Resistance to the Bacteriocin Lcn972 Deciphered by Genome Sequencing.

In view of the current threat of antibiotic resistance, new antimicrobials with low risk of resistance development are demanded. Lcn972 is a lactococcal bacteriocin that inhibits septum formation by binding to the cell wall precursor lipid II in . It has a species-specific spectrum of activity, making Lcn972 an attractive template to develop or improve existing antibiotics. The aim of this work was to identify mutations present in the Lcn972-resistant clone D1-20, previously evolved from the sensitive strain MG1614. Whole-genome sequencing and comparison over the reference genome MG1363 identified several unexpected mutations in the parental strain MG1614, likely selected during in-house propagation. In the Lcn972R clone, two previously identified mutations were mapped and confirmed. Additionally, another transposition event deregulating cellobiose uptake was identified along with three point mutations of unknown consequences for Lcn972 resistance. Two new independent evolution experiments exposing MG1614 to Lcn972 revealed transposition of 981 into the locus as the predominant mutation selected by Lcn972. This event occurs early during evolution and was found in 100% of the evolved clones, while other mutations were not selected. Therefore, activation of coding for a putative anti-ECF (extra-cytoplasmic function) sigma factor is regarded as the main Lcn972 resistance factor in MG1614.