Infectious bursal disease virus is an icosahedral polyploid dsRNA virus

Abstract

Viruses are a paradigm of the economy of genome resources, reflected in their multiplication strategy and for their own structure. Although there is enormous structural diversity, the viral genome is always enclosed within a proteinaceous coat, and most virus species are haploid; the only exception to this rule are the highly pleomorphic enveloped viruses. We performed an in-depth characterization of infectious bursal disease virus (IBDV), a non-enveloped icosahedral dsRNA virus with a bisegmented genome. Up to 6 natural populations can be purified, which share a similar protein composition but show higher sedimentation coefficients as particle density increases. Stoichiometry analysis of their genome indicated that these biophysical differences correlate with the copy number of dsRNA segments inside the viral capsid. This is a demonstration of a functional polyploid icosahedral dsRNA virus. We show that IBDV particles with greater genome copy number have higher infectivity rates. Our results show an unprecedented replicative strategy for dsRNA viruses and suggest that birnaviruses are living viral entities encompassing numerous functional and structural characteristics of positive and negative ssRNA viruses.

Fig. 1.

Purification of IBDV natural populations. (A) A typical CsCl linear gradient for IBDV purification illuminated from the bottom after centrifugation to equilibrium, containing at least 6 IBDV fractions (denoted E1 to E6, from top to bottom). E1–E6 bands represent 3, 5, 12, 16, 48, and 16% of the total virion particles. (B) E1–E6 bands were collected by side puncture, analyzed by SDS/PAGE and developed by Coomassie staining. IBDV structural proteins are indicated. (C) Electron microscopy of purified E1–E6 populations negatively stained with 2% uranyl acetate. T = 7 capsid-like (arrows) and T = 1 capsid-like particles (arrowheads) are indicated for the E1 population. (Scale bar, 100 nm.)

Fig. 2.

Genome stoichiometry of IBDV populations. Agarose gel electrophoresis and autoradiography of intact (A) or SDS- and proteinase K–treated (B) [³³P]HPO₄⁻²-labeled E1–E6 IBDV populations. The same amount of total protein was analyzed for each IBDV population. vRNA corresponds to viral RNA packaged inside intact virion particles; dsRNA-A and dsRNA-B correspond to purified dsRNA segments A and B, obtained after SDS and proteinase K treatments. (C) Relative IBDV dsRNA content based on ³³P labeling for E2–E6 populations. Counts in the bands were measured and normalized with respect to the E2 population value. Average and standard deviation values were calculated from triplicate experiments.

Fig. 3.

Titration of IBDV populations on QM7 monolayers. Number of particles was estimated from protein concentration measurements of purified E1–E6 populations. Error bars indicate the standard deviations of titrations of 3 independent experiments.