PROJECT SUMMARY Of the foodborne bacterial, protozoal and viral diseases, non-typhoidal Salmonella enterica cause the largest burden of illness and death worldwide. The most common human clinical isolates are Salmonella enterica serovars Typhimurium (S. Typhimurium, STm) and Enteriditis (S. Enteriditis). Infection can cause either a self- limiting gastroenteritis or a life-threatening, invasive disease in immunocompromised individuals. During enteric infection, STm adopts both extracellular and intracellular lifestyles, colonizing the intestinal lumen as well epithelial cells and phagocytes in the intestinal mucosa. Here we will study how host-mediated restriction of transition metals, known as nutritional immunity, affects STm colonization of the gut. The NRAMP family of proteins are “promiscuous” transporters of divalent cations that play a major role in metal ion homeostasis from bacteria to man. In mammals, there are two NRAMP genes, SLC11A1 and SLC11A2 in humans and Slc11a1 and Slc11a2 in mice. Slc11a1 is restricted to the myeloid lineage whereas Slc11a2 is ubiquitously expressed. Whilst the role of Slc11a1 in controlling intracellular bacterial and parasitic infections in macrophages and mice is undisputed, whether Slc11a2 also contributes to antimicrobial functions is unknown. Our general hypothesis is that intracellular metal ion availability differs in epithelial cells and phagocytes in the gut due to the distinct cell-type expression of SLC11A1 and SLC11A2. First, we will use microbial biosensors of metal ion concentrations to infect bovine ligated ileal loops, a highly relevant model of enteric salmonellosis in humans, and map the spatiotemporal distribution of metal ion deprivation in the gut. Second, we will define whether SLC11A2, the sole NRAMP family member expressed in intestinal epithelial cells, defends against invading pathogens via metal nutrient limitation. Completion of this proposal will close a significant knowledge gap about host-microbe competition for metal ions in the gut during enteric infection.