Multi-epitope mRNA vaccine and protein vaccine protect mice against Toxoplasma gondii

Background Toxoplasma gondii , an obligate intracellular protozoan parasite, infects almost one-third of the world’s population and all warm-blooded animals, posing a substantial threat to public health. Accordingly, the development of effective vaccines against T. gondii has become an urgent priority. In this study, we constructed a multi-epitope chimeric antigen T-SGR targeting three key protective antigens of T. gondii (SAG1, GRA7, and ROP16), and developed both a messenger RNA (mRNA) lipid nanoparticle (LNP) vaccine and a recombinant protein vaccine based on T-SGR. The immunogenicity and protective effects were further evaluated in C57BL/6 mice. Methods The T-SGR mRNA–LNP vaccine was prepared via in vitro transcription followed by LNP encapsulation, while the T-SGR protein vaccine was obtained via prokaryotic expression and purification. Mice were administered a two-dose immunization regimen. Serum levels of specific IgG, IgG1, and IgG2a antibodies and cytokine levels were measured by enzyme-linked immunosorbent assay (ELISA). T lymphocyte subsets and lymphocyte proliferation were assessed by flow cytometry and Cell Counting Kit-8 (CCK-8) assay. Protective efficacy was evaluated by monitoring survival rates after challenge with highly virulent T. gondii RH strain tachyzoites and moderately virulent ME49 strain tachyzoites. Results Both T-SGR mRNA and protein vaccines induced robust humoral and cellular immune responses in mice. Notably, the IgG antibody titer induced by the mRNA–LNP vaccine was significantly higher than that of the protein vaccine ( P < 0.05). Both vaccines drove a Th1-biased immune response, as evidenced by markedly higher IgG2a levels relative to IgG1. Compared with the phosphate-buffered saline (PBS) control group, both vaccine groups significantly promoted splenocyte proliferation ( P < 0.05). The mRNA vaccine induced significantly higher secretion of IFN-γ, IL-10, IL-12, and IL-2 than the protein vaccine. Both vaccines conferred significant protection against T. gondii infection and prolonged mouse survival. Strikingly, the T-SGR mRNA–LNP vaccine provided 100% protection against the T. gondii ME49 strain, outperforming the recombinant protein vaccine. Conclusions We successfully developed a multi-epitope T-SGR mRNA–LNP vaccine and a recombinant protein vaccine against T. gondii . The T-SGR mRNA–LNP vaccine elicited stronger humoral and cellular immune responses and conferred superior protective efficacy, representing a promising candidate vaccine against toxoplasmosis.