| Avian influenza virus is one of the major infectious diseases in poultry of China and sometimes it could infect human.Avian influenza causes huge losses in the poultry industry each year.In addition,poultry is a host and susceptible animal of avian influenza virus,but how it interacts with avian influenza virus and which host antiviral proteins participate in this interaction are still unclear.Thus,we analyzed the genetic characteristics,pathogenicity of H5N6 avian influenza viruses in South China,2015-2016,and the expression of antiviral factors such as TRIM protein in chicken infected the viruses.The results showed that the HA genes of 8 H5N6 avian influenza viruses had five continuous basic amino acids at the cleavage site（-RRRKR↓G-）,which is a characteristic of highly pathogenic AIVs.Their NA genes had an 11-amino acid deletion of residues58-68.In the PB2 genes,their amino acids of the 8 viruses at 627 site were E,which was the characteristic of AIVs from birds.In the M1 genes,the amino acid at position 30 and215 were aspartic acid（D）and alanine（A）,respectively.Phylogenetic analysis showed that the HA genes of the 8 viruses belonged to clade 2.3.4.4.Their HA,PB2,PA,M,NP and NS genes were classified into MIX-like group and the NA genes of them fell into Eurasian-lineage viruses.The PB1 genes of GS140,GS142,CK146,CK152,CK274 and GS474 viruses were classified into VN 2014-like and the others belonged to MIX-like,which indicated that the 8 viruses belonged to different genotypes,and some of these viruses occurred recombination with different genotypes.To observe the pathogenicity and transmission of these H5N6 viruses in chickens,chickens were inoculated intranasally with 105 EID500 of the CK146 and GS474 viruses in a0.1 ml volume,respectively.The resluts showed that the mean time of death in the CK146and GS474 inoculated groups were 3 and 3.38 days,respectively.In chickens inoculated with the two viruses,the virus replicated in all tested tissues.The mean time of death in the CK146 and GS474 contact groups were 4.25 and 6.2 days,respectively.In all contact chickens,the virus also replicated in all tested tissues.Overall,these H5N6 viruses from clade 2.3.4.4 had high pathogenicity and could be shed by the digestive tract and respiratory tract in all chickens.However,they had different transmissibility in chickens.To understand the host antiviral factors of chicken infected H5N6 avian influenza virus,we measured the expression of antiviral factors in chicken lungs infected with CK146virus.The results showed that the expression of host antiviral proteins（TLR3,MDA5,MAVS,MX1,OAS,TRIM13,TRIM25）and cytokines（IL-6,IFN-β）increased.Then,we cloned and identified the chicken TRIM25 protein.The results of overexpressed chTRIM25revealed that the expression of TLR-3,MDA5,MX1,OAS and IFN-βwas significantly upregulated,which indicatd chTRIM25 could promote type I interferon-mediated antiviral immune response.8 H5N6 avian influenza viruses in our study were highly pathogenic avian influenza viruses,and some of them occurred recombination with different genotypes.CK146 and GS474 viruses had high pathogenicity and transmissibility in chickens,and they could be completely lethal in chickens upon inoculation and naive contact.The expression levels of MDA5,MAVS,TRIM13,TRIM25 and IFN-βwere all upregulated in chicken and CEF infected H5N6 avian influenza virus.We cloned and identified the chicken TRIM25 protein,and we found that chTRIM25 could promote type I interferon-mediated antiviral immune response.Therefore,our study provided experimental data for understanding the genetic characteristics of the H5N6 avian influenza virus and the host antiviral immune response of chickens. |
