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

Prevalence of TGE and PED may lead to severe economic losses in swine-raising countries. So far, the most effective prevention measure is vaccination. DNA vaccines can induce complete immune responses, provide heterologous cross protection and can be easily prepared as polyvalent vaccines [23] . It is documented that the S1domain of TGEV/PEDV contains the main neutralizing epitopes [13, 20, 34, 39] . One report indicated that the immunogenicity of DNA plasmid bearing TGEV S1 gene is superior to another version containing the full-length S gene [29] . In contrast, there are no reports regarding comparative analysis on the efficiency of S and S1-based DNA plasmids. Therefore, this study aimed at evaluating immune response induced by DNA plasmids encoding the TGEV S protein and the PEDV S or S1 proteins using a mouse model. In pIRES vector, there is an internal ribosomal entry site (IRES), which is derived from the encephalomyocarditis virus (ECMV); two multiple cloning sites (MCS) A and B, allow each product of transcription to be translated independently with the participation of ribosome at the same time. In addition, pIRES vector has a CMV promoter for high expression of foreign genes and a SV40 enhancer/promoter that allows the enhancement of gene expression in many hosts [33] . Therefore, the genes of interest in this Figure 7 . Antibody levels in mice treated with pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S). Anti-PEDV serum antibodies (A) and anti-TGEV serum antibodies (B) in pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S) immunized mice were detected by indirect ELISA at different time points following the injection of the plasmid DNAs. The OD 490 was monitored as a function of time over a period of 42 days. w: p,0.01 (highly significant), compared with PBS, pIRES treatment groups. doi:10.1371/journal.pone.0057468.g007 study were cloned into the pIRES vector. Both immunufluorescence assays and Western blot indicated that the in vitro expression of TGEV S, PEDV S or PEDV S1 proteins were successful. It is clear that the level of neutralizing antibodies is an important indicator to evaluate the effect of the vaccine. In this study, IgG levels of PEDV antibodies in pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S) groups increased between 21-42 dpi, and the differences between both groups were not significant. In addition, the values of the pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S) groups peaked at 35 dpi and decreased thereafter, but the differences between these two groups were also not significant. The IgG antibody levels against TGEV of these two groups began to increase at 28 dpi and reached the peak at 35 dpi. Between 35-42 dpi, the antibody levels decreased dramatically (p,0.01) in pIRES-(TGEV-S1-PEDV-S1) group, whereas no significant changes was observed (p.0.05) in the pIRES-(TGEV-S1-PEDV-S) group. This result showed that the recombinant plasmid pIRES-(TGEV-S1-PEDV-S) containing the full-length S protein of PDEV was better in eliciting immune responses to PEDV than is pIRES-(TGEV-S1-PEDV-S1) encoding only the S1 portion of the S protein. The IgG antibody levels and virus neutralizing assays showed that both pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S) groups could elicit humoral immune responses against PEDV and TGEV S proteins, respectively. Plasmid pIRES-(TGEV-S1-PEDV-S) was more efficient in inducing neutralizing antibodies than pIRES-(TGEV-S1-PEDV-S1).