The recombinant Erns and truncated E2-based indirect enzyme-linked immunosorbent assays to distinguishably test specific antibodies against classical swine fever virus and bovine viral diarrhea virus.

BACKGROUND: Classical swine fever (CSF) virus is the causative agent of an economically important, highly contagious disease of pigs. CSFV is genetically and serologically related to bovine viral diarrhea virus (BVDV). BVDV infection in pigs can mimic CSF clinical signs, which cause difficulty in differentiation. Serological test for detection of virus specific antibodies is a valuable tool for diagnosis and surveillance of CSFV and BVDV infections in animals. The aim of this study was to develop the CSFV Erns and BVDV tE2 -based ELISAs to distinguishably test specific antibodies against CSFV and BVDV. METHODS: The CSFV Erns and truncated E2 (tE2, residues 690-865) of BVDV were expressed in E. coli and purified by Ni-NTA affinity chromatography, respectively. Employing Erns or tE2 protein as diagnostic antigen, indirect ELISAs were developed to distinguishably test specific antibodies against CSFV and BVDV. The specificity and sensitivity of ELISAs were evaluated using a panel of virus specific sera of pigs, immunized rabbits and immunized mice. A total 150 clinical serum samples from farm pigs were measured by the developed ELISAs and compared with virus neutralizing test (VNT). RESULTS: Indirect ELISA was established based on recombinant CSFV Erns or BVDV tE2 protein, respectively. No serological cross-reaction between antibodies against CSFV and BVDV was observed in sera of immunized rabbits, immunized mice or farm pigs by detections of the Erns and tE2 -based ELISAs. Compared to VNT, the CSFV Erns -based ELISA displayed a high sensitivity (93.3%), specificity (92.0%) and agreement rate (92.7%), and the sensitivity, specificity and agreement rate of BVDV tE2 -based ELISA was 92.3%, 95.2% and 94.7%, respectively. CONCLUSION: The newly developed ELISAs are highly specific and sensitive and would be valuable tools for serological diagnosis for CSFV and BVDV infections.

Synergistic roles of the E2 glycoprotein and 3' untranslated region in the increased genomic stability of chimeric classical swine fever virus with attenuated phenotypes.

The E2 glycoprotein and 3' untranslated region (UTR) of classical swine fever virus (CSFV) are virulence determinants. To investigate the synergistic roles of E2 and 3'UTR for pathogenicity and genomic stability, a series of chimeric CSFVs were constructed by replacing the E2 gene and/or 3'UTR of virulent CSFV strain Shimen with the corresponding sequence of the lapinized 'Chinese' strain (C-strain) using a reverse genetic approach. The in vitro growth characterization and in vivo pathogenicity of the chimeric CSFVs were investigated. Our results demonstrated that the E2 glycoprotein mediates virus cell-to-cell spread and viral particle release and that the 3'UTR regulates viral RNA replication. The CSFV E2 and 3'UTR synergistically modulate infectious virus production, viral genomic stability in vitro, and attenuation in swine. This work contributes to our understanding of the structure and function of the CSFV genome and virus pathogenicity and will be useful for the development of a novel CSF vaccine.