Genetic control of resistance to experimental infection with virulent Mycobacterium tuberculosis

Abstract

Over 2 billion people are estimated to be infected with virulent Mycobacterium tuberculosis, yet fewer than 10% progress to clinical tuberculosis within their lifetime. Twin studies and variations in the outcome of tuberculosis infection after exposure to similar environmental risks suggest genetic heterogeneity among individuals in their susceptibility to disease. In a mouse model of tuberculosis, we have established that resistance and susceptibility to virulent M. tuberculosis is a complex genetic trait. A new locus with a major effect on tuberculosis susceptibility, designated sst1 (susceptibility to tuberculosis 1), was mapped to a 9-centimorgan (cM) interval on mouse chromosome 1. It is located 10-19 cM distal to a previously identified gene, Nramp1, that controls the innate resistance of mice to the attenuated bacillus Calmette-Guérin vaccine strain. The phenotypic expression of the newly identified locus is distinct from that of Nramp1 in that sst1 controls progression of tuberculosis infection in a lung-specific manner. Mice segregating at the sst1 locus exhibit marked differences in the growth rates of virulent tubercle bacilli in the lungs. Lung lesions in congenic sst1-susceptible mice are characterized by extensive necrosis and unrestricted extracellular multiplication of virulent mycobacteria, whereas sst1-resistant mice develop interstitial granulomas and effectively control multiplication of the bacilli. The resistant allele of sst1, although powerful in controlling infection, is not sufficient to confer full protection against virulent M. tuberculosis, indicating that other genes located outside of the sst1 locus are likely also to be important for controlling tuberculosis infection.

Figure 1

Survival of C3HeB/FeJ and C57BL/6J mice and their F₁ and F₂ hybrids after i.v. infection with 10⁶ cfu of M. tuberculosis Erdman strain.

Figure 2

Segregation of sst1 alleles in the progeny of recombinant males. Males that carried recombinations within the sst1-containing segment of chromosome 1 were identified by genotyping with microsatellite markers using DNA obtained from tail biopsies. Recombinant males were crossed with C3H females, and their progeny was genotyped and tested for susceptibility to i.v. infection with M. tuberculosis. Each column of boxes represents a genotypic class. The number of recombinant males per each genotypic class is denoted by the digit under each column. Each box represents the genotype for loci corresponding to chromosome 1 map on the left. Solid boxes represent homozygotes for B6 alleles, striped boxes are heterozygotes, and open boxes are homozygotes for the C3H allele. The survival times of the progeny heterozygous or homozygous for each marker was compared by using Student's t test. Significant differences between those groups (P < 0.0001) indicated heterozygosity of the recombinant male for the sst1-resistant allele. The sst1 genotypes inferred by this method are at the top of the figure. The two horizontal lines drawn between D1Mit334 and D1Mit45 designate the location of the sst1-containing segment.

Figure 3

Multiplication of M. tuberculosis Erdman and BCG in spleens (Upper) and lungs (Lower) of the backcross 3 (BC3H) mice segregating at the sst1 locus. The animals were infected with equal doses (2 × 10⁴ cfu) of either virulent or vaccine strain of mycobacteria, and five to seven mice were killed per group at each time point.

Figure 4

Histopathology of lungs from M. tuberculosis-infected backcross 3 (BC3H) mice 6 weeks after infection with 2 × 10⁴ cfu of M. tuberculosis (hematoxylin/eosin staining). (A) sst1^s/s mice. (B) sst1^r/s mice.