Enhancing Mycobacterial Clearance via Heterologous BCG Prime Boost Strategy Using Recombinant Mycobacterium smegmatis
Ji, H.; Kang, K.; Kim, J.; Kwon, Y.; Kang, H.; Choi, U.; Kang, J.; CHOI, G.
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Bacille Calmette-Guerin (BCG) remains the only licensed tuberculosis (TB) vaccine, yet its protective efficacy against adult pulmonary TB is limited. To overcome this, we evaluated a novel recombinant Mycobacterium smegmatis (rMs) booster vaccine expressing Ag85B and ESAT-6 via the pMyong2 vector, combined with oat-derived {beta}-glucan adjuvant, to enhance BCG-primed immunity. BALB/c mice were primed with BCG and boosted twice with homologous BCG or varying doses of rMs with or without {beta}-glucan. Five weeks post-booster, mice were challenged intraperitoneally with M. tuberculosis H37Ra. Systemic bacterial clearance in liver tissue, cell-mediated immunity IFN-{gamma}, IL-17a, and antibody responses IgG, IgG2c, IgA were evaluated 3 weeks post-challenge. All rMs booster groups (rMs, high-dose rMs H, low-dose with {beta}-glucan, rMs {beta}) achieved complete systemic clearance of H37Ra with no detectable CFUs (colony forming unit), significantly outperforming mock-vaccinated and repeated BCG groups (**p < 0.01). Repeated BCG revaccination showed higher bacterial loads than single BCG (***p < 0.001), confirming no booster effect. Low-dose rMs with {beta}-glucan maintained 100% survival and total bacterial elimination despite a 50% dose reduction. Immune markers did not strictly correlate with clearance. Homologous BCG revaccination provides no additive protection, whereas pMyong2-based rMs confers robust systemic bacterial clearance. Adjuvanting with {beta}-glucan enables dose reduction while preserving maximum efficacy, establishing rMs as a promising heterologous TB booster candidate.
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