Investigating Listeria monocytogenes colonization of the gallbladder

Abstract

Listeriosis, caused by the bacterium Listeria monocytogenes, is the third leading cause of death among foodborne illnesses in the United States. Following oral inoculation of the host, L. monocytogenes transverses the intestinal epithelium and rapidly disseminates to the mesenteric lymph nodes, liver, and spleen via the lymph and bloodstream. Bacteria colonizing the liver replicate intracellularly in hepatocytes, and as infected cells lyse some bacteria migrate to the gallbladder via hepatic ducts. Once in the gallbladder, L. monocytogenes replicates extracellularly to high bacterial densities. The gallbladder then becomes the primary bacterial reservoir and the source of fecally excreted bacteria. Despite its importance in L. monocytogenes pathogenesis, little is known about how L. monocytogenes survives and replicates in the gallbladder. The goal of this dissertation is to characterize the gallbladder as an infectious niche for L. monocytogenes, which will lead to a more complete understanding of its multi-organ infection cycle and the determinants of intra- vs. extracellular infection. In this work, I investigated both bacterial and host factors that contribute to gallbladder colonization. First, I developed a novel genetic screen using an ex vivo primate gallbladder infection model to reveal L. monocytogenes genes that are required for replication. Further characterization candidates from the ex vivo screen showed that many are also required for intracellular infection and virulence in an oral murine model of listeriosis. Of note, this study identified sugar transport by phosphoenolpyruvate-dependent phosphotransferase systems (PTS) as being important for replication in the gallbladder. These findings represent novel insights into L. monocytogenes carbon metabolism during infection and suggests a role for these transporters for infection in extracellular niches. Next, using a L. monocytogenes mutant deficient in gallbladder colonization, I found that STING-mediated innate immunity plays a role in gallbladder colonization following oral infection. Finally, I investigated the roles peroxide resistance mechanisms and iron transport systems during intracellular replication, which revealed extensive redundancy in the stress responses employed by L. monocytogenes during infection. Overall, my dissertation research highlights the gallbladder as a replicative niche for L. monocytogenes and genetic determinants required for L. monocytogenes replication, which will lend further insight into the gallbladder’s role in its multi-organ pathogenic life cycle.