Biological Identification Of Paramyxovirus Isolated From Ducklings And Duck-embryos

Duck Paramyxovirus Disease (duck-origin Newcastle disease) is an acute septic infectious disease that caused by duck paramyxovirus (duck-origin Newcastle disease virus). The disease is mainly characterized by diarrhea, neurological symptoms, focal glandular gastric bleeding or ulcers, intestinal mucosal bleeding and necrosis of the pancreas or the spleen.Ducks are usually infected concentrated at 5 to 8 weeks of age. And the rates of morbidity and mortality are separately up to 20% to 60% and 10% to 15%, even both are as much as 90% in one or two duck fowles. It is generally believed that the paramyxovirus is disseminated by horizontal transmission primarily through diseased poultry carcasses, offals, excrements or secretions and contaminated feed, water, grass, appliances, etc. via the digestive tract, respiratory organ or conjunctiva. One strain of a virus has been isolated from a 1 day old duckling which symptom was suspected Paramyxovirus disease and dead duck embryos in our lab in 2008. And this strain of virus has been identified as paramyxovirus through the biological characteristics test and amplified F gene fragments by RT-PCR. Therefore, we suspect that there exists such a duck paramyxovirus which can be transmitted vertically through hatching eggs.In this study, the paramyxovirus was isolated and identified from a 1 day old duckling which symptom was suspected Paramyxovirus disease and dead duck embryos during 2008 to 2009. And the biological characteristics and genetic characteristics of the 4 strains isolates of duck paramyxovirus had been studied. The pathogenicity index MDT and ICPI were separatedly 50.4-58.0 and 1.68-1.75, and these showed the isolates was velogen strain.The pathogenicity test showed that the representative strains had strong pathogenicity to the ducklings of different ages and the morbidity and mortality were sepqrately 70% to 100% and 20% to 100%. F gene of the four isolates amplified and sequence analyzed by using RT-PCR showed that the homology between the four strains was 99.8%-100%, and homology between the four strains and the strain SDFCH isolated from 1 day old ducklings from Feicheng region's in our laboratory before was 99.5%-99.8%, and homologies between the four strains and other ducks, chicken, goose, pigeon NDV were 88.1%-98.4%, 88.4%- 98.7%, 88.6%-97.6% and 90.8%-94.8%. Genetic evolution analysis showed that 4 strains duck paramyxovirus and SDFCH in the same branch, belonged to the geneâ…¦d-type. In view of the highly homeology between isolated strains and SDFCH strain, highly similar genetic relationship, we conjectured that it may be related to the way transmitted vertically through hatching eggs.19 pairs of primers were designed and synthesized according to the genome sequence of NDV published in GenBank. And the genome of isolated strain SDWF02 was amplified and sequence analyzed by using RT-PCR. The results show that, the length of gene of SDWF02 was 15192bp and which was the same with NDV geneâ…¤-â…¦andâ…¨published in GenBank.And there existed 6bp insertion at the 5' non-coding region of the NP gene.The whole genome includes the 3' leader sequence, 5' trailing sequence and 6 open reading frame, and the gene order is 3'-NP-PMF-HN-L-5'. The first 12 nucleotides of the SDWF02 gene 3 ' and last 12 nucleotides of the SDWF02 gene 5' are complementary pairs. the starting sequences and terminator sequences of each have highly homology with other relevant NDV sequences. The length of the intergenic sequence of NP-P, P-M, M-F, F-HN and HN-L are 1bp, 1bp, 1bp, 31bp, and 47bp. The results of homology analysis showed that, SDWF02 strain has highly homology with genotypeâ…¦strains especially chicken/China/Guangxi9/2003, chicken/China/Guangxi11/2003, GM, Muscovy duck/China(Fujian)/FP1/02, Laying duck/JSD0812/2008, goose/ZJ1, goose/SF02 that isolated from home and abroad in recent years, and the homology is 97.0%-97.3%; SDWF02 strain has low homology with other genotypes NDV strains and traditional strains Lasota, F48E9 and Mukteswar, and the homology is only 82.8%-89.1%. In addition, compared with other strains, SDWF02 strain has several its own specific amino acids emerged on the NP, P, M, HN and L proteins.We conjectured that those amino acids may cause the strains to be passed by duck embryos.