Evaluating the immune response and protection conferred by vaccine candidate toxoplasma gondii GRA8 protein

Toxoplasma gondii is a zoonotic protozoan that can infect all warm-blooded animals including humans and may lead to serious clinical manifestations especially in immunocompromised and pregnant humans and animals. Almost 30% of the world population is infected with T. gondii. Along with the treatment cost of the disease, the agent may cause large economic losses in farm animals. Therefore, it is very important to develop a vaccine against T. gondii. For this purpose, GRA8 protein that showed high immunogenicity in our previous study was firstly examined by bioinformatics. After removing intracellular and transmembrane fragments of the protein, a signal peptide was added. The naked GRA plasmid and the CpG+Escort+GRA8 plasmid DNA vaccines were developed by cloning the gene fragment into the pcDNA.3.3 vector. Swiss Webster mice were vaccinated twice, at three weeks intervals. The sıvısal immune response was determined both Western Blot and ELISA, and the cellular immunity was determined both flow cytometer and MTT assay. Protection efficacy of DNA vaccines was determined by challenging each mice orally with 7-8 cysts of T. gondii PRU strain. According to the results, GRA8 plasmid and CpG+Escort+GRA8 plasmid vaccine groups induced strong IgG response compared to controls (P<0.001). As we examined the IgG1 and IgG2a responses, a balanced Th1-Th2 polarization was seen. IFN-γ secreting CD4 T lymphocyte ratios increased 1.45 to 2 and 1.6 to 2 times compared to the controls. In the vaccine groups, IFN-γ secreting CD4 T and CD8 T lymphocyte ratios increased 1.31 to 1.75 and 1.17 to 1.70 times compared to the controls, respectively. Stimulation index were found to be high in the vaccine groups which was between 7% and 34% compared to the control groups. The number of tissue cysts in the GRA8 plasmid group was 440.74±343.35 that decreased significantly compared to PBS and CpG+Escort+Empty plasmid groups (P<0.0001, ****). In CpG+Escort+GRA8 plasmid group, the number of tissue cysts found to be 804.76±715.87, which decreased significantly compared to PBS (P=0.0086, **) and CpG+Escort+Empty plasmid groups (P=0.0007, ***). The amount of tissue cysts in the GRA8 plasmid vaccine group was significantly lower than the CpG+Escort+GRA8 plasmid group (P<0.05, *). Overall, DNA vaccines developed from vaccine candidate GRA8 antigen which was selected by protein microarray screening showed strong sıvısal and sıvısal immune response and provide partial protection against toxoplasmosis mice model. Furthermore, development of multi-antigenic DNA vaccines using GRA8 antigen and other T. gondii antigens future studies may provide complete protection against toxoplasmosis. Keywords: Toxoplasma gondii; GRA8 protein; DNA vaccine; Nano-adjuvant; CpG