Natural resistance-associated macrophage protein (NRAMP) encoded by the Slc11a1 gene is a membrane-associated transporter of divalent metal ions. Murine Slc11a1 has two known alleles, a functional Slc11a1(Gly169), which is found in DBA2/J, NOD/LtJ, and 129p3/J and related mouse strains, and a non-functional Slc11a1(Asp169), that is found in C56Bl/6J (B6) and BALB/cJ mice. B6 mice congenic for Slc11a1(Gly169) (B6-Slc11a1(G169)) are markedly resistant to the intracellular pathogens Salmonella, Leishmania, and Mycobacterium tuberculosis. We examined the host cell response and replication of Francisella in B6-Slc11a1(G169) mice. Bone marrow-derived macrophages from either B6-Slc11a1(G169) or B6 mice were both effectively invaded by Francisella live vaccine strain (LVS). However, at 16 hours post-infection (hpi), the number of LVS bacteria recovered from B6 macrophages had increased roughly 100-fold, while in B6-Slc11a1(G169) mice the number decreased 10-fold. When the mice were challenged intranasally (i.n.) B6 mice lost significant amounts (similar to 15%) of weight, where as B6-Slc11a1(G169) mice lost no weight. Three days after infection in B6-Slc11a1(G169) mice, we failed to recover viable Francisella from the lungs, livers, or spleens. By contrast, B6 mice had bacterial burdens approaching 1x10(6) CFU/organ in all three organs. To further examine the degree of resistance imparted by Slc11a1(Gly169) expression, we challenged mice deficient in TLR2, TLR4, and TLR9, but expressing the functional Slc11a1 (B6-Slc11a1(G169) Tlr2/4/9(-/-)). Surprisingly, B6-Slc11a1(G169) Tlr2/4/9(-/-)mice had no notable weight loss. Eighty percent of B6-Slc11a1(G169) Tlr2/4/9(-/-)mice yielded no detectable Francisella in any organ tested. Additionally, Slc11a1(G169) produced little detectable cytokine either in the lung or serum compared to B6 mice. Mice expressing Slc11a1(Gly169) survived even high doses (similar to 80 LD50) of LVS inoculation. These data taken together serve to highlight that functional Slc11a1(Gly169) can compensate the lack of TLR2/4/9. Thus Slc11a1 is a critical player in murine resistance to pulmonary Francisella infection, but not footpad infection.