Enterocin AS-48RJ: a variant of enterocin AS-48 chromosomally encoded by Enterococcus faecium RJ16 isolated from food

The bacteriocinogenic strain RJ16 isolated from goat cheese has been identified as Enterococcus faecium by species-specific PCR, DNA-rRNA hybridization and rDNA sequencing. Purified bacteriocin from strain RJ16 is a carboxypeptidase A-resistant peptide with a molecular mass (7125 Da) very close to the cyclic peptide enterocin AS-48. Bacteriocin from strain RJ16 and AS-48 show identical antibacterial spectra, although the former is slightly less active on strains of Listeria monocytogenes and Bacillus cereus. Producer strains show cross-immunity. PCR amplification of total DNA from strain RJ16 with primers for the AS-48 structural gene and sequencing of the amplified fragment revealed an almost identical sequence (99.5%), except for a single mutation that predicts the change of Glu residue at position 20 of AS-48 to Val. Therefore, bacteriocin produced by E. faecium RJ16 should be considered a variant of AS-48, which we call AS-48RJ. PCR amplification revealed that strain RJ16 contains the complete as-48 gene cluster. Hybridization with probes for as-48 gene cluster revealed a chromosomal location of as-48 genes in strain RJ16, being the first example of a chromosomal location of this bacteriocin trait. Strain RJ16 produced enzymes of interest in food processing (esterase, esterase lipase and phytase activities), and did not decarboxylate amino acids precursors for biogenic amines. Strain RJ16 did not exhibit haemolytic or gelatinase activities, and PCR amplification revealed the lack of genes encoding for known virulence determinants (aggregation substance, collagen adhesin, enterococcal surface protein, endocarditis antigens, as well as haemolysin and gelatinase production). Strain RJ16 was resistant to ciprofloxacin (MIC>2mgl-1) and levofloxacin (MIC>4mgl-1) and showed intermediate resistance to nitrofurantoin and erythromycin, but was sensitive to ampicillin, penicillin, streptomycin, gentamicin, rifampicin, chloramphenicol, tetracycline, quinupristin/dalfopristin, vancomycin and teicoplanin. Altogether, results from this study suggest that this broad-spectrum bacteriocin-producing strain may have a potential use in food preservation.