Bacilysin within the Bacillus subtilis group: gene prevalence versus antagonistic activity against Gram-negative foodborne pathogens

Abstract

The Bacillus subtilis group comprises species known for their ability to produce a wide variety of antimicrobial peptides. This work focuses on bacilysin, a broad-spectrum active dipeptide, and its prevalence in the B. subtilis group. In silico genome analysis of strains from Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus licheniformis, Bacillus pumilus and B. subtilis subspecies inaquosorum, spizizenii and subtilis revealed that the bacilysin gene cluster is present in all species except for B. licheniformis. This observation was corroborated by PCR detection of the bacilysin genetic determinants on a collection of 168 food and environmental strains from the B. subtilis group. Phylogenetic analyses also demonstrated that the bacilysin gene cluster sequence showed more than 80 % identity within each species of the B. subtilis group. An in vitro screening of the strain collection was performed against foodborne pathogens. Twenty-three strains were selected for their ability of their Cell-Free Supernatant to inhibit foodborne pathogens. After an ammonium sulphate precipitation of their supernatant, eight strains, all belonging to B. velezensis, exhibited antimicrobial activity against Gram-negative pathogens. Using Ultra High Performance Liquid Chromatography - Mass Spectrometry, the presence of bacilysin was confirmed in these eight precipitates. These findings provide evidence that bacilysin is a major player in the antagonistic activity of B. velezensis against Gram-negative foodborne pathogens.