Detection And Sequence Analysis Of Influenza A Virus And Newcastle Disease Virus From Wild Ducks Wingting In Wetlands

Avian influenza virus (AIV) and Newcastle disease virus(NDV) are pathogens of social and economical importance known to be disseminated throughout the world by migratory birds. To improve the efficacy and the sensitivity of different diagnosis for NDV and ATV, a duplex reverse transcription-polymerase chain reaction (RT-PCR) technique was established in the present study. To do this, two sets of primers were designed according to the sequences of AIV and NDV published in GenBank. One pair of primer was 190bp and the other was 363bp in length, were expected to be amplified, respectively. After the two sets of primers were combined and used for amplification trials, the conditions of PCR for detection of the two viruses were optimized by orthogonal assay. Based on the single RT-PCR assay that detected each of the two viruses respectively, a duplex RT-PCR system for both of them in one tube was established to distinguish the two viruses. The results suggested that the established duplex RT-PCR system was much more sensitive, specific, rapidly and could be applied for early clinical diagnosis of AIV and NDV. Furthermore, it could also be considered to be used on the research of pathogenesis of the two viruses.70 cloacal swabs from 70 feral ducks including 8 types wintering in Hongze Lake in Jiangsu province collected from February 2007 were checked for the presence of influenza A viruses and Newcastle disease virus using duplex RT-PCR. Results suggest that the prevalence of influenza viruses is up to 14.29%(10/70). The prevalence in Ruddy Shelduck, Spotbilled duck is up to 25%. The results reveal that enormous pools of influenza viruses are present in migratory ducks in these areas. Sequences analysis of NS gene suggest that these viruses maybe a great threat to public and poultry health. The prevalence of Newcastle disease virus is up to 1.43%(1/70). Sequences analysis of F gene showed that the identities with Lasota,B1 strains were high.Analysis of M gene nucleotide sequences and deduced amino acid sequences from five isolates suggest that M gene of spdkXY605,spdkXY705,spdkXY905,maXY704, btHZ1405 are closely related to HPATV H5N1 from poultry in Hebei,Jilin Province which indicate that migratory ducks wintering in Jiangsu Province maybe involved in long distance transmission of avian influenza viruses .