Phylogenetic And Molecular Characterization Of H6 Subtype Avian Influenza Virus In Wild Birds In Shanghai, 2016-2017

The H6 subtype avian influenza virus(AIV),belonging to the LPAIV,was first isolated from a turkey in Massachusetts in 1965 and is now widely distributed globally.H6 subtype AIVs have a broader host range and can be easily isolated from migratory waterfowl,as well as domestic aquatic and mammals.In addition to providing internal gene fragments for highly pathogenic avian influenza viruses such as H5N6,a number of reports have also proved that H6 subtype AIVs can break through species limitations to infect humans and threaten human health.Therefore,in recent years,people pay more attention to H6 subtype AIVs.As the natural host of avian influenza virus,wild birds play a vital role in virus reassortment and transmission,besides,Shanghai is a crucial stopover or wintering site for migratory wild birds along the East Asian-Australasian Flyway.Considering the widespread prevalence of H6 subtype AIVs in China and the danger posed to human health by the continuously participation of H6 subtype AIVs in the recombinatio of highly pathogenic AIVs,we want to understand the situation of H6 subtype AIVs carried by wild birds in Shanghai.A total of 1514 cloacal and tracheal swab samples were collected from Jiuduansha Natural Reservation Zone and Nanhui Dongtan wetland of Pudong,Shanghai.30 samples were H6 subtype AIVs,and 8 strains were selected for further detection and analysis.Four H6 subtype AIVs(H6N1,H6N2,H6N5,and H6N8)were found to be prevalent.The results of homology analysis showed that the 8 H6 strains we isolated shared high homology with different sublineages isolated from domestic ducks or wild birds in different countries along the East Asian-Australasian Flyway route,and on the basis of the source of the genes,we divided the eight strains into seven genotypes,which highlighted wild birds' ability to spread AIVs and encourage the evolution and emergence of new genotypes.The results of cell experiment showed that the eight H6 subtype strains can proliferate in MDCK,A549 and MLE–12 cells.Receptor binding analysis showed that these viruses preferentially bound to avian receptors and did not have the ability to spread in the crowd.However,the possibility that the H6 subtype AIVs recombinating with highly pathogenic avian influenza virus can't be excluded.Continued surveillance of H6 subtype AIVs in wild birds in eastern China is important to understand their transmission and evolution better.