Characterization Of H5N1HPAIVs In China In 2005～2007

Since late 2003, H5N1 highly pathogenic avian influenza (HPAI) has spread in an unprecedented manner in southeastern Asia. The outbreak occurred occasionally in China since the first case reported in 26th Jan in 2004. In May 2005, an infection occurred among wild birds in Qinghai lake in China, henceforth, the viruses were spread to Europe and Africa through the migration of wild birds. In fact, the viruses continue to circulate among poultry in Asia and remain a threat to both veterinary and human public health. The cumulative number of confirmed human cases of avian influenza A/(H5N1) reported to WHO were 240 deaths of 384 infected on 17 April 2008.The present study performed extensively characterization of H5N1 HPAIVs circulated in 2005~2007 in China, which correlative to cases of poultry and human. The phylogenic tree of 8 segments showed that the viruses evoluted from the reassortant of wild birds viruses with GS/GD/1/96, Phylogenitically, the viruses established 4 clades, which were clade ST(soutern China,ST), isolates mainly circulating in Hunan, Hubei, Anhui and Fujian provinces; clade XJ(Xinjiang, XJ), isolates from Xinjiang province, clade SX(Shanxi, SX), viruses prevailed in Shanxi and Ningxia provinces; clade WB(Wild birds, WB), formed by the viruses from Liaoning provinces with wild birds isolates in Qinghai and Tibet. Different clades possess their own property of genome. SX clade acquired the resistance to amantadine and antigenic-drifted. Four clades represent 11 genotypes, 3 genetypes in clade ST, 2 genetypes in clade XJ, clade SX possess 2 genetypes, 4 genotypes in clade WB. Clade ST was found to be closely related to that of the virus from other countries in Asia. 3 human viruses were origin from avian influenza virus for their genetic and phylogenetic property same to clade ST.Cross HA-HI test were performed to identify the antigenic-drift of the heterogeneous viruses, the index of antigenic-drift correlation showed that clade SX drifted from the remained clades. Experimental infection of chickens and ducks to evaluate the pathogenicity of viruses from 4 clades in poultry, all the viruses were highly pathogenic avian influenza viruses for IVPI≥1.2. Different clades exhibited diverse pathogenicity in ducks. Clades ST,XJ and WB can replicate efficiently in ducks and produce clinical symptoms, whereas clade SX producing no evidence of disease.Classical mammalian models BALB/C mice infected with viruses to test the pathogenicity The results revealed that the virus acquired the ability to replicate in mice without prior adaption. 4 clades showed a big diversity of pathogenicty in mice, clade ST showed high, moderate and low pathgenicity in mice, clade WB was lethal, and that of clade XJ and SX were low.To trace the origin of 3 H5N1 HPAIV isolated from the washes of the eggs smuggled to China by Vietnamese in 2005, 8 segments were compared with the reference sequences in Genbank, the largest homology with HK/156/97 and also at the same linage in the phylogenic tree demonstrated that 3 Vietnam viruses were HK/97-like virus. They are highly pathogenic avian influenza virus in chickens, but not lethal to ducks. They replicated in mice with a limited tissue tropism.We also present the characterization of a H7N2 avian influenza virus CK/HeB/1/02 the only one H7 subtype virus found in China presently. The analysis of genetic and phylogenitic revealed that CK/HeB/1/02 may have been introduced in chickens from wild birds. Both IVPI and the amino acids in the cleavage site demonstrated the virus was low pathogenic. The data of virus shedding from oropharyngeal and cloacal and replication in tissues post administrated intravenously and intranasal showed that CK/HeB/1/02 has not adapted in chickens. The virus replicated in mice with a wider organ tropism without preadaptation.In summary, the evolution and biological characterization of H5N1 HPAIV prevailing in China in 2005 ~ 2007 were presented. The virus evoluted from the reassortment of wild birds virus with GS/GD/1/96 according to the analysis of genetic and phylogenitic. The viruses established 4 clades with geographic relativeness. Different genotypes could be found in each clade and possessed distinctive characteristic in genome. We demonstrated that clade SX acquired the resistant to amantadine and antigenic-drifted. Different clades exhibited a big pathogenic diversity in mice. The origin of 3 human viruses were traced from avian influenza. The evolution of 3 H5N1 HPAIVs from Vietnam illustrated that HK/97-like virus still exist naturally. We also present the characterization of the only one H7N2 subtype avian influenza virus in China, which was low pathogenicity in chicken, duck and mammalian. The study would provide basic data and information for avian influenza virus research, especially research on H5N1 HPAIV in China.