H5N6 Avian Influenza Virus Isolation,Phylogenetic Analysis In Nanchong Sichuan Province,Nanchang Jiangxi Province And Using High-Throughout Sequencing To Explore The Molecular Of Evolution H10N8 Avian Influenza Viruses Adapt Mouse-lung

Avian influenza is caused by type A influenza virus of avian infectious diseases,according to the virion surface haemagglutinin(HA)and neuraminidase(NA)to distinguish different Asian type;according to the virulence can be divided into highly pathogenic and low pathogenic.Since December 2003,H5N1 subtype avian influenza virus epidemic outbreak began in Hong Kong,the avian influenza virus growing endangerment to our poultry industry,and to the health of our people caused a great threat.In Sichuan Province and Jiangxi Province we investigate influenza a virus in live poultry markets in 2014,11 avian influenza A H5N6 virus were isolated from live poultry markets(LPMs),including a poultry drinking water source,a pigeon source,three chicken source,and six ducks source.To study of these 11 H5N6 avian influenza virus genetic and phylogenetic,we analysis their molecular characteristics.The results show that 11 HA gene of the virus were novel reassortants between H5N1 clade2.3.4 and H6N6 virus,Jiangxi and Sichuan samples had evolved into two different distinct lineages.Meanwhile,the human H5N6 virus was closely related to the avian-source virus of Sichuan lineage;not only includes the T160A substitution in the haemagglutinin protein,but also an 11 amino acids deletion in the neuraminidase stalk;which may aid in enhancing viral affinity for human-like receptors and virulence mammals.After the occurrence of the H5N6 infection in human cases,once again reminded of the potential threat to public health of people H5 subtype for a pandemic,but also illustrates the importance of long-term surveillance for live poultry epidemiology of avian influenza investigation.In this study,an H10N8 subtypes of avian influenza virus(A/water/Dongting Lake/Hunan/3-9/2007(H10N8))in Mouse-lung adapt strains P0,P2,P4 and P6.for the study,the virus lethal in mice showed an increase in the phenomenon in Mouse-lung passaging.Used the genome sequencing were performed by high-throughput sequencing(Illumina)to analysis the rat lung-passaged virus primary(P0),the second generation(P2),the fourth generations(P4),sixth generation(P6)were sequenced.Hoping to understand the virus genome change during the adaptation in mice.The Illumina part Received a total clean reads 1784920(PO),1726484(P2),1931171(P4),2484253(P6).Effectively Influenza reads accounted for 82.33%of clean reads.The 3730 sequencing data and Illumina sequencing results with a high degree of consistency,and used the results of Illumina sequencing to analysis the mutation of the H10N8 genome in each passage.The results of visible changes in the influenza virus genome key amino acid sites with death rate in mice showed a direct correlated.especially in the PB2 gene 627 glutamate(E)to lysine(K)mutation makes the P4 generation increased pathogenicity in mice(in multiple organs virus is detected),and before the P0,P2 generation on the same sites emerged entropy corresponding rise.Similarly,other sites such as the PA gene 653 leucine(L)to proline(P)and 277 phenylalanine(P)to serine(S)step changes the amino acid,reveals that H10N8 avian influenza virus in mice gradual process of adaptation.The results provides an important reference.For the understanding of how viruses evolve to adapt to the host and enhances virulence mechanism.