(335d) Development of Single-Virion Fusion Tool for Assessing Influenza Virus Pandemic Risk

Pandemic influenza remains a formidable threat to human health, but it is challenging, if not impossible, to predict which influenza virus subtype will cause the next pandemic. An alternative way is to assess the pandemic risk of all animal influenza viruses that have infected humans, identify those that show elements of pandemic potential, and work to alleviate that potential risk. To achieve these goals, a risk assessment framework was created to evaluate the risk elements including: genomic variation, receptor-binding properties, transmissibility in animal models, and antiviral treatment susceptibility[1]. In recent years, our group has shown that the membrane fusion process is an essential entry step of influenza virus that varies significantly with subtype and may also be an important factor that determines its pathogenicity [2]. Single-virion fusion assays combine total internal reflective fluorescence microscopic (TIRFM) and a microfluidic device coated with supported lipid bilayers that mimic host cell surfaces, to allow the direct observation of fluorescence-labeled viral particles binding to, and fusing with, the target cell membranes. This assay quantitatively characterizes: 1) the highest pH that initiates fusion, 2) the fusion kinetics across pH values, from early to mature endosomes, 3) the extent of fusion of the population, and 4) acid stability of hemagglutinin (HA). These fusion data characterize the release of viral genomes along the endosomal pathway, and we have shown that there are clear differences among various strains, e.g., clinical strains vs. attenuated strains. Hence, it may be necessary to include virus fusion evaluation into the influenza risk assessing framework.

Here, we performed single-virion fusion assay to assess the pandemic potential of influenza virus H10N8, a strain that transmitted from chickens to humans in China in 2013. This strain was lethal to three out of four infected human cases. Considering its high fatality, we used it as a model for studying a newly transmitted influenza virus. In our work, we employed pseudoviruses that contain the H10 and N8 proteins on a benign backbone for the purpose of classifying the fusion properties of this HA. In this presentation, we describe the particle construction, the fusion assay, and the analysis of the data to build a picture of this virus’s pandemic potential as it depends on its fusion properties. We believe fusion could be a key factor, that when taken with the other risk elements, may create a more complete framework to help scientists identify high-risk strains.

1. Trock, S.C., S.A. Burke, and N.J. Cox, Development of framework for assessing influenza virus pandemic risk. Emerging infectious diseases, 2015. 21(8): p. 1372.

2. Costello, D.A., G.R. Whittaker, and S. Daniel, Variations in pH Sensitivity, Acid Stability, and Fusogenicity of Three Influenza Virus H3 Subtypes. Journal of virology, 2015. 89(1): p. 350-360.