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Protective immunity induced in mice by multiantigenic DNA vaccine with genes encoding SAG1 and MIC8 of Toxoplasma gondii.

OBJECTIVE
To observe the immunoprotection induced by multiantigenic SAG1-MIC8 DNA vaccine of Toxoplasma gondii in C57BL/6J mice.


METHODS
The sequences of genes encoding SAG1 and MIC8 protein were inserted into the eukaryotic expression vector pcDNA3.1 and the multiantigenic recombinant plasmid pcDNA3.1-SAG1-MIC8 was constructed. Then the recombinant plasmid was transfected into Hela cells to test its expression and the recombinant protein characterized by Western blotting 70 mice were divided into 5 groups randomly: PBS, pcDNA3.1, pcDNA3.1-SAG1, pcDNA3.1-MIC8 and pcDNA3.1-SAG1-MIC8. Each mouse was injected intra-muscularly by 100 microg recombinant plasmid for 3 times every two weeks. Mice were bled on day 0, 13, 27, 41, and 55. Four weeks after the final inoculation (on day 56), spleens from seven immunized mice per group were collected. Another seven immunized mice per group were intraperitoneally challenged with 1 x 10(4) tachyzoites of RH T. gondii and the survival time was observed. Serum IgG antibody and cytokines IFN-gamma and IL-4 were demonstrated by ELISA and the T lymphocyte proliferation assay were carried out with 3H-TdR incorporation.


RESULTS
Western blotting showed that the mature protein extracts in Hela cells upon transfection with pcDNA3.1-SAG1 (Mr 34,000), pcDNA3.1-MIC8 (Mr 74,000) and pcDNA3.1-SAG1-MIC8 (Mr 109,000) were effectively expressed in cells. The results of IgG antibodies (on day 41 and 55), IgG2b, IgG2c, IFN-gamma (on day 55) and T lymphocyte proliferation assay (on day 56) were more obvious in mice immunized with pcDNA3.1-SAG1-MIC8 multiantigenic DNA vaccine than those in mice with single-gene plasmids (P < 0.05). There was no significant difference in IgG1 and IL-4 levels between vaccinated and control mice after the final inoculation (on day 55) (P > 0.05). The median survival time was 3, 4, 7, 7, and 10d, respectively, with considerable difference among the groups (P < 0.01).


CONCLUSION
The multiantigenic DNA vaccine elicits a stronger immuno-protection in mice than the monovalent DNA vaccine.