An attenuated quadruple gene mutant of <i>Mycobacterium tuberculosis</i> imparts protection against tuberculosis in guinea pigs

Previously we had developed a triple gene mutant of Mycobacterium tuberculosis ( Mtb Δ mms ) harboring disruption in three genes, namely mptpA , mptpB and sapM Though vaccination with Mtb Δ mms strain induced protection in the lungs of guinea pigs, the mutant strain failed to control the hematogenous spread of the challenge strain to the spleen. Additionally, inoculation with Mtb Δ mms resulted in some pathological damage to the spleens in the early phase of infection. In order to generate a strain that overcomes the pathology caused by MtbΔmms in spleen of guinea pigs and controls dissemination of the challenge strain, MtbΔmms was genetically modified by disrupting bioA gene to generate MtbΔmmsb strain. Further, in vivo attenuation of MtbΔmmsb was evaluated and its protective efficacy was assessed against virulent M. tuberculosis challenge in guinea pigs. Mtb Δ mmsb mutant strain was highly attenuated for growth and virulence in guinea pigs. Vaccination with Mtb Δ mmsb mutant generated significant protection in comparison to sham-immunized animals at 4 and 12 weeks post-infection in lungs and spleen of infected animals. However, the protection imparted by Mtb Δ mmsb was significantly less in comparison to BCG immunized animals. This study indicates the importance of attenuated multiple gene deletion mutants of M. tuberculosis for generating protection against tuberculosis.