RSM based optimization of bioactive metabolites in culture supernatant derived from Bacillus stercoris strain GRS6 demonstrates significant enhancement of bioactive potential against Escherichia coli and Listeria monocytogenes

The present work focuses on the bioactivity of secondary metabolites secreted by Bacillus stercoris strain GRS6 against E. coli MTCC118 and L. monocytogenes MTCC657 which are commonly considered as gastrointestinal pathogens. This strain was isolated from the sediments of river Ganga at Haridwar (India) and its cell free supernatant demonstrated anti-microbial action against these two test organisms. As bioactivity of microbial metabolites is also a consequence of media composition and physiological conditions, therefore an attempt was made to optimize media and growth conditions of B. stercoris for subsequent enhancement of biological activity. The optimization studies were conducted by following One Factor at time (OFAT) and response surface methodology - Central composite design. The OFAT approach investigated the effect of various carbon and nitrogen sources along with different growth media. Apart from these, the critical physiological parameters such as pH (5-9), temperature (25 to 50℃) and incubation period (24 to 120 hrs.) were also carried out. Based on OFAT findings, A CCD matrix comprising of 26 experimental runs was generated as combinations of different growth parameters by Design Expert version 13.0. The percentage improvement against E. coli calculated as inhibition zone (IZ) was 21.5% in RSM optimization compared to that observed in OFAT approach. Similarly, RSM optimization improved bioactivity against L. monocytogenes by 24.9% as compared to OFAT. Spectroscopic and chromatographic identification of GRS6 bioactive metabolites revealed the presence of 3,3-([1,3] Dioxolan)-4,4-Dimethyl-7-Oxa-Bicyclo [4.1.0] Heptan and dodecanamide. The present study signifies the importance of RSM approach for enhancing the bioactivity against gastrointestinal pathogens.