Evaluation of Immunogenicity of Mycobacterium tuberculosis ag85ab DNA Vaccine Delivered by Pulmonary Administration

<b>Background:</b> Tuberculosis (TB) is a respiratory infectious disease, and the current TB vaccine has low local lung protection. We aim to optimize immune pathways to improve the immunogenicity of vaccines. <b>Methods:</b> In the immunogenicity study, 50 BALB/c mice were randomly divided into the following: (1) phosphate buffered saline (PBS)+intramuscular injection combined with electroporation (EP) group (100 μL), (2) pVAX1+EP group (50 μg/100 μL), (3) <i>ag85ab</i>+EP group (50 μg/100 μL), (4) pVAX1+pulmonary delivery (PD) group (50 μg/50 μL), and (5) <i>ag85ab</i>+PD group (50 μg/50 μL). Immunization was given once every 2 weeks for a total of three times. The number of IFN-γ-secreting lung and spleen lymphocytes was determined by enzyme-linked immunospot assay (ELISPOT). The levels of Th1, Th2, and Th17 cytokines in the culture supernatants of lung and spleen lymphocytes were detected with the Luminex method. The proportion of FoxP3 regulatory T cells in splenocytes was determined by flow cytometry. The levels of IgG-, IgG1-, and IgG2a-specific antibodies in plasma and IgA antibody in bronchoalveolar lavage fluid (BALF) were determined by enzyme-linked immunosorbent assay (ELISA). <b>Results:</b> The PD and EP routes of <i>Mycobacterium tuberculosis</i> (<i>M. tb</i>) <i>ag85ab</i> DNA vaccine can effectively induce the responses of IFN-γ-secreting lung and spleen lymphocytes, and induce dominant Th1 and Th17 cell immune responses. The PD route can induce earlier, greater numbers and stronger responses of pulmonary effector T cells, with higher levels of the specific antibody IgA detected in BALF. High levels of the specific antibodies IgG, IgG1, and IgG2α were detected in the plasma of mice immunized by the EP route. <b>Conclusions:</b> The PD route of DNA vaccines can more effectively stimulate the body to produce strong cellular and mucosal immunity than the EP route, especially local cellular immunity in the lungs, which can provide early protection for the lungs. It can significantly improve the immunogenicity of the <i>ag85ab</i> DNA vaccine, suggesting a feasible and effective approach to DNA immunization.