Isolation and preliminary characterization of a bacteriocin-producer Bacillus strain inhibiting methicillin resistant Staphylococcus aureus.

Abstract

In a multivalent approach to discover new antimicrobial substance, a total of 160 Bacilli were isolated from termitarium soil, characterized on the basis of their morphological and physiological characters and screened for their antimicrobial activity by agar well diffusion method against certain drug resistant pathogenic bacteria such as Staphylococcus aureus, Methicillin resistant Staphylococcus aureus and common food contaminating bacteria Listeria monocytogenes. After preliminary screening, sixteen isolates showed inhibitory activity against test pathogens. Among them Bacillus isolate TSH58 exhibited maximum inhibitory activity against MRSA, Staphylococcus aureus and Listeria monocytogenes. Based on morphological, physiological, biochemical and 16S rDNA characteristics isolate TSH58 was identified as a member of the Bacillus cereus species group. Various nutrient sources and culture conditions were optimized, the partially purified antimicrobial metabolite was subjected to various treatments such as heat, pH and proteolytic enzymes. Complete loss in the activity observed when the crude metabolite was treated with proteolytic enzymes suggesting its proteinaceous nature and termed as bacteriocin like inhibitory substance (BLIS). Minimal inhibitory concentration of the partially purified bacteriocin determined by microtiter plate assay was 80 μg/ml for MRSA and 40 μg/ml for L. monocytogenes. Tricine SDS PAGE analysis revealed that the partially purified bacteriocin produced by the Bacillus strain TSH58 had an apparent molecular weight of about 4.0 KDa.