The Development and Use of Reporter Influenza B Viruses

Abstract

Influenza B viruses (IBVs) are major contributors to total human influenza disease, responsible for ~1/3 of all infections. These viruses, however, are relatively less studied than the related influenza A viruses (IAVs). While it has historically been assumed that the viral biology and mechanisms of pathogenesis for all influenza viruses were highly similar, studies have shown that IBVs possess unique characteristics. Relative to IAV, IBV encodes distinct viral proteins, displays a different mutational rate, has unique patterns of tropism, and elicits different immune responses. More work is therefore required to define the mechanisms of IBV pathogenesis. One valuable approach to characterize mechanisms of microbial disease is the use of genetically modified pathogens that harbor exogenous reporter genes. Over the last few years, IBV reporter viruses have been developed and used to provide new insights into the host response to infection, viral spread, and the testing of antiviral therapeutics. In this review, we will highlight the history and study of IBVs with particular emphasis on the use of genetically modified viruses and discuss some remaining gaps in knowledge that can be addressed using reporter expressing IBVs.

Keywords: Influenza B virus; molecular virology; reporter virus; reverse genetics; viral genetic engineering.

Figure 1

Schematic illustrating unique influenza B virus proteins. The RNA genome of the segmented influenza B virus (not to scale) with the unique gene products (relative to other influenza viruses) shown in color. Abbreviations: PB2—polymerase basic protein 2, PB1—polymerase basic protein 1, PA—polymerase acidic protein, HA—hemagglutinin, NP—nucleoprotein, NB—glycoprotein NB, NA—neuraminidase, M1—matrix protein, BM2—BM2 protein, NS1—non-structural protein 1, NEP—nuclear export protein.

Figure 2

Schematic outlining influenza virus reverse genetic systems. (a) Bi-cistronic plasmid that drives expression of positive-sense viral mRNA for gene transcription and negative-sense viral RNA for genomic replication. (b) Transfection of an eight-plasmid system (one plasmid per viral genomic segment) allows production of recombinant wild-type virus. Alternatively, genomic segment plasmids containing reporter constructs can be introduced in place of wild-type (WT) segments to generate viruses harboring reporter genes.

Figure 3

Designs of published influenza B virus genomic segments harboring reporter genes. (a–c) mNeon expressed in each of the polymerase subunit segments (d) NanoLuc expressed in the PB1 segment (e–g) Fluorescence proteins expressed in the NS1/NEP segment (h) Cre recombinase expressed in the PB1 segment (i) mNeon expressed in the HA segment. Abbreviations: PB2—polymerase basic protein 2, PB1—polymerase basic protein 1, PA—polymerase acidic protein, HA—hemagglutinin, NS1—non-structural protein 1, NEP—nuclear export protein. GFP—green fluorescent protein, UTR—untranslated region, *PS—silently mutated packaging signal, PS—duplicated packaging signal, PTV-1 2A—porcine teschovirus 2A sequence for co-translational separation, KDEL—endoplasmic reticulum retention sequence to prevent secretion, SD—Splice donor site, SA mutated—mutated Splice acceptor site.