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

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.

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.

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.

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.