Theme and variations in the evolutionary pathways to virulence of an RNA plant virus species

The diversity of a highly variable RNA plant virus was considered to determine the range of virulence substitutions, the evolutionary pathways to virulence, and whether intraspecific diversity modulates virulence pathways and propensity. In all, 114 isolates representative of the genetic and geographic diversity of Rice yellow mottle virus (RYMV) in Africa were inoculated to several cultivars with eIF(iso)4G-mediated Rymv1-2 resistance. Altogether, 41 virulent variants generated from ten wild isolates were analyzed. Nonconservative amino acid replacements at five positions located within a stretch of 15 codons in the central region of the 79-aa-long protein VPg were associated with virulence. Virulence substitutions were fixed predominantly at codon 48 in most strains, whatever the host genetic background or the experimental conditions. There were one major and two isolate-specific mutational pathways conferring virulence at codon 48. In the prevalent mutational pathway I, arginine (AGA) was successively displaced by glycine (GGA) and glutamic acid (GAA). Substitutions in the other virulence codons were displaced when E48 was fixed. In the isolate-specific mutational pathway II, isoleucine (ATA) emerged and often later coexisted with valine (GTA). In mutational pathway III, arginine, with the specific S2/S3 strain codon usage AGG, was displaced by tryptophane (TGG). Mutational pathway I never arose in the widely spread West African S2/S3 strain because G48 was not infectious in the S2/S3 genetic context. Strain S2/S3 least frequently overcame resistance, whereas two geographically localized variants of the strain S4 had a high propensity to virulence. Codons 49 and 26 of the VPg, under diversifying selection, are candidate positions in modulating the genetic barriers to virulence. The theme and variations in the evolutionary pathways to virulence of RYMV illustrates the extent of parallel evolution within a highly variable RNA plant virus species.
Author Summary Parallel changes in independently evolving lineages are important, but their contribution to pathogen evolution has not been assessed at the species level. We investigated the extent of phenotypic and genotypic parallel evolution in a highly variable RNA plant virus species, Rice yellow mottle virus (RYMV). Isolates representative of the genetic and geographic diversity of RYMV in Africa were inoculated to several rice cultivars with eIF(iso)4G-mediated Rymv1-2 resistance. The theme and variations in the evolutionary pathways to gain virulence found in the VPg of RYMV illustrate the frequency of parallel evolution. The repeated occurrence of the R48E substitution in the VPg of most strains, whatever the Rymv1-2 background and plant growth conditions, showed the specificity of parallel evolution that operated through the same pathway, locus, and mutation. The frequency and specificity of parallel mutations indicate, respectively, that RYMV is able to rapidly explore the adaptive landscape, fixing favorable mutations to virulence, and that there are a limited number of pathways across the adaptive landscape. Our results provide insights into the ways an RNA virus species explores the adaptive landscape and into the constraints restricting the number of mutational pathways.