Abstract
BACKGROUND: Toxoplasma gondii can cause congenital infections and abortions in humans. TgIST and TgNSM play critical roles in intracellular cyst formation and chronic infection. However, no studies have explored their potential to induce protective immunity against T. gondii infection. OBJECTIVE: To evaluate the immune efficacy of DNA vaccines encoding TgNSM and TgIST genes against T. gondii infection, using the acute and chronic ME49 strain (Type II). METHODS: DNA vaccines, including eukaryotic plasmids pVAX-IST and pVAX-NSM, were constructed. A cocktail DNA vaccine combining these two genes was formulated. The expression and immunogenicity were determined using the indirect immunofluorescence assay (IFA). Mice were immunized with DNA vaccines encoding either TgIST or TgNSM, as well as with the cocktail DNA vaccine. Humoral and cellular immune responses were analyzed by detecting antibody levels, cytotoxic T cell (CTL) responses, cytokines, and lymphocyte surface markers. Mouse survival and brain cyst counts were assessed 1 to 2 months post-vaccination in experimental toxoplasmosis models. The adjuvant efficacy of plasmid pVAX-IL-36γ in enhancing DNA vaccine-induced protective immunity was also evaluated. RESULTS: DNA immunization with pVAX-IST and pVAX-NSM elicited strong humoral and cellular immune responses, characterized by increased Toxoplasma-specific IgG2a titers, Th1 responses (including production of IFN-γ, IL-2, IL-12p40, and IL-12p70), and cell-mediated activity with elevated frequencies of CD8+ and CD4+ T cells, and CTL responses. This provided significant protective efficacy against acute and chronic T. gondii infection. Mice immunized with the two-gene cocktail (pVAX-IST + pVAX-NSM) showed greater protection than those immunized with single-gene vaccines. Co-administration of the molecular adjuvant pVAX-IL-36γ further enhanced the protective immunity induced by the cocktail DNA vaccine. CONCLUSIONS: TgIST and TgNSM induce effective immunity against T. gondii infection, making them promising vaccine candidates against toxoplasmosis. Additionally, IL-36γ is a promising genetic adjuvant that enhances protective immunity in a vaccine setting against T. gondii, and it should be evaluated in strategies against other apicomplexan parasites.