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Development and Characterization of a Reverse-Genetics System for Influenza D Virus

文献类型： 外文期刊

作者： Yu, Jieshi 1 ; Liu, Runxia 1 ; Zhou, Bin 2 ; Chou, Tsui-wen 3 ; Ghedin, Elodie 3 ; Sheng, Zizhang 4 ; Gao, Rongyuan 1 ; Z 1 ;

作者机构： 1.South Dakota State Univ, Dept Biol & Microbiol, Brookings, SD 57007 USA

2.Ctr Dis Control & Prevent, Influenza Div, Natl Ctr Immunizat & Resp Dis, Atlanta, GA USA

3.NYU, Dept Biol, Ctr Genom & Syst Biol, New York, NY 10003 USA

4.Columbia Univ, Zuckerman Mind Brian Behav Inst, New York, NY USA

5.Guangdong Acad Agr Sci, Anim Dis Diagnost Ctr, Inst Anim Hlth, Guangdong Key Lab Anim Dis Prevent,Guangdong Open, Guangzhou, Guangdong, Peoples R China

6.BioSNTR, Brookings, SD 57007 USA

7.SD CBRC, Brookings, SD 57007 USA

关键词： influenza D virus; reverse-genetics system

期刊名称：JOURNAL OF VIROLOGY （ 影响因子：5.103； 五年影响因子：5.078 ）

ISSN： 0022-538X

年卷期： 2019 年 93 卷 21 期

页码：

收录情况： SCI

摘要： Influenza D virus (IDV) of the Orthomyxoviridae family has a wide host range and a broad geographical distribution. Recent IDV outbreaks in swine along with serological and genetic evidence of IDV infection in humans have raised concerns regarding the zoonotic potential of this virus. To better study IDV at the molecular level, a reverse-genetics system (RGS) is urgently needed, but to date, no RGS had been described for IDV. In this study, we rescued the recombinant influenza D/swine/Oklahoma/1314/2011 (D/OK) virus by using a bidirectional seven-plasmidbased system and further characterized rescued viruses in terms of growth kinetics, replication stability, and receptor-binding capacity. Our results collectively demonstrated that RGS-derived viruses resembled the parental viruses for these properties, thereby supporting the utility of this RGS to study IDV infection biology. In addition, we developed an IDV minigenome replication assay and identified the E697K mutation in PB1 and the L462F mutation in PB2 that directly affected the activity of the IDV ribonucleoprotein (RNP) complex, resulting in either attenuated or replication-incompetent viruses. Finally, by using the minigenome replication assay, we demonstrated that a single nucleotide polymorphism at position 5 of the 3' conserved noncoding region in IDV and influenza C virus (ICV) resulted in the inefficient cross-recognition of the heterotypic promoter by the viral RNP complex. In conclusion, we successfully developed a minigenome replication assay and a robust reverse-genetics system that can be used to further study replication, tropism, and pathogenesis of IDV. IMPORTANCE Influenza D virus (IDV) is a new type of influenza virus that uses cattle as the primary reservoir and infects multiple agricultural animals. Increased outbreaks in pigs and serological and genetic evidence of human infection have raised concerns about potential IDV adaptation in humans. Here, we have developed a plasmid-based IDV reverse-genetics system that can generate infectious viruses with replication kinetics similar to those of wild-type viruses following transfection of cultured cells. Further characterization demonstrated that viruses rescued from the described RGS resembled the parental viruses in biological and receptor-binding properties. We also developed and validated an IDV minireplicon reporter system that specifically measures viral RNA polymerase activity. In summary, the reverse-genetics system and minireplicon reporter assay described in this study should be of value in identifying viral determinants of cross-species transmission and pathogenicity of novel influenza D viruses.

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