Garvieacin Q

Lactococcus garvieae BCC 43578 produces a novel class II bacteriocin, garvieacin Q (GarQ), 70 amino acids in length and containing a 20-amino-acid N-terminal leader peptide. It is cleaved at the Gly-Gly site to generate the mature GarQ (5,339 Da), which is especially inhibitory against Listeria monocytogenes ATCC 19115 and other L. garvieae strains.

Bovine mastitis infection in dairy cattle is a significant economic burden for the dairy industry globally. To reduce the use of antibiotics in treatment of clinical mastitis, new alternative treatment options are needed. Antimicrobial peptides from bacteria, also known as bacteriocins, are potential alternatives for combating mastitis pathogens. In search of novel bacteriocins against mastitis pathogens, we screened samples of Norwegian bovine raw milk and found a Streptococcus uberis strain with potent antimicrobial activity toward Enterococcus, Streptococcus, Listeria, and Lactococcus. Whole-genome sequencing of the strain revealed a multibacteriocin gene cluster encoding one class IIb bacteriocin, two class IId bacteriocins, in addition to a three-component regulatory system and a dedicated ABC transporter. Isolation and purification of the antimicrobial activity from culture supernatants resulted in the detection of a 6.3-kDa mass peak by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, a mass corresponding to the predicted size of one of the class IId bacteriocins. The identification of this bacteriocin, called ubericin K, was further confirmed by in vitro protein synthesis, which showed the same inhibitory spectrum as the purified antimicrobial compound. Ubericin K shows highest sequence similarity to the class IId bacteriocins bovicin 255, lactococcin A, and garvieacin Q. We found that ubericin K uses the sugar transporter mannose phosphotransferase (PTS) as a target receptor. Further, by using the pHlourin sensor system to detect intracellular pH changes due to leakage across the membrane, ubericin K was shown to be a pore former, killing target cells by membrane disruption. IMPORTANCE Bacterial infections in dairy cows are a major burden to farmers worldwide because infected cows require expensive treatments and produce less milk. Today, infected cows are treated with antibiotics, a practice that is becoming less effective due to antibiotic resistance. Compounds other than antibiotics also exist that kill bacteria causing infections in cows; these compounds, known as bacteriocins, are natural products produced by other bacteria in the environment. In this work, we discover a new bacteriocin that we call ubericin K, which kills several species of bacteria known to cause infections in dairy cows. We also use in vitro synthesis as a novel method for rapidly characterizing bacteriocins directly from genomic data, which could be useful for other researchers. We believe that ubericin K and the methods described in this work will aid in the transition away from antibiotics in the dairy industry.

The Gram-positive Lacticaseibacillus rhamnosus has been broadly reported as capable of exerting beneficial health effects. Bacterial genomic diversity may promote niche specialization, thus creating subpatterns within populations. As L. rhamnosus advantageous effects have been widely reported at strain level and few is known regarding the distribution of beneficial genes among L. rhamnosus strains, we investigated all publicly available genomes of Lactobacillus and Lacticaseibacillus genera to study the pangenome and general population structure of L. rhamnosus. Core genome multilocus sequence typing detected eight L. rhamnosus phylogroups (PG1 to PG8). L. rhamnosus harbors an open pangenome; PG1, PG3, PG4, and PG5 exhibited highly conserved gene distribution patterns. Genes significantly associated to the PG1, which comprises L. rhamnosus GG, are mainly phage-related. The adhesion operon spaCBA-srtC1 was found in 44 (24.7%) genomes; however, considering only the PG1, the prevalence was of 65%. In PG2 the spaCBA-srtC1 prevalence was of 43%. Nevertheless, both human and milk-derived strains harbored this operon. Further, two main types of bacteriocin clusters were found (Bact1 and Bact2). Bact1 predictions indicate the presence of garQ, encoding the class II bacteriocin garvieacin Q, that is mainly present in the closely related PG8A and a PG2 subcluster. PG2 harbors two distinct subclusters, harboring either spaCBA-srtC1 or Bact1. Our findings provide novel insights on the distribution of biotechnological relevant genes across L. rhamnosus population, uncovering intra-species patterns that may bring forth the development of more efficient probiotic products.