations yield a myriad of glucosinolate-derived bioactive components which have not been described earlier. MethodsThis present study employed integrated 16S rRNA amplicon sequencing and LC-MS-based metabolomics to elucidate the bacterial diversity and metabolome of the two fermented mustard seed food products. Results and DiscussionUnivariate and multivariate analyses of metabolomics data revealed differential abundances of a few therapeutically-important metabolites viz., sinapine, indole-3-carbinol, gamma-linolenic acid in Kahudi, and metabolites viz., beta-sitosterol acetate, 3-butylene glucosinolate, erucic acid in Kharoli. A metagenomic investigation involving the 16S rRNA (V3-V4) amplicon sequencing showed the dominance of Firmicutes (99.1 +/- 0.18%) in Kahudi, and Firmicutes (79.6 +/- 1.92%) and Proteobacteria (20.37 +/- 1.94%) in Kharoli. The most abundant genera were Bacillus (88.7 +/- 1.67% in Kahudi; 12.5 +/- 1.75% in Kharoli) followed by Lysinibacillus (67.1 +/- 2.37% in Kharoli; 10.4 +/- 1.74% in Kahudi). Members of both these genera are well known for proteolytic and endospore-forming abilities which could have helped in colonizing and thriving in the stringent fermentation environments.