Serosurvey Of Major Bovine Resporitary Viruses And Identification Of BVDV Isolates And Vaccine Development

Bovine respiratory disease complex (BRDC) is a serious disease in cattle industry worldwide.Viruses including bovine herpesvirus1(IBRV), bovine virus diarrhea virus (BVDV), bovine respiratorysyncytial virus (BRSV) and bovine parainfluenza virus type3(BPIV3) are recognized as the majorprimary pathogens inducing the BRDC. The aim of the study was to investigate the seroprevalence ofthe four viruses in major dairy and beef cattle breeding and farming regions in China.Serum samples (n=3503) collected from147farms of14provinces in China were examined byvirus neutralization test (VNT) and the results were analyzed by the SPSS software. Our surveillancestudy showed that the overall sample seropositive rates in the investigated areas for BVDV, BoHV-1,BPIV3and BRSV were56.5%,33.3%,67.2%and69.6%, respectively. The seroprevalence indifferent provinces ranged from5.2%to84.9%for BVDV,3.1%to95.8%for BoHV-1,27.8%to97.3%for BPIV3and41.2%to94.4%for BRSV.A new isolate of bovine viral diarrhea virus (BVDV), named HN01, was isolated from the nasalswabs using MDBK cell culture. The HN01strain caused cytopathic effect (CPE) in MDBK cellcultures after two passages. The virus specifically reacted to BVDV1-specific monoclonal antibody inan immunofluorescence assay. A fragment of288bp of genome from this isolate was amplified by theRT-PCR. Phylogenetic analysis of5′UTR indicated that the virus was BVDV type1strain, belong tosub-genotype1a. In the pathogenesis study, four calves experimentally infected with the BVDV straindeveloped depression, cough and other clinical signs. Calves showed high temperature over40°C, andwhite blood cell counts dropped more than40%. The experimental infection showed that the virus wasmoderate pathogenic to cattle and can be used as a BVDV challenge virus to evaluate the efficacy ofBVDV vaccines in the target animals.A noncytopathic BVDV2b strain named SD1301was firstly isolated and identified inChina. Thevirus was sequenced, and the virulence was analyzed. The genome of the virus is12258nt in length andcontains a5′UTR, one open reading frame (ORF) encoding a polyprotein of3897amino acids and3′UTR. The complete genome was submitted to GenBank with the accession number KJ000672.Phylogenetic analysis of5′UTR, Npro, E1and E2gene demonstrated the virus had high similarity withthe strain Hokudai-Lab/09. In the experimental infection study, all calves developed clinical signs withrectal temperature higher than the base temperature1℃at least1day, and white blood cells (WBC)and platelet decreased by over40%. Grossing anatomy found that the lymph lode leukopenia,lymphopenia, neutropenia, thrombocytopenia. The virulence of the isolate suggested that the virus weresimilar with a moderate virulent strain. This was the first confirmed isolation of ncpBVDV2b foundcirculating in China.The immunogenicity and efficacy of an inactivated Chinese BVDV1a NM01vaccine strain wasevaluated by challenging with a Chinese BVDV1b JL strain. Five2-4month old calves were intramuscularly vaccinated with a single dose of the vaccine strain and boosted with same dose threeweeks after the first vaccination, with five mock immunized calves serving as a control group.Twenty-one days post the second vaccination; all calves were challenged with6×106.8TCID50of thechallenge strain JL. The median serum neutralizing antibody titer of the immunized calves reached1:410, while the control calves were seronegative at21days post the second vaccination. The clinicalsigns, such as the temperature and leukopenia of the immunized calves and viral shedding, weresignificantly less than the mock immunized calves after challenging with the virulent BVDV1b strain,indicating that the BVDV1a vaccine strain elicited efficacious protection against the endemic BVDV1bstrain in China. The vaccine candidate developed with predominant BVDV1a and1b strains may beeffective in protecting against both BVDV1a and BVDV1b infections in China.