Reverse Genetics of Influenza B Viruses

Abstract

Annual influenza epidemics are caused not only by influenza A viruses but also by influenza B viruses. Initially established for the generation of recombinant influenza A viruses, plasmid-based reverse genetics techniques have allowed researchers the generation of wild type and mutant viruses from full-length cDNA copies of the influenza viral genome. These reverse genetics approaches have allowed researchers to answer important questions on the biology of influenza viruses by genetically engineering infectious recombinant viruses. This has resulted in a better understanding of the molecular biology of influenza viruses, including both viral and host factors required for genome replication and transcription. With the ability to generate recombinant viruses containing specific mutations in the viral genome, these reverse genetics tools have also allowed the identification of viral and host factors involved in influenza pathogenesis, transmissibility, host-range interactions and restrictions, and virulence. Likewise, reverse genetics techniques have been used for the implementation of inactivated or live-attenuated influenza vaccines and the identification of anti-influenza drugs and their mechanism of antiviral activity. In 2002, these reverse genetics approaches allowed also the recovery of recombinant influenza B viruses entirely from plasmid DNA. In this chapter we describe the cloning of influenza B/Brisbane/60/2008 viral RNAs into the ambisense pDP-2002 plasmid and the experimental procedures for the successful generation of recombinant influenza B viruses.

Broad and Protective Influenza B Virus Neuraminidase Antibodies in Humans after Vaccination and their Clonal Persistence as Plasma Cells

“…In order to determine if the inhibitory activity of the IBV NA hMAbs was specific for IBV of the Victoria lineage or if they also have the ability to inhibit IBV of the Yamagata lineage, we used a plasmid-based reverse genetic system (43, 47) to generate a recombinant mCherry-expressing virus containing the six internal genes (PB2, PB1, PA, NP, M and NS-mCherry) from B/Brisbane/60/2008 virus and the HA and NA from influenza B/Yamagata/16/1988 virus, a representative member of the Yamagata lineage. Then, the ability of the six IBV NA hMAbs to inhibit infection was evaluated using our fluorescence-based microneutralization assay as described above.…”

“…Recombinant influenza virus A/California/4_NYICE_E3/2009 (pH1N1), B/Yamagata/16/1988, and B/Brisbane/60/2008 WT or mCherry-expressing viruses have been previously described (43, 56, 57). A reassortant IBV containing the six internal genes (PB2, PB1, PA, NP, M, and NS-mCherry) from B/Brisbane/60/2008 virus and the HA and NA from B/Yamagata/16/1988 (reB/Yamagata/16/1988 mCherry) virus was generated using plasmid-based reverse genetics techniques (43, 47). Two recombinant B/Brisbane/60/2008 virus strains containing amino acid substitution E117A or H273Y (E119A or H274Y [N2 numbering]) (6, 48) in the NA were generated using plasmid-based reverse genetics techniques (43, 47).…”

“…A reassortant IBV containing the six internal genes (PB2, PB1, PA, NP, M, and NS-mCherry) from B/Brisbane/60/2008 virus and the HA and NA from B/Yamagata/16/1988 (reB/Yamagata/16/1988 mCherry) virus was generated using plasmid-based reverse genetics techniques (43, 47). Two recombinant B/Brisbane/60/2008 virus strains containing amino acid substitution E117A or H273Y (E119A or H274Y [N2 numbering]) (6, 48) in the NA were generated using plasmid-based reverse genetics techniques (43, 47). IBV B/Malaysia/2506/2004 (NR-9723), B/Ohio/01/2005 (NR-41801), B/Nevada/03/2011 (NR-44023), B/Sydney/507/2006 (NR-36526), B/Texas/06/2011 (NR-44024), and B/Lee/1940 (NR-3178) were obtained from BEI Resources, and the IBV B/Wisconsin/01/2010 (FR-806) was obtained from International Reagent Resources (IRR).…”

Functional Characterization and Direct Comparison of Influenza A, B, C, and D NS1 Proteins in vitro and in vivo

“…four types of influenza viruses: IAV, IBV, ICV and the newly identified IDV (Wright et al, 2007;Chen and Holmes, 2008;Wanitchang et al, 2012;Tong et al, 2013;Baker et al, 2014;Yoon et al, 2014;Hengrung et al, 2015;Matsuzaki et al, 2016;Wang et al, 2016;Foni et al, 2017;Nogales et al, 2017c;Su et al, 2017;Nakatsu et al, 2018;Asha and Kumar, 2019;Zhang et al, 2019). Despite the divergence of their genomes and host range, all types of influenza viruses express NS1 proteins.…”

“…Influenza viruses belong to the Orthomyxoviridae family and are enveloped viruses which contain a segmented genome of singlestranded RNA molecules of negative polarity (Wright et al, 2007;Nogales and Martinez-Sobrido, 2016;Martinez-Sobrido et al, 2018;Blanco-Lobo et al, 2019). Currently, there are four recognized influenza virus types: A, B, C, and D (IAV, IBV, ICV, and IDV, respectively) (Wright et al, 2007;Chen and Holmes, 2008;Wanitchang et al, 2012;Tong et al, 2013;Baker et al, 2014;Yoon et al, 2014;Hengrung et al, 2015;Matsuzaki et al, 2016;Wang et al, 2016;Foni et al, 2017;Nogales et al, 2017c;Su et al, 2017;Nakatsu et al, 2018;Asha and Kumar, 2019;Zhang et al, 2019). IAV and IBV contain eight genomic viral (v)RNA segments (Wright et al, 2007), and two major glycoproteins in the virion surface, the hemagglutinin (HA) and neuraminidase (NA), which are responsible for viral binding and release, respectively, of the virus from infected cells (Wright et al, 2007).…”