The Influence of the Toxin/Antitoxin mazEF on Growth and Survival of Listeria monocytogenes under Stress

A major factor in the resilience of Listeria monocytogenes is the alternative sigma factor B (σB). Type II Toxin/Antitoxin (TA) systems are also known to have a role in the bacterial stress response upon activation via the ClpP or Lon proteases. Directly upstream of the σB operon in L. monocytogenes is the TA system mazEF, which can cleave mRNA at UACMU sites. In this study, we showed that the mazEF TA locus does not affect the level of persister formation during treatment with antibiotics in lethal doses, but exerts different effects according to the sub-inhibitory stress added. Growth of a ΔmazEF mutant was enhanced relative to the wildtype in the presence of sub-inhibitory norfloxacin and at 42 °C, but was decreased when challenged with ampicillin and gentamicin. In contrast to studies in Staphylococcus aureus, we found that the mazEF locus did not affect transcription of genes within the σB operon, but MazEF effected the expression of the σB-dependent genes opuCA and lmo0880, with a 0.22 and 0.05 fold change, respectively, compared to the wildtype under sub-inhibitory norfloxacin conditions. How exactly this system operates remains an open question, however, our data indicates it is not analogous to the system of S. aureus, suggesting a novel mode of action for MazEF in L. monocytogenes

Sublethal Concentrations of Antibiotics Cause Shift to Anaerobic Metabolism in Listeria monocytogenes and Induce Phenotypes Linked to Antibiotic Tolerance

The human pathogenic bacterium Listeria monocytogenes is exposed to antibiotics both during clinical treatment and in its saprophytic life tyle. As one of the keys to successful treatment is continued antibiotic sensitivity, the purpose of this study was to determine if exposure to sublethal antibiotic concentrations would affect the bacterial physiology and induce antibiotic tolerance. Transcriptomic analyses demonstrated that each of the four antibiotics tested caused an antibiotic-specific gene expression pattern related to mode-of-action of the particular antibiotic. All four antibiotics caused the same changes in expression of several metabolic genes indicating a shift from aerobic to anaerobic metabolism and higher ethanol production. A mutant in the bifunctional acetaldehyde-CoA/alcohol dehydrogenase encoded by lmo1634 did not have altered antibiotic tolerance. However, a mutant in lmo1179 (eutE) encoding an aldehyde oxidoreductase where rerouting caused increased ethanol production was tolerant to three of four antibiotics tested. This shift in metabolism could be a survival strategy in response to antibiotics to avoid generation of ROS production from respiration by oxidation of NADH through ethanol production. The monocin locus encoding a cryptic prophage was induced by co-trimoxazole and repressed by ampicillin and gentamicin, and this correlated with an observed antibiotic-dependent biofilm formation. A monocin mutant (ΔlmaDCBA) had increased biofilm formation when exposed to increasing concentration of co-trimoxazole similar to the wild type, but was more tolerant to killing by co-trimoxazole and ampicillin. Thus, sublethal concentrations of antibiotics caused metabolic and physiological changes indicating that the organism is preparing to withstand lethal antibiotic concentrations