Epidemic Investigation Of Influenza Virus In Poultry Markets In Yueyang,Hunan Province From 2010 To 2012

Epidemics of avian influenza not only cause substantial economic losses to the poultry business,but also pose a serious threat to public health.In China,the highly pathogenic avian influenza(HPAI)H5N1 viruses have undergone extensive genetic recombination with viruses from different sources and have produced numerous H5N1 genotypes.The domestic ducks from South China,as an important mixer for influenza viruses,greatly facilitate the genetic and antigenic changes of the viruses.Poultry markets have long been considered a potential bird flu "hotbed",as it is easy for the avian viruses to have gene reassortments and multiple subtypes have been detected in the markets.Thus the surveillance of influenza in poultry markets is vitally important for prevention and control of avian influenza viruses.During the epidemic investigation of avian influenza from poultry markets in Yueyang,Hunan province from 2010 to 2012,seventy-three strains of influenza viruses were isolated from the live poultry markets.Among them,there are one strain of H5N1/H9N2 mixed-subtype,one strain of H9N2 subtype and four strains of H5N1 subtype from chickens,three strains of H5N1/H9N2 mixed-subtype,three strains of H5N1/H6N6 mixed-subtype,one strain of H9N2 and 59 strains of H5N1 from ducks,and one strain of H5N1 from a pigeon.We chose one strain of H5N1/H6N6 mixed-subtype,which was isolated from a duck in December 2011,for further analysis.Its whole genome was sequenced using next generation sequencing(NGS)(Illumina)and 10 segments were obtained,includingPB2,PB1,PA,H5,H6,NP,N1,N6,M and NS.For it contained 2 HA subtypes(H5 and H6)and 2 NA subtypes(N1 and N6),it was denominated as A/duck/Hunan/747/2011(mixed).Phylogenetic analysis indicated that the mixed-subtype virus H5 HA gene belonged to clade 2.3.2.1 and H6 HA belonged to the group ? lineage,but that the segments coding for inner proteins had more or less recombination.To evaluate the replication capacity and virulence of the mixed-subtype virus,in vivo studies were performed.The growth curve inchick embryo reflected good replication capacity of the mixed-subtype.When mice were inoculated with the virus,they all died.Residual virus could be detected in lungs,brains,spleens and kidneys when the.4 organs of the infected mice were examined.The results indicated that the mixed-subtype had strong replication and high virulence in mice.