Isolation and characterization from raw milk of Enterocin B-producing Enterococcus faecium: A potential dairy bio-preservative agent

Abstract

• A bacteriocin-producing Enteococcus faecium strain was isolated form raw milk • Cell free supernatant exhibited antimicrobial activity against Listeria monocytogenes • Genomics analysis of RM12 strain revealed several enterocin-encoding genes • Enterocin B was detected through liquid chromatography coupled with mass spectrometry Six Enterococcus faecium strains from raw milk and ripened cheeses showed an effect in impairing L. monocytogenes cell growth. One of the strains, i.e. the RM12, was selected based on its cell-free supernatant (CFS) antilisterial activity due to the presence of a bacteriocin-like substance, exhibiting sensibility to proteolytic enzymes and a partially reduced activity after α-amylase treatment, suggesting its glycoprotein nature. The bacteriocin showed stability at a broad pH range and at high temperatures, and its production was demonstrated in both commercial and dairy based media containing lactose (i.e. pasteurized milk, whey and scotta). The antimicrobial activity peak correlated with the strain growth kinetics, showing a bactericidal mode of action versus Listeria monocytogenes . At genomic level, three genes encoding for enterocins were found (namely entA, entB and entP ), whereas an approach based on CFS peptides fractionation, followed by liquid chromatography coupled with mass spectrometry, lead to the identification of only one constitutive protein, the Enterocin B, responsible for the antimicrobial activity. E. faecium RM12 was susceptible to the whole tested antibiotic panel, except for gentamicin, and did not show any aminoacidic decarboxylating nor haemolytic activities. The analyses here conducted thus provide a comprehensive characterization of Enterocin B-producing E faecium RM12 and his potential usage as bio-preservative agent in fermented dairy products.

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