Construction and safety,immunogenicity analysis of attenuated Salmonella typhimurium harbouring TGEV DNA vaccine

Abstract:

[Objective] To investigate the feasibility of using attenuated Salmonella typhimurium as carrier for oral immunization of TGEV DNA vaccine. [Methods] 2.1 Kb fragments of the TGVE SC-H strain S gene that encompasses all the four major antigenic domains was amplified by RT-PCR and cloned into eukaryotic expression vector pVAX1. The recombinant plasmid pVAX-S was transfected into COS7 cells and the expression of recombinant plasmids was identified by indirect immunofluorscence assay. Then pVAX-S was transformed by electroporation into attenuated Salmonella typhimurium SL7207, The recombinant was screened and designated as SL7207(pVAX-S). Mouse peritoneal macrophages were infected with SL7207(pVAX-S), the transcription and expression of S gene were detected by RT-PCR and indirect immunofluorscence . BALB/c mouse were inoculated orally with SL7207(pVAX-S) at dosage of 5×108、1×109、2×109CFU for safety analysis. In a vaccination test, BALB/c mouse were immunized orally with recombinant bacterial at dosage of 1×109 CFU, for three times and specific serum IgG and intestinal mucosa1 IgA antibody were detected by indirect ELISA. [Results] Recombinant plasmid pVAX-S was constructed correctly and expressed in COS7 cells. The transcription and expression of S gene was detected after mouse peritoneal macrophages were infected with SL7207(pVAX-S). The recombinant bacterium was safe to mouse at dosage of 2×109CFU. Specific serum IgG and intestinal mucosa1 IgA antibody against TGEV S protein were detected in SL7207 (pVAX-S) immunized group at 2 weeks post-boosting, and there were significant difference (P<0.05) in serum IgG and most significant difference (P<0.01) in intestinal mucosa1 IgA at 2 weeks post-three immunization compared with SL7207(pVAX) control group. [Conclusion] The recombinant Salmonella carried TGEV S gene DNA vaccines had good immunogenicity and safety in mouse.