Development of a vaccine against Toxoplasma gondii, the cause of abortion in sheep

Project no. 25279 · Status: Kapanmış (Yayın bekleniyor) · 2023

Summary

Toxoplasma gondii is an important zoonotic protozoan that causes abortus, stillbirth and loss of offspring in sheep. Infected animals not only cause economic losses but also pose a risk to public health. The only licensed commercial vaccine against this infection, TOXOVAX® (MSD, New Zealand), contains the live attenuated S48 strain and is effective in preventing congenital toxoplasmosis. However, the vaccine's limited shelf life, cold chain requirements, and the risk of the live parasite gaining virulence highlight the need for safer vaccine platforms. In this context, this project aims to develop a pDNA vaccine capable of inducing an immune response against T. gondii infection in sheep and to evaluate its immunogenicity. To achieve this goal, a DNA vaccine design was first developed. The GRA8 protein, which is expressed throughout all life stages of the parasite, was selected as the vaccine antigen. Valine was added to the amino acid sequence to increase protein stability, and the protein sequence was optimised using sheep codons. The sheep codon-optimised GRA8 (scoGRA8) gene region was cloned into the pVAX1 vector to obtain the pVAX1-scoGRA8 DNA vaccine. The in vitro expression of the vaccine was performed in HEK293T cells, and the protein expression level of the vaccine was confirmed by Western blot, immunofluorescence staining (IFAT), and quantitative real-time PCR tests. Sheep were administered a total of three doses of the vaccine via IM and EP routes. The humoral immune response induced by the vaccine was evaluated using ELISA and Western blot tests after three doses of vaccination. The cellular immune response was analysed by measuring IFN-γ levels in the supernatants of peripheral mononuclear blood cells (PKMHs) isolated from blood samples taken two weeks after the third dose and stimulated with tachyzoite lysate antigen (TLA), as well as in serum samples taken at the same time. The data obtained showed that the pVAX1-scoGRA8 vaccine successfully expressed the recombinant GRA8 protein, elicited antigen-specific IgG antibody responses in both vaccine groups, and increased IFN-γ production from CD4+ and CD8+ T cells. Furthermore, immunological response analyses revealed no statistically significant difference between the IM and EP vaccine groups. These findings indicate that the developed DNA vaccine effectively induces both humoral and cellular immune responses and represents a promising vaccine candidate against sheep toxoplasmosis.

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