| Twenty-five H9N2 subtype avian influenza viruses (AFVs) isolated from the Eurasian continent were sequenced and molecular evolutionarily analyzed in this study. These viruses from 1995 to 2000 isolated from different regions,including the Northeast,North,Middle,South,Southwest and Northwest of China,Korea in Eastern Asia,Pakistan in Southern Asia. Iran and Saudi Arabia in the Middle East,Ireland and Germany in Europe,and South Africa They were isolated from various types of avian. in relation to chickens,ducks,ostriches and pheasants.The nucleic acids of the all influenza viruses conducted in the research were extracted from the viruses propagated in specific-pathogen-free chicken embryos. All of the eight segments were amplified by RT-PCR,and the purified PCR products were done cycle sequencing with specific primers,furthermore,the sequencing products were purified and run gel on ABI Prism 377 DNA sequencing machine. The specific primers of the eight genes for PCR and cycle sequencing were designed using the OHgo (4.0 version) and Genedoc (2.3 version) software. Finally,the WISCONSIN Sequence Analysis Package (GCG,10.2 version),Phylogenetic Analysis Using Parsimony (PAUP,4.0 version) and Treeview (1.5 version) software were used for data analysis,sequence translation,phylogenetic tree drawing and molecular evolution analysis.The phylogenetic trees revealed that all of the genes of the H9N2 subtype AFVs conducted in this study belonged to Eurasian avian lineage,in which there were Y280-like,Gl-like and Y439-like sublineages in the HA,PB1,PA,MR,M and NS genes,Y280-like,G1 -like,Y439-like and G9-like sublineages in the NA gene,and Y280-like,GI-like,Y439-like and KOR-like sublineages in the PB2 gene,respectively. And the sublineages could differentiate into several clades,even subclades.The result of this investigation displayed that the H9N2 subtype human isolates constituted Gl-HK clade in Gl-like sublineage with Gl-like AFVs isolated from Hong Kong together,and this kind of viruses,with possible abilities to infect human and induce human influenza pandemic,were not found in the mainland of China The viruses from China Mainland belonged to Y280-like sublineage,except some individual genes of CK/SC/5/97,DK/NJ/2797 and CK/HN/62/00 were situated in Gl-like sublineage,and only CK/SZ79/97 isolated from Shenzhen was classical G9-like virus which prevailed in the live poultry market in Hong Kongin 1997,so this virus might come from Hong Kong Specific Administrative Region of China. The viruses isolated from the mainland of China developed from the earlier viruses circulating in China,and they were related to each other closely and evolved slowly.This research illustrated that CK/PA/4/99 and CK/PA/5/99 isolated from Pakistan were located in Gl-PA clade in Gl-like sublineage,which had high homology with Gl-HK clade,and the phylogenetic trees suggested that Gl-PA clade involved into the identical sublineage earlier than Gl-HK cladeb obviously,so Gl-PA viruses were possible origin of Gl-HK viruses. CK/IR/1 IT/99 and CK/SAR/532/99 were reassortants of Gl-like and Y439-Iike viruses. DK/GE/1 13/95,CK/GE/R35/98,PH/EE/PV18/97 and OS/SAF/9508 103/95 were part of Y439-like sublineage in principle,except the NS genes of PH/E/PV 18/97 and DK/GE/1 13/95 belonged to B group and Eurasian avian lineage of A group,respectively. Only CK/KOR/99029/99 was classical KOR-like virus.In conclusion,this study elucidated the molecular evolution of H9N2 AIVs isolated from the Eurasian continent,clarified the category of the recent epidemical strains of H9N2 AFVs circulating in the mainland of China,and made a foundation for the further investigation. The possible source of Gl-like viruses was proposed in this thesis,which provided the theory-support for the preventive research on the infection to human being from H9N2 and H5N1 subtype AFVs.This investigation was supported by the national key basic research and development grants (973) G199901 1902 and 199901 1905.Ph.D. candidate:LI JianweiSpecialty:Preventive veterina...
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