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Tuberculosis (TB) remains a significant global health challenge, with approximately 1.5 million deaths per year. The Bacillus Calmette-Guérin (BCG) vaccine against TB is used in infants but shows variable protection. Here, we introduce a novel approach using a double gene knockout mutant (DKO) from wild-type Mycobacterium tuberculosis (Mtb) targeting fbpA and sapM genes. DKO exhibited enhanced anti-TB gene expression in mouse antigen-presenting cells, activating autophagy and inflammasomes. This heightened immune response improved ex vivo antigen presentation to T cells. Subcutaneous vaccination with DKO led to increased protection against TB in wild-type C57Bl/6 mice, surpassing the protection observed in caspase 1/11-deficient C57Bl/6 mice and highlighting the critical role of inflammasomes in TB protection. The DKO vaccine also generated stronger and longer-lasting protection than the BCG vaccine in C57Bl/6 mice, expanding both CD62L-CCR7-CD44+/-CD127+ effector T cells and CD62L+CCR7+/-CD44+CD127+ central memory T cells. These immune responses correlated with a substantial ≥ 1.7-log10 reduction in Mtb lung burden. The DKO vaccine represents a promising new approach for TB immunization that mediates protection through autophagy and inflammasome pathways. [ABSTRACT FROM AUTHOR]

The vaccinia virus (VACV) strain Western Reserve C16 protein has been characterized and its effects on virus replication and virulence have been determined. The C16L gene is present in the inverted terminal repeat and so is one of the few VACV genes that are diploid. The C16 protein is highly conserved between different VACV strains, and also in the orthopoxviruses variola virus, ectromelia virus, horsepox virus and cowpox virus. C16 is a 37.5 kDa protein, which is expressed early during infection and localizes to the cell nucleus and cytoplasm of infected and transfected cells. The loss of the C16L gene had no effect on virus growth kinetics but did reduce plaque size slightly. Furthermore, the virulence of a virus lacking C16L (vDeltaC16) was reduced in a murine intranasal model compared with control viruses and there were reduced virus titres from 4 days post-infection. In the absence of C16, the recruitment of inflammatory cells in the lung and bronchoalveolar lavage was increased early after infection (day 3) and more CD4(+) and CD8(+) T cells expressed the CD69 activation marker. Conversely, late after infection with vDeltaC16 (day 10) there were fewer T cells remaining, indicating more rapid clearance of infection. Collectively, these data indicate that C16 diminishes the immune response and is an intracellular immunomodulator.