Dissemination of influenza B virus to the lower respiratory tract of mice is restricted by the interferon response.

The global burden of disease caused by influenza B virus (IBV) is substantial; however, IBVs remain overlooked. Understanding host-pathogen interactions and establishing physiologically relevant models of infection are important for the development and assessment of therapeutics and vaccines against IBV. In this study, we assessed an upper respiratory tract (URT)-restricted model of mouse IBV infection, comparing it to the conventional administration of the virus to the total respiratory tract (TRT). We found that URT infections caused by different strains of IBV disseminate to the trachea but resulted in limited dissemination of IBV to the lungs. Infection of the URT did not result in weight loss or systemic inflammation even at high inoculum doses and despite robust viral replication in the nose. Dissemination of IBV to the lungs was enhanced in mice lacking functional type I IFN receptor (IFNAR2), but not IFNγ. Conversely, in mice expressing the IFN-inducible gene Mx1, we found reduced IBV replication in the lungs and reduced dissemination of IBV from the URT to the lungs. Inoculation of IBV in both the URT and TRT resulted in seroconversion against IBV. However, priming at the TRT conferred superior protection from a heterologous lethal IBV challenge compared to URT priming, as determined by improved survival rates and reduced viral replication throughout the respiratory tract. Overall, our study establishes a URT-restricted IBV infection model, highlights the critical role of IFNs in limiting dissemination of IBV to the lungs, and also demonstrates that the lack of viral replication in the lungs may impact protection from subsequent infections. IMPORTANCE: Our study investigated how influenza B virus (IBV) spreads from the nose to the lungs of mice and the impact this has on disease and protection from re-infection. We found that when applied to the nose only, IBV does not spread very efficiently to the lungs in a process controlled by the interferon response. Priming immunity at the nose only resulted in less protection from re-infection than priming immunity at both the nose and lungs. These insights can guide the development of potential therapies targeting the interferon response as well as of intranasal vaccines against IBV.

Triton X-100-treated virus-based ELLA demonstrates discordant antigenic evolution of influenza B virus hemagglutinin and neuraminidase.

Neuraminidase (NA)-specific antibodies have been associated with protection against influenza and thus NA is considered a promising target for next-generation vaccines against influenza A (IAV) and B viruses (IBV). NA inhibition (NI) by antibodies is typically assessed using an enzyme-linked lectin assay (ELLA). However, ELLA can be confounded by anti-hemagglutinin (anti-HA) antibodies that block NA by steric hindrance (termed HA interference). Although strategies have been employed to overcome HA interference for IAV, similar approaches have not been assessed for IBV. We found that HA interference is common in ELLA using IBV, rendering the technique unreliable. Anti-HA antibodies were not completely depleted from sera by HA-expressing cell lines, and this approach was of limited utility. In contrast, we find that treatment of virions with Triton X-100, but not Tween-20 or ether, efficiently separates the HA and NA components and overcomes interference caused by anti-HA antibodies. We also characterize a panel of recombinant IBV NA proteins that further validated the results from Triton X-100-treated virus-based ELLA. Using these reagents and assays, we demonstrate discordant antigenic evolution between IBV NA and HA over the last 80 years. This optimized ELLA protocol will facilitate further in-depth serological surveys of IBV immunity as well as antigenic characterization of the IBV NA on a larger scale.IMPORTANCEInfluenza B viruses (IBVs) contribute to annual epidemics and may cause severe disease, especially in children. Consequently, several approaches are being explored to improve vaccine efficacy, including the addition of neuraminidase (NA). Antigen selection and assessment of serological responses will require a reliable serological assay to specifically quantify NA inhibition (NI). Although such assays have been assessed for influenza A viruses (IAVs), this has not been done of influenza B viruses. Our study identifies a readily applicable strategy to measure the inhibitory activity of neuraminidase-specific antibodies against influenza B virus without interference from anti-hemagglutinin (anti-HA) antibodies. This will aid broader serological assessment of influenza B virus-specific antibodies and antigenic characterization of the influenza B virus neuraminidase.