Evaluation on the Efficacy and Immunogenicity of Recombinant DNA Plasmids Expressing Spike Genes from Porcine Transmissible Gastroenteritis Virus and Porcine Epidemic Diarrhea Virus

Meng, Fandan; Ren, Yudong; Suo, Siqingaowa; Sun, Xuejiao; Li, Xunliang; Li, Pengchong; Yang, Wei; Li, Guangxing; Li, Lu; Schwegmann-Wessels, Christel; Herrler, Georg; Ren, Xiaofeng

To determine the immune response of mice to the DNA vaccination, plasmids were applied by intramuscular injection. At different times after vaccination, animals were sacrificed and analyzed for various parameters of the immune reaction. The proliferation of spleen T lymphocytes upon stimulation with purified S1 protein from either TGEV or PEDV was analyzed by MTT assays. As shown in Figure 4A , when stimulated with PEDV-S1 protein at 28 dpi, the proliferation levels of spleen T lymphocytes from mice injected with pIRES-(TGEV-S1-PEDV-S1) and pIRES-(PEDV-S1) were increased compared to those of control cells and the increase was highly significant (p,0.01). By 42 dpi, the proliferation levels of cells from mice injected with pIRES-(TGEV-S1-PEDV-S1) or pIRES-(PEDV-S1) were somewhat lower but still were significantly increased when compared to the control cells (p,0.01). By contrast, the proliferation levels of lymphocytes from animals immunized with pIRES-(TGEV-S1-PEDV-S) or pIRES-(PEDV-S) were highest at 42 dpi (p,0.01); no significant differences (p.0.05) were observed between the groups of pIRES-(TGEV-S1-PEDV-S) and pIRES-(PEDV-S). The proliferation values upon stimulation by TGEV-S1 protein are shown in Figure 4B . By 28 dpi the proliferation levels of spleen T lymphocytes from animals injected with pIRES-(TGEV-S1-PEDV-S1), pIRES-(TGEV-S1-PEDV-S) or pIRES-(TGEV-S1) were all significantly increased (p,0.01) relative to the PBS or pIRES controls. Furthermore, the values of the pIRES-(TGEV-S1-PEDV-S1) group were somewhat higher than those of the pIRES-(TGEV-S1-PEDV-S) group (p,0.05). By 42 dpi, the changes in the proliferation levels relative to controls remained highly significant (p,0.01) irrespective of the plasmid used for vaccination, pIRES-(TGEV-S1-PEDV-S1) or pIRES-(TGEV-S1-PEDV-S). No significant differences (p.0.05) were observed between the 28 dpi and 42 dpi values. Assays using PBMC to determine the proliferation of peripheral blood lymphocytes upon stimulation by PEDV-S1 protein were performed. At 14 dpi the proliferation of blood lymphocytes from mice immunized with pIRES-(TGEV-S1-PEDV-S1) or pIRES-(TGEV-S1-PEDV-S) was significantly increased (p,0.05) relative to the PBS and pIRES controls ( Figure 4C ) and this difference remained significant (p,0.01) when cells were analyzed at 42 dpi. The values of the two time points were similar in the case of the mice immunized with pIRES-(TGEV-S1-PEDV-S). In the pIRES-(TGEV-S1-PEDV-S1) group, the proliferation values at 42 dpi were slightly but significantly lower (p,0.05). As shown in Figure 4D , when stimulated with TGEV-S1 protein, the proliferation of PBMC was increased at 28 and 42 dpi in mice immunized with pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S) (p,0.01) relative to PBS and pIRES controls. In the group pIRES-(TGEV-S1-PEDV-S), the proliferation significantly increased from 28 to 42 dpi whereas the proliferation of PBMC from mice immunized with pIRES-(TGEV-S1-PEDV-S1) was similar at both time points.