Abstract:

A RT-PCR-RFLP assay was developed for the detection and differentiation between wild-type and rabbit attenuated vaccine viruses of classical swine fever virus (CSFV). A pair of specific primers was designed based on the Shimen strain, and used to verify the assay. Eight epidemic wild-type strains isolated from 20 clinical pathological samples of suspected swine fever and 2 vaccine strains were amplified by RT-PCR, cloned and sequenced. The results showed that a fragment of 825 bp was amplified from genomic RNA of CSFV wild-type which could be digested with ApaⅠ into two fragments of 322 bp and 503 bp when analyzed with RFLP, while the vaccine strain could not be digested with ApaⅠ. The lowest concentration of RNA could be detected in the present assay is 0.028 6 μg·mL^－1. Eight epidemic wild-type strains contained the ApaⅠ recognition sequence GGGCCC while the two vaccine strains contained the sequence GAGCCC. Eight epidemic wildtype strains belonged to Genomic Group 2. On the contrary, two vaccine strains with close genetic relationship to HCLV belonged to Genomic Group 1. The developed RT-PCR-RFLP could be used to detect and differentiate wild-type CSFV from pig vaccinated with the vaccine viruses.