Bacteriocin-induced mutations activate antibiotic resistance modules in Lactococcus lactis

Abstract

[Background] Adaptive evolution experiments under cell envelope stress exerted by the lipid IIbinding bacteriocin Lcn972 selected for evolved Lcn972R L. lactis mutants resistant to bacitracin and nisin. Genome sequencing identified mutations in the ysaDCB operon coding for a putative bacitracin ABC-transporter (BceAB-like) involved in sensing and resistance to antimicrobial peptides. [Objectives] Our goal was to assess the role of the mutations found in the YsaB permease. [Methods] The genes and their mutated versions were cloned and expressed on a defective ysaB L. lactis. Their contribution to resistance was determined by IC50 assays and their ability to sense bacitracin and activate transcription was quantified by promoter fusions. [Results]: Expression of the wild type genes resulted in a 4-fold increase in resistance to bacitracin, demonstrating their function as a bona-fide BceAB-like ABC transporter in L. lactis. Mutated versions, however, were more effective increasing up to 9- and 5-fold bacitracin IC50 values. Moreover, counter mutations could be selected that restored the antibiotic sensitive phenotype. Promoter fusions revealed that mutated versions had an increased signalling activity and, therefore, a higher basal expression of the ysaDCB operon in the absence of bacitracin, i.e. without inducer. To sum up, bacteriocin pressure may select for constitutively expressed and functionally enhanced efflux pumps that interfere with antibiotic activity.