Deletion of BCG_2432c from the Bacillus Calmette-Guerin vaccine enhances autophagy-mediated immunity against tuberculosis

Background Mycobacterium bovis bacillus Calmette-Guerin (BCG) is an attenuated live vaccine that provides insufficient protection against tuberculosis (TB), the underlying mechanisms for which remain unknown. Assuming that the BCG vaccine inherits immune evasive strategies from virulent parent M. bovis strains, we aimed to identify the associated genes and assess their effects on the vaccine efficacy. Methods Three genes, BCG_3174, BCG_1782, and BCG_2432c, associated with immune evasion were first identified via bioinformatics analysis and then confirmed in the genome of M. bovis and 12 commercial BCG vaccine substrains using Polymerase Chain Reaction (PCR) and DNA sequencing. These genes were disrupted to develop mutant strains, and their effects on autophagy and their protective efficacy were further compared with the BCG vaccine in vitro and in vivo. Results Of the three identified genes, only the disruption of BCG_2432c, namely Delta BCG_2432c, conferred stronger protection against intranasal TB in vaccinated mice, when compared with the BCG vaccine. Delta BCG_2432c showed a stronger ability to trigger intracellular ROS-mediated complete autophagic flux in infected THP-1 cells that resulted in higher antigen presentation. The improved protection could be attributed to early and increased IFN-gamma(+) CD4(+) T-EM and IL-2(+)CD4(+)T(CM) cells in the spleens and lungs of Delta BCG_2432c-vaccinated mice. Conclusions The insufficient efficacy of the BCG vaccine is attributable to the important autophagy-inhibition gene BCG_2432c that blocks the autophagosome-lysosome pathway of antigen presentation. Delta BCG_2432c provides a promising platform to either replace the current BCG vaccine or develop vaccines that are more effective against TB.