Analysis Of The Complete Genome Sequence And Studies On Pathogenicity Of Duck Hepatitis Virus Type3SD1201Strain

Duck virus hepatitis (DVH), caused by duck hepatitis virus (DHV), is a highly contagiousand fatal disease in young ducklings. The disease mainly infects duckings which age is fromone week to three weeks. With the development of waterfowl industry in China, themorbidity and mortality rates of DHV are rising. DVH has become one of the major diseasesfor affecting healthy development of duck industry. In recent years, DVH caused by duckhepatitis A virus type3(DHAV-3) is widely popular in China and South Korea. For theclinical symptom, pathological changes and epidemiological characteristics, DHAV-3issimilar with DHAV-1, so it is difficult to identify the pathogen in diagnosis of DVH. In thisresearch, the complete genome sequence and the pathogenicity of SD1201strain DHAV-3,which has been separated from the Shandong area, were tested. This research mainly includesthe following contents:1. Sequence and analysis of the whole genome of DHAV-3SD1201strainThe genome sequence of DHAV-3SD1201strain was amplified by RT-PCR using primerswhich were designed according to the conserved region of DHAV-3genome. Sequenceanalysis showed that the genomic structure of DHAV-3SD1201strain is5’UTR-VP0-VP3-VP1-2A1-2A2-2B-2C-3A-3B-3C-3D-3’UTR, which was consistented withthe genome structure characteristics of Picornaviridae. The full genomic length of SD1201contained7791nt, with a large open reading frame (ORF) encoded2252amino acids.Compared with genome sequences, SD1201strain shared93.3%to98.8%of nucleotidehomology with other DHAV-3strains published in GeneBank, with the highest homologywith G strain and the minimum homology with B strain.2. Study on the pathogenicity of DHAV-3SD1201strainTo determines the virulence of DHAV-3strain SD1201, the median duck embryonal lethaldose (ELD50) and median lethal dose to ducklings (LD50) were tested. The result showed thatthe ELD50of SD1201strain was5-3.60/0.2mL and LD50was2-5.37/0.5mL. In animal experiment,120two-day-old ducklings were randomly divided into four groups and eachgroup was consisted of thirty ducklings. The ducklings in three groups were injected byintramuscular with0.5mL DHAV-3strain SD1201, corresponding to0.2LD50/each,2LD50/each and20LD50/each of DHAV-3respectively. The ducklings in the forth group wereinjected by intramuscular with0.5mL normal salin as control. At1h,2h,6h,12h,18h,24h,36h,48h,72h and96h post inoculation, the various tissues of ducklings were collected andtested the amount of viral load of DHAV-3by a real-time fluorescent quantitative RT-PCRmethod established by our laboratory. The result showed that DHAV-3could be detected inheart, liver, spleen, kidney, thymus and bursa, and the highest amount of virus was in the liver.It was found that the viral load had relationship with the different time of post inoculation andand the inoculation dose of DHAV-3.