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

Abstract

Formulations of ready-to-eat (RTE) foods with antimicrobial compounds constitute an important safety measure against foodborne 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, -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 -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 ␤-glucuronidase (GUS) reporter fusion with the dlt promoter indicated that nisin does not specifically induce VirRdependent upregulation of dltABCD. Lastly, prior exposure of L. monocytogenes parent strain H7858 and the ⌬virR mutant to 2% potassium lactate enhanced subsequent resistance against nisin and -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. Control of Listeria monocytogenes in ready-to-eat (RTE) foods is an important food safety goal due to the high mortality rate associated with listeriosis, particularly in susceptible populations, such as pregnant women, the elderly, and those with a compromised immune system (1). L. monocytogenes is of particular concern for those RTE foods that support growth of this pathogen to high levels during refrigerated storage, which can potentially cause a life-threatening disease. L. monocytogenes harbors a variety of stress coping mechanisms that allow it to survive under suboptimal environmental conditions associated with foods (e.g., acidic, osmotic, and/or temperature stress) (2). The ability of L. monocytogenes to tolerate and grow under such a wide range of adverse conditions elevates the likelihood of foodborne transmission to a human host. Thus, a multipronged approach (e.g., prevention of postprocessing contamination and reformulation of RTE foods with antimicrobials) to limit L. monocytogenes in foods along the farm-to-fork continuum is critical to reduce the potential for foodborne illnesses involving this organism (3).Natural antimicrobials are commonly applied to RTE foods to control foodb...

Genetic and environmental factors influence Listeria monocytogenes nisin resistance

“…Cell envelope stress response systems, including those regulated by VirR (Kang et al . ) and LiaR (Bergholz et al . ), are known to contribute to nisin resistance at low temperatures and in the presence of NaCl.…”

“…Based on the significant differences in nisin resistance phenotypes that we present here, it is possible that genetic background plays a larger role in overall nisin resistance phenotypes than simply the presence or absence of SSI-1. Cell envelope stress response systems, including those regulated by VirR (Kang et al 2015) and LiaR , are known to contribute to nisin resistance at low temperatures and in the presence of NaCl. Variation in regulation or expression of these systems could also contribute to the varying nisin resistance phenotypes observed.…”

Transcriptomic Analysis of the Adaptation of Listeria monocytogenes to Growth on Vacuum-Packed Cold Smoked Salmon

“…H7858 is a lineage I, serotype 4b strain (representing epidemic clone II) isolated from RTE meat and was linked to a multistate listeriosis outbreak from 1998 to 1999 (24,25). We selected H7858 for this study because we had previously quantified the phenotypic and transcriptomic responses of this strain to a number of stresses relevant to food products, including organic acids and bactericidal additives, and know that it can grow to high levels on cold smoked salmon (26)(27)(28). L. monocytogenes H7858 was streaked from frozen BHI stock stored at Ϫ80°C in 15% glycerol onto a BHI agar plate, followed by incubation at 37°C for 24 h. A single colony was subsequently inoculated into 5 ml of BHIB (in 16-mm tubes), followed by incubation at 37°C with shaking (230 rpm) for 18 h (series 25 incubator; New Brunswick Scientific, Edison, NJ).…”

The Role of Stress and Stress Adaptations in Determining the Fate of the Bacterial Pathogen Listeria monocytogenes in the Food Chain

“…Some examples of antimicrobials which have been proven to be active against L. monocytogenes include lauric arginate, chitosan, pediocin, and nisin (Kaur et al, 2013; Kang et al, 2015). Nisin is one of the most common antimicrobials used in the food industry especially within dairy products and acidic foods (Delves-Broughton et al, 1996).…”

“…Different systems including two component regulatory systems and the GSR have been implicated in L. monocytogenes resistance to nisin. Kang et al (2015) showed that a mutant deficient in the response regulator VirR had a greater loss of membrane integrity compared to the wild-type strain, while Begley et al (2006) found that a sigB mutant had decreased growth and survival in response to nisin. However, Palmer et al (2009) reported data which conflicted these results.…”