VirR-Mediated Resistance of Listeria monocytogenes against Food Antimicrobials and Cross-Protection Induced by Exposure to Organic Acid Salts

Formulations of ready-to-eat (RTE) foods with antimicrobial compounds constitute an important safety measure against food-borne pathogens such as Listeria monocytogenes. While the efficacy of many commercially available antimicrobial compounds has been demonstrated in a variety of foods, the current understanding of the resistance mechanisms employed by L. monocytogenes to counteract these stresses is limited. In this study, we screened in-frame deletion mutants of two-component system response regulators associated with the cell envelope stress response for increased sensitivity to commercially available antimicrobial compounds (nisin, lauric arginate, epsilon-polylysine, and chitosan). A virR deletion mutant showed increased sensitivity to all antimicrobials and significantly greater loss of membrane integrity when exposed to nisin, lauric arginate, or epsilon-polylysine (P < 0.05). The VirR-regulated operon, dltABCD, was shown to be the key contributor to resistance against these antimicrobial compounds, whereas another VirR-regulated gene, mprF, displayed an antimicrobial-specific contribution to resistance. An experiment with a beta-glucuronidase (GUS) reporter fusion with the dlt promoter indicated that nisin does not specifically induce Vir-dependent upregulation of dltABCD. Lastly, prior exposure of L. monocytogenes parent strain H7858 and the Delta virR mutant to 2% potassium lactate enhanced subsequent resistance against nisin and epsilon-polylysine (P < 0.05). These data demonstrate that VirRS-mediated regulation of dltABCD is the major resistance mechanism used by L. monocytogenes against cell envelope-damaging food antimicrobials. Further, the potential for cross-protection induced by other food-related stresses (e.g., organic acids) needs to be considered when applying these novel food antimicrobials as a hurdle strategy for RTE foods.