Molecular characterization of class IIa, heat-stable enterocin<i style=""> </i>produced by <i style="">Enterococcus faecium</i> MTCC 5153

Abstract

<i style=""> </i>The objective of this study was to provide molecular evidences for enterocin biosynthesis by <i>Enterococcus faecium</i> MTCC 5153. The culture filtrate (CF) of <i style="">E. faecium</i> MTCC 5153 exhibited a broad inhibitory spectrum against several enterococci, food-spoilage lactic acid bacteria (LAB) as well as pathogenic Gram-positive bacteria. The antimicrobial compound present in the CF showed the properties of class IIa bacteriocin. PCR was employed for taxonomic identification and detection of genes coding for virulence factors and different enterocins. Based on amplification of enterocin A (<i style="">ent</i>A) gene by PCR, additional characterization of enterocin A operon was carried out. Southern hybridization confirmed the presence of chromosomally encoded enterocin A (<i style="">ent</i>A) gene. These results suggest distribution of conserved enterocin A operon among different strains of <i style="">E. faecium</i>. Tricine SDS-PAGE activity gel assay in combination with mass spectral analysis indicated the production of single enterocin of 4.8 kDa in size. Production of enterocin was carried out using different sugars in tryptone, yeast extract (TYE) medium, under microaerophilic condition and the antimicrobial activity in the range of 10-50 x10³ arbitrary units per milliliter (AU mL^-1) was observed. This study indicates the potentiality of native isolate to produce broad spectrum bacteriocin which can be used to deal with spoilage bacteria and food-borne pathogens like listeria.