Infections caused by highly antibiotic-resistant microbes are an increasing problem in hospitalized patients. Many of these infections occur following antibiotic therapy, an intervention that promotes colonization with highly antibiotic-resistant bacteria. Although elimination of commensal bacteria opens up physical and nutrient niches that enhance growth of antibiotic resistant microbes, results from our laboratory suggest that antibiotics also compromise innate immune defenses on mucosal surfaces by interfering with homeostatic stimulation of innate immune receptors by commensal flora. The focus of this application is to characterize the relationship between intestinal microbes and the mucosal innate immune system. Our experimental approach will be to manipulate the commensal flora or the innate immune system and measure the host's ability to resist colonization and infection by Vancomycin-resistant enterococcus (VRE) and carbapenem-resistant Klebsiella pneumoniae (KPC), two highly antibiotic resistant bacteria that are emerging causes of severe, frequently lethal infections. Our first aim is to characterize the effect of distinct antibiotics on the commensal flora of the murine gut and the downstream effect of antibiotics on the expression of innate immune effector molecules by the intestinal epithelium. The second aim of our studies is to determine the effect of innate immune receptor deficiencies on the flora of the murine intestinal tract. These studies will take advantage of a large panel of TLR and signaling adaptor mutant mouse strains in our animal colony. Our third aim is to test the ability of different microbial molecules or analogs to stimulate innate immune defense against VRE and KPC in mice depleted of commensal flora by antibiotic treatment. We believe these studies will provide important insights into the two-way relationship between the microbial flora and the intestinal epithelium. Our studies are likely to provide new approaches to prevent or limit infections caused by highly antibiotic-resistant bacteria. PUBLIC HEALTH RELEVANCE: Infections with highly antibiotic-resistant bacteria are an increasingly common problem in patients being treated with antibiotics. We have discovered that antibiotics, by eliminating normal bacteria inhabiting the intestine, compromise the ability of the intestinal lining to resist attack by antibiotic-resistant bacteria. Our experiments will determine which normal intestinal bacteria are important for the maintenance of antimicrobial resistance in the intestine, and which mammalian molecules are responsible for detecting normal bacteria in the intestine. Our third aim is to discover whether administration of molecules derived from bacteria can reverse the increased susceptibility to infection induced by antibiotic administration.