Recombinant Eukaryotic Plasmid Of ROP16_?/? Failed To Provide Efficient Protection Against Toxoplasmosis In BALN/c Mice

To construct the recombinant eukaryotic expression plasmids of Toxoplasma ROP16?/?, and to assess the immune protection in BALB/c mice induced by ROP16?/? DNA vaccine.Methods:T. gondii R0P16?/? coding gene obtained by PCR amplification was inserted into the eukaryotic expression vector pEGFP-C2 to generate recombinant pEGFP-ROP16?/?. T293 cells were transfected by Liposome method in vitro and the fusion protein expression was identified by Western blotting. Thirty-six mice were randomly divided into 3 groups with 12 in each:PBS control group; empty plasmid group; and pEGFP-ROP16?/? group, respectively. Immunization was completed in each mouse by intramuscular injection once 2 weeks for 3 times. Sera at each time point was collected for detection of antibodies against Toxoplasma by indirect ELISA. After two weeks of the last immunization, mouse splenocytes were harvested and cultured for tests of cytokines in the supernatant. The remaining mice were injected with Wh3 tachyzoites, with 1000 parasites for each, followed by observation of survival time and mortality of the animals.Results:The recombinant eukaryotic expression vector of pEGFP-ROP16?/? was successfully generated and efficient expressions were noted both in the transfected T293 cells and the muscles in which the plasmid vaccine was administered. All vaccinated mice, however, did not present the increased IgG antibodies and cytokine levels after pEGFP-ROP16?/? immunization. Correspondingly, no difference was seen of either survival or mortality of the animals compared with the control.Conclusion:Our conclusion is that Toxoplasma ROP16?/? DNA vaccine failed to provide BALB/c mice with efficient immune protection against virulent tachyzoites challenge due to its inaccessibility to host immune components and its lack of linear B epitopes.