Characterization of bacteriocin related genes discovered in a novel probiotic isolate Enterococcus faecium W1

Aims: To isolate and characterize the bacteriocin activity identified by the genomic analysis of a novel enterococcal isolate, Enterococcus faecium W1 Methods and Results: Culture fermentates of a novel E. faecium isolate, W1 showed high levels of antimicrobial activity against Listeria monocytogenesisand a variety of other Gram+ve and Gram-ve bacteria. Ammonium sulphate precipitation of the fermentates followed by hydrophobic interaction chromatography showed bactericidal activity correlated with fractions containing only low molecular weight proteins. The same fractions had activity on human cancer cell line HT-29 measured by cytotoxicity assay (MTT) and apoptosis induction. Whole-genome sequencing of E. faecium W1 revealed five bacteriocin related gene clusters, a lack of virulence factors and lack of antibiotic resistance genes. To confirm that antimicrobial activity was correlated with the bacteriocin genes identified, selected bacteriocin genes were cloned and expressed as His-tagged proteins in E. coli. Bactericidal activity against L. monocytogenes was associated with protein purified by metal affinity chromatography. A structure-function analysis to investigate the role of key residues in activity is ongoing. Conclusion: E. faecium W1 is a promising candidate for use as a probiotic supplement or in food preservation. The bacteriocin related genes are functional and could be improved for future use as antimicrobial and cancer cell therapeutics. Significance and Impact of the Study: There is a need for new approaches to the growing problem of antibiotic resistance. Expression of bacteriocins identified in E. faecium W1 could contribute to this endeavour in addition to its use as a probiotic organism.