ZIKA Virus Amplification And The Viral-induced Immune Response In Mice

ZIKA virus(ZIKV)is an enveloped virus with a positive sense single stranded RNA genome,which belongs to a member of the Flavivirus genus of the Flaviviridae family.ZIKV infection tends to cause Guillain-Barre syndrome and neonatal congenital microcephaly.Due to lack of ideal animal infection models,the virological characteristics of ZIKV infection,the immune response induced by infection and the pathogenic mechanisms have not been studied in detail.this paper observed and analyzed the replication characteristics of the Asian epidemic strain ZIKV/H/PF/2013in vitro.ZIKV amplification and the viral-induced immune response in mice were further analyzed through footpad injection.Firstly,the characteristic of ZIKV amplification in vitro was analyzed.C6/36 cell line was used to amplify virus,the titer is up to 5×10~7-10~8 PFU/ml.Then the duplication curve of ZIKV was analyzed in both BHK-21 and A549 cell lines.The result showed that ZIKV amplified linearly from 12 to 48 hours post infection and the virus production was peaked at 48 hours post infection.Four siRNAs were designed based on ZIKV's genome and their antiviral activity was tested in the A549 cell line.The result showed that the four specific siRNAs had certain antiviral activity compared to the control siRNA,in which the No.3 siRNA had the strongest antiviral activity and the effect could reach 60%at 6.25 nM.Secondly,to know the characteristic of ZIKV amplification in mice,the amplification of ZIKV from local to system,the body weight changes,survival rate,tissue tropism,and histopathology were analyzed systemically in type I and II IFN receptor deficient AG6 mice and the wild type control C57BL/6 mice by footpad injection.The results showed that ZIKV could cause fatal infections in AG6 mice but not establish systemic infection in wild-type mice;ZIKV has broad tissue tropism which can infect the heart,liver,spleen,lung,kidney,brain and other tissues of mice;ZIKV also caused significant histopathological changes in the infected organs.Thirdly,the expression of the viral envelope protein(ZIKV-E),composition and activation of major immune cell subsets in mouse footpads,axillary lymph nodes,peripheral blood and spleen were analyzed by fluorescence flow cytometry.The results showed that in ZIKV-infected AG6 mice,a variety of immune cell subsets including neutrophils were positive for ZIKV-E protein and significant neutrophil were recruited.Moreover,ZIKV infection induced DC activation in peripheral blood,CD4+and CD8+T cells activation in peripheral blood and spleen while inhibited DC activation in the draining lymph nodes.In summary,this study analyzed the characteristics of ZIKV amplification in mice deeply.To our knowledge,it's the first time to find that ZIKV can infect a variety of immune cell subpopulations including neutrophils and induce siginificant neutrophils recruitment.This study provides a foundation for further study on the spreading and pathogenic mechanism of ZIKV in vivo.