Immunogenicity and protective efficacy of recombinant BCG expressing a multi-epitope vaccine against mycobacterium tuberculosis.

INTRODUCTION: Mycobacterium tuberculosis (Mtb) infects approximately one-quarter of the global population, of whom 5%-10% develop active tuberculosis (TB). Although Bacillus Calmette-Guérin (BCG) remains the cornerstone vaccine against severe infant TB, its efficacy in adults is limited, underscoring the need for novel TB vaccines. This study aimed to design a multi-epitope vaccine, tMEV, and validate its protective potential through bioinformatic prediction and in vivo experiments.

METHODS: tMEV was constructed from dominant T- and B-cell epitopes derived from Mtb ESAT-6, PE35, PPE38, and PE4 proteins, fused with an adjuvant. Its vaccine candidacy was assessed via physicochemical and solubility profiling, secondary and tertiary structure modeling, TLR-4 molecular docking, molecular dynamics simulation, and immune simulation. The tMEV coding sequence was cloned into pMV361 and transformed into BCG to generate recombinant BCG (rBCG-tMEV). Immunogenicity, antigenicity, and protective efficacy were evaluated in mouse immunization and Mtb challenge models.

RESULTS: tMEV exhibited favorable vaccine-like properties in silico. In mice, both tMEV and rBCG-tMEV induced Mtb-specific immune responses. Compared with PBS, tMEV significantly enhanced cellular and humoral immunity with a Th2-biased profile. Relative to conventional BCG, rBCG-tMEV elicited a superior mixed Th1-Th2 response and higher antigen-specific immunity, accompanied by increased secretion of IFN-γ, IL-2, TNF-α, granzyme B, and perforin. Moreover, rBCG-tMEV markedly reduced bacterial loads in the lungs and spleens of Mtb-infected mice, conferring partial protection.

DISCUSSION: These findings demonstrate that tMEV and rBCG-tMEV possess robust immunogenicity and protective potential against Mtb, representing promising novel TB vaccine candidates and providing a rationale for future vaccine development and optimization.