Engineering stress as a motivation for filamentous virus morphology

Abstract

Many viruses are pleomorphic in shape and size, with pleomorphism often thought to correlate with infectivity, pathogenicity, or virus survival. For example, influenza and respiratory syncytial virus particles range in size from small spherical virions to filaments reaching many micrometers in length. We have used a pressure vessel model to investigate how the length and width of spherical and filamentous virions can vary for a given critical stress and fluorescence super-resolution microscopy along with image analysis tools to fit imaged influenza viruses to the model. We have shown that influenza virion dimensions fit within the theoretical limits of the model, suggesting that filament formation may be a way to increase an individual virus's volume without particle rupture. We have also used cryoelectron microscopy to investigate influenza and respiratory syncytial virus dimensions at the extrema of the model and used the pressure vessel model to explain the lack of alternative virus particle geometries. Our approach offers insight into the possible purpose of filamentous virus morphology and is applicable to a wide range of other biological entities, including bacteria and fungi.

Fig. 1

Schematics of the cross sections of interest and the pressure/stress relations derived for increasing lengths of virus particles. Viruses are modeled as cylinders with hemispherical caps. For different lengths of cylinders, L = 0, L > 0, and L→ꝏ, the pressure/stress relation is given along with the cross section of interest. In the limits of L→0 and L→ꝏ, the model used for filamentous viruses tends to the known results for spheres and pipes of an infinite length.

Fig. 2

Super-resolution imaging of spherical and filamentous influenza particles fit against the predicted theory from pressure vessel analysis. (A) A representative direct stochastic optical reconstruction microscopy (dSTORM) field of view (FOV) of labeled A/Udorn/72 influenza where the hemagglutinin protein is imaged in the red channel. Scale bar, 10 μm. (B and C) Zoomed-in images from (A) showing individual filaments and spherical particles. Scale bar, 200 nm. (D) A representative dSTORM FOV of a virus-negative sample, imaged in the red channel. Scale bar, 10 μm. (E) The major/minor axis contour plot with lines at major axis = minor axis (red) and with the line as given in Eq. 6 describing the derived allowable relation between major and minor axes with r₀ = 70 nm (yellow) with a maximum frequency of 100 for clarity. (F) The major/minor axis scatterplot with lines at minor axis = 140 nm (red) and major axis = minor axis (red) and with the line as given in Eq. 6 describing the derived allowable relation between major and minor axes with r₀ = 70 nm (yellow).

Fig. 3

Cryo-electron tomography of filamentous and spherical virus particles and measurements of their diameter and membrane thickness. (A) Representative tomogram of filamentous A/Udorn/72 influenza virus. Scale bar, 100 nm. (B) Representative tomogram of filamentous A2 respiratory syncytial virus. Scale bar, 100 nm. (C and D) Bar graphs of the average outer and inner diameters and the membrane thicknesses of the two filamentous viruses. (E) Representative tomogram of spherical A/WSN/33 influenza virus particles. Scale bar, 100 nm. (F) Bar graphs of the average outer and inner diameters and the membrane thicknesses of A/WSN/33 spheres.

Fig. 4

Schematic of the proposed shapes formed from symmetric enlargement in three, two, or one dimensions and the outcome for virions with those geometries. The three possible shapes of the virus when they are enlarged symmetrically in either one, two, or three dimensions are shown. Three-dimensional enlargement is predicted to result in virion rupture when the maximum stress is reached, while growth in two dimensions is predicted to result in ballooning that will lead to a similar fate. Our data suggest that a filamentous shape with enlargement along a single plane is not likely to encounter this issue.