EsxG·EsxH (TB9.8·TB10.4) mycobacterial peptides enhanced BCG-induce protection in a murine model of pulmonary Tuberculosis

The attenuated <i>Mycobacterium bovis</i> bacillus Calmette-Guérin (BCG) vaccine is the only validated vaccine against tuberculosis (TB); however, its efficiency is limited in adults because it induces wide variability in protection and has limited immunologic memory. A prior <i>in silico</i> study using bioinformatic analysis and molecular dynamic simulations identified four peptides (G1, G2, H1, and H2) derived from the mycobacterial antigens TB10.9·TB10.4 (EsxG·EsxH) with predicted capacity to bind MHC-II molecules and potentially activate T and B cells. The present study aimed to experimentally validate these peptides as potential candidates for subunit vaccine boosters. We evaluated their cytotoxicity and protective efficacy of these peptides against <i>Mycobacterium tuberculosis</i> (Mtb) in a murine model, where they were administered as a booster following BCG immunization. Two months after BCG vaccination, mice received a subcutaneous boost with G1, G2, H1, or H2 peptides. One month later, the mice were challenged with the reference strain H37Rv. Specific IgG antibody titers were quantified by ELISA. Boosting BCG with these peptides enhanced protection compared to BCG alone, as evidenced by reduced pulmonary bacterial load and induction of peptide-specific IgG antibodies. These findings support the potential of epitope-based subunit vaccines as BCG booster. This approach provides a foundation for developing targeted booster strategies against TB.