Altering the size distribution of influenza virion populations

Harry Smith Vacation Studentship Laboratory-adapted influenza viruses produce predominantly spherical virions. In contrast, clinical and veterinary isolates produce a mixture of virions of different sizes, from 0.1 µm spheres to filaments which can reach tens of microns in length. Filamentous influenza virions were discovered in 1946, but the bulk of influenza research has analysed only spherical forms of the virus and the role of filaments in influenza infections is unclear. Functional studies of filaments require the development of methods to manipulate the ratio of spherical to filamentous virions, and we reasoned that this could be achieved by filtration. To test this, we infected MDCK cells with the filamentous Udorn strain of influenza A virus. We collected virus-containing growth media and passed this through filters with 5 µm, 0.45 µm and 0.2 µm pores. Filtrates and unfiltered virus were compared, using Western blotting to measure their protein composition, plaque assays to measure their infectivity and negative stain transmission electron microscopy to measure individual particle sizes. We found that filtration through a filter with 5 µm pores had little effect on composition, infectivity and the ratio of spherical to filamentous particles. In contrast, sub-micron filters, particularly those with 0.2 µm pores, caused a general depletion of virions but increased the sphere to filament ratio. We therefore concluded that sub-micron pore sizes can be used to preferentially remove filaments from populations of pleomorphic influenza virions, providing a useful tool for subtractive studies of the contribution filaments make to influenza virus infections.