Recombinant vaccinia virus expressing MIC8, AMA1, or RON4 induce protection against Toxoplasma gondii ME49 strain infection

Toxoplasma gondii (T. gondii) infection is a globally prevalent and potentially severe disease, particularly in infants and immunocompromised individuals. Despite its widespread impact, no licensed vaccine is currently available for human use. The T. gondii antigens MIC8, AMA1, and RON4 have been identified as candidates capable of inducing immunity against toxoplasmosis. In this study, we generated recombinant vaccinia viruses (rVV) expressing MIC8 (MIC8-rVV), AMA1 (AMA1-rVV), or RON4 (RON4-rVV), and evaluated the vaccine efficacy of each rVV in mice. Mice were intranasally immunized twice with rVV and subsequently challenged with T. gondii ME49 strain. All three rVV vaccines induced T. gondii-specific immunoglobulin (Ig)G and IgA responses, as well as significant activation of CD8⁺ T cells and total B cells. All immunized mice 100% survived a lethal challenge, whereas the naïve control group did not. Among the three vaccines, MIC8-rVV elicited the strongest CD8⁺ T cell and B cell responses, resulting in the highest reduction in brain cyst counts following T. gondii ME49 strain challenge. These findings demonstrate that rVV-based delivery of MIC8, AMA1, and RON4 antigens can confer protective immunity against T. gondii ME49 strain infection, with MIC8 emerging as a particularly promising vaccine candidate.