The devR-devS two-component system of Mycobacterium tuberculosis was identified earlier and partially characterized in our laboratory. A devR::kan mutant of M. tuberculosis was constructed by allelic exchange. The devR mutant strain showed reduced cell-to-cell adherence in comparison to the parental strain in laboratory culture media. This phenotype was reversed on complementation with a wild-type copy of devR. The devR mutant and parental strains grew at equivalent rates within human monocytes either in the absence or in the presence of lymphocytic cells. The expression of DevR was not modulated upon entry of M. tuberculosis into human monocytes. However, guinea pigs infected with the mutant strain showed a significant decrease in gross lesions in lung, liver and spleen; only mild pathological changes in liver and lung; and a nearly 3 log lower bacterial burden in spleen compared to guinea pigs infected with the parental strain. Our results suggest that DevR is required for virulence in guinea pigs but is not essential for entry, survival and multiplication of M. tuberculosis within human monocytes in vitro. The attenuation in virulence of the devR mutant in guinea pigs together with DevR-DevS being a bona fide signal transduction system indicates that DevR plays a critical and regulatory role in the adaptation and survival of M. tuberculosis within tissues.