Development and characterization of a reverse-genetics system for influenza D virus

Jieshi Yu, Runxia Liu, Bin Zhou, Tsui wen Chou, Elodie Ghedin, Zizhang Sheng, Rongyuan Gao, Shao lun Zhai, Dan Wang, Feng Lia

Research output: Contribution to journalArticlepeer-review

16 Scopus citations

Abstract

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-plasmid-based 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.

Original languageEnglish
Article numbere01186-19
JournalJournal of Virology
Volume93
Issue number21
DOIs
StatePublished - 2019

Bibliographical note

Publisher Copyright:
Copyright © 2019 American Society for Microbiology. All Rights Reserved.

Funding

This study was supported by NIH R01AI141889, SDSU-AES 3AH-673, USDA/NIFA 2016-67016-24949, National Science Foundation/EPSCoR (http://www.nsf.gov/od/iia/programs/epscor/index.jsp) award IIA-1335423, and the SD-CBRC supported by the State of South Dakota’s Governor’s Office of Economic Development. The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention. This study was supported by NIH R01AI141889, SDSU-AES 3AH-673, USDA/NIFA 2016-67016-24949, National Science Foundation/EPSCoR (http://www.nsf.gov/od/iia/ programs/epscor/index.jsp) award IIA-1335423, and the SD-CBRC supported by the State of South Dakota’s Governor’s Office of Economic Development. The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention.

FundersFunder number
SD-CBRC
National Science Foundation Arctic Social Science Program
National Institutes of Health (NIH)R01AI141889, USDA/NIFA 2016-67016-24949, SDSU-AES 3AH-673
National Institutes of Health (NIH)
Centers for Disease Control and Prevention
Office of Experimental Program to Stimulate Competitive ResearchIIA-1335423
Office of Experimental Program to Stimulate Competitive Research
South Dakota Governor's Office of Economic Development

    Keywords

    • Influenza D virus
    • Reverse-genetics system

    ASJC Scopus subject areas

    • Microbiology
    • Immunology
    • Insect Science
    • Virology

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