Replication Characteristics Of A Bovine Viral Diarrhea Virus(SDO803) From Pig Passaged On MDBK

Bovine viral diarrhea virus (BVDV) causes infection of ruminant animals, and swine werereported to be infected by BVDV in China in middle of90s. So far, few researches about thepathogenesis mechanism and disease hazard of pig BVDV are demonstrated. In this study,191clinicalsamples collected from the provinces of Shandong, Zhejiang, Anhui, Guangxi, Jiangsu, Tianjin, Hubeiand Shanghai city and30commercial vaccines (PRRSV, CSFV, PRV) were detected by RT-PCR. Theresults showed that23out of191samples were BVDV positive, and10out of30vaccine samples werepositive.8in108healthy pigs were detected to be positive (8.33%), and15in85sick pigs weredetected to be positive (17.6%), which indicated that BVDV may play a role in the process ofpathogenesis of disease pigs. The results also showed that commercial swine vaccines in China werecontaminated by BVDV at a high level, suggesting that pigs inoculated with BVDV-infected vaccinesmay be one of most important transmission routes of BVDV infection. The RT-PCR productions of sixpositive samples were sequenced and analyzed by bioinformatics software. The results showed that thehomology of six nucleotide sequences was99.4%-100%. But compared with standard reference strainssequences, the homology was between71.8%and95.8%. And six samples were phylogeneticallyanalyzed to be BVDV-1b. These results further indicated pigs inoculated with BVDV-infected vaccinesmay be one of most important transmission routes of BVDV infection.To obtain a cell-adapted BVDV stain, BVDV SD0803strain was serially passaged in MDBK celllines up to forty times (P0-P40). RT-PCR and IFA were performed to confirm the replication of BVDVin cells. To identify genetic changes associated with adaptation of the BVDV SD0803strain, wedetermined the nucleotide changes that arose during MDBK passage (40) of the BVDV SD0803strain.Sequencing results showed that the nucleotides homology for BVDV P40and parental virus was99.8%,and amino acid homology was99.6%. The BVDV P40strain contained a23nucleotide mutationrandomly across the genome, resulting in15aa mutation located mostly in E2and NS5B. To determinethe growth characteristics between the parent virus and passaged viruses, supernatant samples werecollected at the indicated time points, and virus growth was measured by quantitation of virion RNAreleased from MDBK cells. The growth curve showed that both parent and passaged viruses had highreplication efficiency in MDBK cells, and passaged viruses did not produce higher virus replicationefficiency than parent virus. Although some mutations happened during virus passaging, it did notimproved virus titer in cells. This study provides significant meanings for candidate vaccine of BVDVin pigs.