A Surveillance Study Of Avian Influenza Viruses In Qinghai Lake Region

Objective:Qinghai Lake is located in the qinghai-tibet plateau in northwest of China, being migratory birds’wintering and natural habitats during the breeding season.There are three migratory birds’fly way:Central Asia migration flyway, east Asia-Australia’s flyway and east Africa-west Asia migration flyway, and the three major international migratory bird migration flyways across Qinghai province. In order to understand the distribution of avian influenza virus in Qinghai Lake region, we conducted an active surveillance program on influenza A viruses in Qinghai Lake region.Methods:A total of3680environmental samples were collected from Qinghai Lake area. All samples were inoculated into specific pathogen free (SPF) chicken embryo eggs for virus culture.The isolates with positive hamegglutinin assay were indentified as influenza nucleic acid positive by real-time PCR method. The full genome of the isolates were sequenced and analyzed.Results:1. We have collected3680environmental samples in this study. A total of27strains of influenza A virus were separated, the isolation rate was0.73%.13strains are H9N2subtype viruses.7strains are H5N1subtype viruses.4strains are H4N6and2strains are H13N8.1strain is H3N2subtype virus. The proportion of each subtype were48.15%(H9N2),25.93%(H5N1),14.81%(H4N6),7.4%(H13N8),3.7%(H3N2) respectively. 2. The positive rate of the poultry samples was different in different month. The positive rate of the samples collected in September2012was the highest (1.3%), and the positive rate of November was the lowest (0.2%).3. The analysis of different sampling sites showed that the positive rate was different. Ledu live poultry market has the highest positive rate of influenza A virus (4.18%), and No virus was isolated from Gangca live poultry market, Haiyan farm and Sha island.4. The analysis of different sample types showed the positive rate was different. specimens of cage surface has the highest positive rate of influenza A virus (2.51%), stool specimens was the lowest (0.52%).5. The phylogenetic analysis showed that H017, QH011, QH025, QH071, QH368and2strains of subtype H13N8AIVS are Eurasian lineage virus, and5strains of H9N2subtype avian influenza virus H017, QH011, QH025, QH071, QH368were new reassortant viruses of Y280-like lineage, Gl-like lineage and F98-like lineage.Two strains of H13N8viruses were reassortant viruses between gulls and ducks viruses. Amino acids site228of HA of two H13N8subtype viruses has a mutation (G228S). Further biological assay showed that mutation has improved the binding afinity of a-2,6receptor.Conclusions:In addition to the subtype H5N1reported previously, subtype of avian influenza virus H9N2, H3N2, H4N6and H13N8subtype also were popular in Qinghai Lake region. the genome sequence analysis of H13N8and H9N2subtype avian influenza virus showed that two subtype viruses are noval genetype viruses.The positive rate of different month, sample site, and different sample type were different, and the three factors all can affect the positive rate.