LAUSR

The microbial risk involved with natural food fermentation is largely unknown. Here, we report the prevalence of enteric bacterial pathogens in the traditional fermented foods marketed in Northeast region of India. A total of 682 samples of 39 food types (broadly categorized into fermented soybean, bamboo shoot, fish, milk and pork products) collected over four different seasons from seven states of India were analyzed in this study. Cultivation-independent analysis by MiSeq amplicon sequencing of V4-V5 region of the 16S rRNA gene showed the bacterial community structure in the foods. Among the WHO prioritized foodborne bacterial pathogens, we detected the prevalence of phylotypes related to Clostridium botulinum, Bacillus cereus, Staphylococcus aureus, Clostridium perfringens, Listeria monocytogenes, and Escherichia coli in these ethnic foods. We also observed the occurrence of other well known human enteric pathogens like Proteus mirabilis, Clostridium difficile, and Yersinia enterocolitica. Further pathogen-specific qPCR assays confirmed a higher population (>107 cells/g) of B. cereus, P. mirabilis, and a C. botulinum related phylotype in the fermented soybean, fish, and pork products. We noticed a general trend of higher pathogen occurrence during the colder months without any seasonal variation of total bacterial load in the fermented foods. Further qPCR analysis on toxigenic and pathogenic potential, and toxins production by immunoassays showed that all the soybean samples and the isolated B. cereus cultures were positive for diarrheal toxins (Nhe and Hb1), and nearly half of the samples were positive for emetic toxin (cereulide). Similarly, the food samples and associated swarming P. mirabilis cultures were positive with the pathogenic factors like hemolysin (hpm), urease (ure) and multidrug resistance. However, we could not confirm the presence of botulinum neurotoxin (toxins A, B, E, and F) in the C. botulinum positive food samples. This is the first baseline data of the enteric bacterial pathogens prevalent in the traditional fermented foods of India, which will support the sustained effort of WHO to estimate the global foodborne disease burden. The unusual presence of P. mirabilis in the fermented foods marketed in the Indian region with high incidence of urolithiasis cases is a concern. Our study emphasizes the need of the hour to have a coordinated action to control and prevent the spread of enteric bacterial pathogens through fermented foods marketed in India. Moreover, replacing the indigenous process with a defined starter culture based controlled fermentation will enhance the safety of Indian fermented foods.