Strategic deletion of genes in BCG leads to an enhancement in immunogenicity 2267476

Abstract Introduction Tuberculosis (TB) remains one of the most pressing public health crises, attributed to the bacterium Mycobacterium tuberculosis (Mtb) and resulting in a staggering 1.25 million deaths in 2023 alone. The only approved vaccine for TB is the Bacille Calmette-Guérin (BCG), which is typically administered at birth. While BCG is effective in preventing miliary and meningeal TB in children, it offers only limited protection against pulmonary TB in adults. This limitation highlights the urgent need for an improved BCG vaccine or alternative TB vaccines. Methods We hypothesized that the BCG vaccine could be enhanced by deleting genes that encode immune-evasive proteins. To test this hypothesis, we selected three specific genes: sapM, nuoG, and zmp1, whose products interfere with phagolysosomal (PL) fusion, autophagy, and apoptosis in macrophages. Results We created three genetically modified vaccine strains in the BCG Pasteur background: single knockout (SKO; ΔsapM), double knockout (DKO; ΔsapM and ΔnuoG), and triple knockout (TKO; ΔsapM, ΔnuoG, and Δzmp1). We then evaluated these genetically modified BCG strains for their potential to enhance PL fusion, autophagy, antigen presentation, and ex vivo immunogenicity. The results indicated that macrophages process SKO, DKO, and TKO strains more effectively via PL fusion and autophagy, thereby enhancing antigen presentation. The ex vivo immunogenicity results correlated with the in vitro results. Conclusion Our findings indicate that the immunogenicity of BCG can be improved by selectively deleting genes associated with immune evasion. Funding Source NIH Topic Categories Vaccines and Immunotherapy (VAC)