Genetic and functional dissection of resistance to gram-negative infection in wild-derived mice

Infectious diseases continue to be one of the leading causes of morbidity and mortality in the world. Infection of mice with Salmonella typhimurium causes a systemic disease reminiscent of human typhoid fever and constitutes one of the best-described models of infectious disease.We show that wild-derived Mus musculus molossinus (MOLF/Ei) mice are extremely susceptible to infection with S. typhimurium, despite resistance alleles at known Salmonella-susceptibility loci. To identify the genes responsible for the innate susceptibility of MOLF/Ei mice, we performed a genome-wide scan using a panel of (C57BL/6J x MOLF/Ei)F2 mice. Salmonella-resistance was linked to Nramp1 with a LOD score of 18.8 and to mouse chromosome 11 with peak LOD score of 7.0 at D11Mit5. One recessive, susceptibility QTL (quantitative trait loci) was identified on distal chromosome 1 with a maximal LOD score of 4.8 at D1Mit100.In parallel, we undertook a functional dissection of the response of MOLF/Ei mice to infection with S. typhimurium. Kinetics of bacterial proliferation within the reticuloendothelial organs and histological evaluation of tissue sections suggest that MOLF/Ei mice do not succumb to infection following an overwhelming bacterial growth or massive tissue necrosis. We also demonstrate that, in spite of a normal response to Lipopolysaccharide and high expression of IL-1alpha and IL-1beta mRNA during infection, MOLF/Ei do not upregulate Nos2 mRNA expression and have a reduced capacity to release nitrite in vitro.We examined the candidacy of several genes located within the chromosomal interval defined by our genetic analysis and thought to be involved in resistance to Salmonella infection. The newly described role of Toll-like receptors in innate immunity and the location of human TLR5 in a region homologous to the interval containing the mouse chromosome 1 Salmonella-susceptibility QTL prompted us to clone mouse Tlr5. Mouse Tlr5 mRNA encodes for an 859 amino acid protein predominantly expressed in the liver. As expected, Tlr5 maps to the distal chromosome 1. The decreased expression of Tlr5 mRNA in the liver of susceptible MOLF/Ei mice, as compared to other strains, suggests the possibility that Tlr5 may play a role in host response to Gram-negative infection in wild-derived mice.We examined four candidate genes (Il12b, Csfgm, Nos2 and Mpo) within the Salmonella-resistance QTL on mouse chromosome 11. Although sequence analysis and mRNA expression studies reveal differences between the MOLF/Ei and C57BL/6J alleles, no definite conclusions can be drawn as to the roles of these genes in S. typhimurium infection. We created a series of B6.MOLF-Nramp1Gly169/chr11 congenic strains that collectively overlap the entire chromosome 11 QTL to define more precisely the chromosomal interval involved and, ultimately, to identify the causative gene(s). Preliminary testing of these strains confirms the presence of the MOLF/Ei-resistance locus and allows us to exclude the genomic area encompassing D11Mit84 and D11Mit141 from the QTL interval, thus eliminating Il12b as a candidate gene. Further characterisation of these strains will help elucidate which of the candidate genes on chromosome 11 confers resistance to infection with S. typhimurium