Comparative Study Of Two Different Strains Of H7N9 Virus Against Host Innate Immunity

Since the H7N9 influenza virus was reported in 2013,it has caused huge economic losses to the poultry industry.The virus could break through the inter-species barrier and infect humans,causing infection and death.Public health security faced a huge threat.The host's innate immune system plays an important role in the process of combating viral infection.The comparative study of the difference in the process that virus against host innate immunity can lead to a deeper understanding of the interaction between the virus and the host,which could provide new ideas and scientific basis for the development of prevention and control measures.In order to have a further understanding of the difference that H7N9 virus against host innate immunity,two different isolates of H7N9 influenza virus were selected as model viruses,which were avian virus A/Pigeon/Shanghai/S1421/2013(H7N9)[PG/S1421] and human virus A/Anhui/1/2013(H7N9)[AH/1].These two viruses only had one amino acid difference at the 627 position in the PB2 protein,PG/S1421 was 627 glutamic acid(E),while AH/1 was lysine(K),but showed a great difference in pathogenicity to mammals.C57BL/6 mice were infected intranasally with different doses of two influenza viruses,and the body weights and mortality were continuously recorded.The results showed that AH/1 could cause disease and death in mice,while PG/S1421 could not cause disease in mice.There was a difference in the degree of pathological damage in the lungs of mice intranasally infected with the viruses.The level of type I interferon in the lung was detected by ELISA assay and the results showed that both viruses could induce the production of type I interferon in the lung,but AH/1 induced more amount of type I interferon.In order to determine the main pathways for the induction of type I interferon by two strains of influenza virus,mice lacking the key molecules in the type I interferon signaling pathways were used to collect the peritoneal macrophages,then the cells were infected with the viruses,and the production of type I interferon was detected.The results showed that the two strains of influenza virus infection mainly produced type I interferon through the RIG-I/MAVS/IRF3 signal pathway.To further investigate the causes for the different production of type I interferon induced by PG/S1421 and AH/1,firstly,two constructed A549 cell lines expressing PB2 protein of PG/S1421 and AH/1 were infected with two viruses,detecting the production of IFN-?.The mRNA relative expression results showed no significant difference in the production of type I interferon induced by the two viruses,indicating that the single amino acid difference at position 627 of PB2 is not the main cause that AH/1 induction of more type I interferon.Then,two strains of virus were used to infect A549 cells and mouse peritoneal macrophages at the same dose.Indirect immunofluorescence and flow cytometry assay showed no significant difference in infection efficiency,indicating that the difference in type I interferon production was not due to infection efficiency.Finally,the mouse peritoneal macrophages were infected with two strains of influenza virus,and the supernatant and cell protein were collected.The contents of IFN-? in the supernatant were detected by ELISA.The expression of key molecules in the interferon production pathways were detected by western blotting assay.It was found that mouse peritoneal macrophages infected with the two viruses could both produced type I interferon,but AH/1 induced more and AH/1 infection up-regulated RIG-I expression,while PG/S1421 infection had no such significant effect on the expression level of RIG-I.The results of this study indicated that avian virus PG/S1421 infection did not cause disease in mice,human virus AH/1 infection could cause disease and death in mice.Both viruses could induce type I interferon production,however a significantly difference was existed in the aspects against host innate immunity.AH/1 infection could promote the production of type I interferon by up-regulating the expression of RIG-I,and a higher viral titer in the host suggested that AH/1 might escape the host's innate immunity.This study reveals the primary mechanism that human influenza virus promotes the production of type I interferon,providing a new idea for the prevention and control of influenza.