Streptococcus uberis is one of the principal causative agents of bovine mastitis. In this study, we report that S. uberis strain 42 produces a lantibiotic, nisin U, which is 78% identical (82% similar) to nisin A from Lactococcus lactis. The 15.6-kb nisin U locus comprises 11 open reading frames, similar in putative functionality but differing in arrangement from that of the nisin A biosynthetic cluster. The nisin U producer strain exhibits specific resistance (immunity) to nisin U and cross-resistance to nisin A, a finding consistent with the 55% sequence similarity of their respective immunity peptides. Homologues of the nisin U structural gene were identified in several additional S. uberis strains, and in each case cross-protective immunity was expressed to nisin A and to the other producers of nisin U and its variants. To our knowledge, this is the first report both of characterization of a bacteriocin by S. uberis, as well as of a member of the nisin family of peptides in a species other than L. lactis.The lactic acid bacterium Streptococcus uberis, in nature primarily found on the lips and skin of cows, in raw milk, and on udder tissue (11), is also a major cause of bovine mastitis (4). Due to the ubiquity of this bacterium in the environment of the dairy cow, teat-end contamination poses a constant threat of infection, and the current level of disease caused by S. uberis remains a persistent problem impacting both on the economic production of milk and on the welfare of the dairy cow (21).Many lactic acid bacteria produce broad-spectrum proteinaceous antimicrobials called bacteriocins, some of which could provide valuable alternatives to traditional therapeutic antibiotics for the treatment of infectious diseases (30). Two such bacteriocins, nisin and lacticin 3147, which are produced by strains of Lactococcus lactis are potential candidates for mastitis control (5, 30). Nisin is the active ingredient in two commercial products: Consept (Applied Microbiology, Inc., New York, NY) and Wipe-Out (ImmuCell, Portland, OR). Lacticin 3147 has also been evaluated as a teat-seal formulation for the prevention of mastitis during the "dry" period in which the cow is not lactating (30).Nisin and lacticin 3147 both belong to the lantibiotic class of bacteriocins (10, 34). The lantibiotics are ribosomally synthesized, low-molecular-weight, heat-stable peptides characterized by their content of posttranslationally modified amino acids, including lanthionine and/or ␤-methyl-lanthionine (22,26,31). Nisin is the most intensively studied lantibiotic (20,35,37). Nisin Z (25) and nisin Q (45) are two natural variants of the original nisin A, differing in their propeptide components from nisin A by one and four amino acids, respectively. Lantibiotic loci typically comprise a structural gene (lanA) and other genes that encode proteins responsible for posttranslational modification of the prepeptide (lanB and lanC, or lanM), proteolytic processing (lanP), transport (lanT), producer selfprotection (lanI and lanEFG), and regulatio...

This is the first report of intra-species horizontal gene transfer for subtilosin and the first fully characterized bacteriocin isolated from B. amyloliquefaciens. Finally, this is the first report on subtilosin's activity against bacterial vaginosis-associated pathogens.

Salivaricin A (SalA), the first Streptococcus salivarius lantibiotic to be characterized, appears to be inhibitory to most Streptococcus pyogenes strains. A variant of the SalA structural gene (salA1) is present in more than 90% of S. pyogenes strains, but only strains of M serotype 4 and T pattern 4 produce the biologically active peptide. The present study identifies four additional variants (salA2 to salA5) of the SalA structural gene and demonstrates that each of the corresponding inhibitory peptides (SalA2 to SalA5) is produced in vitro. These variants appear to be similar to SalA and SalA1 in their inhibitory activity against Micrococcus luteus and in their ability to act as inducers of SalA production. It had previously been shown that S. pyogenes strain SF370 had a deletion (of approximately 2.5 kb) in the salM and salT genes of the salA1 locus. In the present study, several additional characteristic deletions within the salA1 loci were identified. S. pyogenes strains of the same M serotype all share the same salA1 locus structure. Since S. salivarius is a predominant member of the normal oral flora of healthy humans, strains producing anti-S. pyogenes lantibiotics, such as SalA, may have excellent potential for use as oral probiotics. In the present study, we have used a highly specific SalA induction system to directly detect the presence of SalA in the saliva of humans who either naturally harbor populations of SalA-producing S. salivarius or who have been colonized with the SalA2-producing probiotic S. salivarius K12.

Streptococcus uberis is commonly found in the environment and in association with various bovine body sites and is a major cause of bovine mastitis. Moreover, S. uberis is known to produce a variety of bacteriocin-like inhibitory substances, antimicrobial agents that generally inhibit closely related bacterial species. In this respect, S. uberis strain 42 has previously been shown to produce a novel nisin variant named nisin U. This paper reports that, in addition to nisin U, S. uberis strain 42 produces a second bacteriocin that induces the lysis of metabolically active, susceptible target bacteria and which has therefore been named uberolysin. Isolation of the native active antimicrobial agent revealed that uberolysin is a 7048 Da peptide that is refractory to sequence analysis by Edman degradation. Transposon mutagenesis was used to generate a uberolysin-negative mutant of S. uberis 42 and sequencing of DNA flanking the insertion site revealed, in addition to the structural gene (ublA), several open reading frames likely to be involved in post-translational modification, transport and producer self-protection (immunity), and possibly in regulation of the biosynthetic gene cluster. In addition, a pair of direct repeats that may be involved in bacteriocin acquisition were identified; indeed, ublA could be identified in 18 % of tested S. uberis strains. Enzymic hydrolysis of uberolysin was used to confirm that ublA does indeed encode the precursor of uberolysin, that an unusually short leader sequence of only six amino acids is cleaved during processing of the mature peptide and that uberolysin is post-translationally covalently modified to form a head-to-tail monocycle. Thus, uberolysin is a unique cyclic bacteriocin, belonging to the same family of bacteriocins as enterocin AS-48 and circularin A.

Bacterial vaginosis is a common vaginal infection associated with numerous gynecological and obstetric complications. This condition is characterized by the presence of thick adherent vaginal biofilms, composed mainly of Gardnerella vaginalis. This organism is thought to be the primary aetiological cause of the infection paving the way for various opportunists to colonize the niche. Previously, we reported that the natural antimicrobials subtilosin, ε-poly-L-lysine, and lauramide arginine ethyl ester selectively inhibit the growth of this pathogen. In this study, we used plate counts to evaluate the efficacy of these antimicrobials against established biofilms of G. vaginalis. Additionally, we validated and compared two rapid methods (ATP viability and resazurin assays) for the assessment of cell viability in the antimicrobial-treated G. vaginalis biofilms. Out of the tested antimicrobials, lauramide arginine ethyl ester had the strongest bactericidal effect, followed by subtilosin, with clindamycin and polylysine showing the weakest effect. In comparison to plate counts, ATP viability and resazurin assays considerably underestimated the bactericidal effect of some antimicrobials. Our results indicate that these assays should be validated for every new application.