Acid tolerance response of Listeria monocytogenes: Bioenergetics and mechanisms of resistance to the antimicrobial nisin

Bacterial infections caused by Listeria monocytogenes target mainly newborn, pregnant, elderly and immunocompromised individuals, and are associated with mortality rates around 25%. Control of L. monocytogenes in foods is difficult due to its widespread presence in nature and ability to grow at refrigeration temperatures. The responses of L. monocytogenes to acid, osmotic and thermal stresses contribute to the problem, since they increase pathogen resistance and virulence beyond constitutive levels. The Acid Tolerance Response (ATR) is defined as the abnormal resistance to lethal acid after exposure to mild acidic conditions. The specific acid, rather than the medium's pH, is critical for ATR induction.; Changes in regulation of stress response proteins occur during induction of the ATR. These proteins include chaperones, transcriptional regulators, and, as demonstrated herein, the F0F1-ATPase, a proton pump. The ATR also increases virulence and cross-protects listeriae to other stressors, such as elevated temperatures and antimicrobials. This dissertation reports that the ATR conferred persistent resistance to the food antimicrobial nisin. ATR-induced cells (ATR+), but not the control (ATR -), persist for at least 30 days at 4°C in a model fermented food where Lactococcus lactis had produced lactic acid and nisin.; To elucidadate the mechanism of resistance, this study determined that ATR+ cells had both lower transmembrane electric potential (Deltapsi) and transmembrane pH (DeltapH) relative to ATR- . This resulting low proton motive force (PMF) of ATR+ cells was implicated in their higher resistance to nisin, whose action is enhanced by energized membranes, and was independent of de novo protein synthesis. The reduced PMF of ATR+ cells was paradoxically accompanied by significantly increased intracellular ATP concentrations. Since the listerial F0F1-ATPase enzyme complex hydrolyses ATP to generate PMF, we studied the regulation of this enzyme. Mass spectrometric analyses suggest that ATR+ cells downregulate the F0F 1-ATPase c subunit compared to ATR- cells. This reduction in the number of proton-translocating c subunits results from the reduced PMF of ATR+ cells, which decreased the F0F 1-ATPase activity, spared vital ATP and rendered the cells less sensitive to nisin.; This link between bioenergetics and proteomics, leading to resistance to multiple stressors, provides a novel target for multiple hurdle inhibitory systems against L. monocytogenes and possibly other pathogens.