The Seperation Of H9N2Subtype Aiv And The Variation Analysis Of HA Gene

H9N2subtype avian influenza is popular throughout the world, is one of thediseases which have serious economic losses in poultry farm. This subtype originallyonly spreaded in poultry, and later it was discovered that it can also infect mammalsdirectly, and have an important public health significance. In order to understand thevariation pattern of H9N2subtype AIV, we separated H9N2subtype AIV, cloned theHA gene and analysised the gene sequence and genetic evolution in this study,obtaining the following results:1. Collection samples from the clinical suspected flu poultry farm in Luoyang ofHenan province and Xianyang of Shaanxi province, we had separated five strainsfrom the chickens by embryonated eggs inoculation and passaged, HA test anddetection of RT-PCR method, which named LY35, LY36, LY48, LY49, XY.2. We compounded two pairs of primers according to the reference of HA genesequences in GenBank, amplified HA gene of LY36, XY strains by RT-PCR method.After cloning, sequencing and splicing, the two whole sequenses of HA gene wereobtained and submited to GenBank, which accession number were KF638574,KF638575, respectively.3. The HA nucleotide gene sequense of LY36, XY and reference strains fromGenBank nucleotide sequence editing, analysis, amino acid sequence prediction andalignment were performed using DNAStar software. Phylogenetic trees wereconstructed using MEGA.6software by the neighbor-joining method.The resultsshowed that: the homology of nucleotide sequenses between two viruses were97.1%,belonged to BJ/94branch (Y280subbranch). The amino acid sequence of the HA genecleavage sites of the two strains were PSRSSR/GL, which showed that they had thepropertie of low pathogenic influenza virus. Both the two strains had a mutation ofglutamine to leucine in234amino acid site and were added a glycosylation sites in313amino acid site, which would affect the virulence of the virus and change the range of hosts.In conclusion, LY36, XY H9N2subtype AIV isolates in our laboratory are themajor epidemic strain in recent years in our country, but they have a certain degree ofvariation at the genetic level.