Bacteria producing hydrolytic exoenzymes are of great importance considering their contribution to the host metabolism as well as for their various applications in industrial bioprocesses. In this work hydrolytic capacity of bacteria isolated from the gastrointestinal tract of Bombay duck (Harpadon nehereus) was analyzed and the enzyme-producing bacteria were genetically characterized. A total of twenty gutassociated bacteria, classified into seventeen different species, were isolated and screened for the production of protease, lipase, pectinase, cellulase and amylase enzymes. It was found that thirteen of the isolates could produce at least one of these hydrolytic enzymes among which protease was the most common enzyme detected in ten isolates; lipase in nine, pectinase in four, and cellulase and amylase in one isolate each. This enzymatic array strongly correlated to the previously reported eating behavior of Bombay duck. 16S rRNA gene sequence-based taxonomic classification of the enzyme-producing isolates revealed that the thirteen isolates were grouped into three different classes of bacteria consisting of eight different genera. Staphylococcus, representing ∼46% of the isolates, was the most dominant genus. Measurement of enzyme-production via agar diffusion technique revealed that one of the isolates which belonged to the genus Exiguobacterium, secreted the highest amount of lipolytic and pectinolytic enzymes, whereas a Staphylococcus species produced highest proteolytic activity. The Exiguobacterium sp. expressing a maximum of four hydrolases, appeared to be the most promising isolate of all.

Lactic acid bacteria (LAB) with prominent antimicrobial effects against pathogens have been reported in several milk-based and plant-based foods. Borhani is a popular beverage prepared from the ingredients of both dairy and plant origins and is believed to be highly beneficial for health. Herein, we report the pathogen-inhibitory activity of two borhani-associated lactic acid bacteria (LAB), Limosilactobacillus fermentum strain LAB-1 and Levilactobacillus brevis strain LAB-5. Their antimicrobial activity was primarily assessed using the cell free supernatant (CFS) by agar diffusion technique in which both strains showed strong antimicrobial effects against several pathogenic and spoilage microorganisms including Acinetobacter baumannii, Bacillus cereus, Bacillus subtilis, Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Salmonella abony, Salmonella typhi, Shigella flexneri, and Staphylococcus aureus. The L. fermentum strain with its ability to inhibit all the target pathogens appeared to be more effective with larger inhibition-zone formation as compared to the L. brevis strain which also successfully inhibited all pathogens but had relatively little effects on A. baumannii. The extent of their inhibitory effect was further assessed by co-culture inhibition assay in which growth of the test microbes was monitored for 24 hours in presence of the CFS. The CFS of both lactic acid bacteria could effectively inhibit growth of the pathogenic microbes for a significant period of time. While the L. fermentum strain could almost completely stop growth of all test organisms, the L. brevis strain was particularly effective against Shigella flexneri and the Salmonella species. Our study, therefore, suggests the presence of beneficial lactic acid bacteria in borhani which can be of important use as antimicrobial agents in functional foods and therapeutics to help acquire protection against drug resistant pathogens.

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