Background: Pepino mosaic virus (PepMV) is an emerging plant pathogen that infects tomatoes worldwide. Understanding the factors that influence its evolutionary success is essential for developing new control strategies that may be more robust against the evolution of new viral strains. One of these evolutionary factors is the distribution of mutational fitness effect (DMFE), that is, the fraction of mutations that are lethal, deleterious, neutral, and beneficial on a given viral strain and host species. The goal of this study was to characterize the DMFE of introduced nonsynonymous mutations on a mild isolate of PepMV from the Chilean 2 strain (PepMV-P22). Additionally, we also explored whether the fitness effect of a given mutation depends on the gene where it appears or on epistatic interactions with the genetic background. To address this latter possibility, a subset of mutations were also introduced in a mild isolate of the European strain (PepMV-P11) and the fitness of the resulting clones measured.
Results: A collection of 25 PepMV clones each containing a single nucleotide nonsynonymous substitution was created by site-directed mutagenesis and the fitness of each mutant was determined. PepMV-P22 genome showed a high degree of robustness against point mutations, with 80% of mutations being either neutral or even beneficial and only 20% being deleterious or lethal. We found that the effect of mutations strongly depended on the gene in which they were introduced. Mutations with the largest average beneficial effects were those affecting the RdRp gene, in contrast to mutations affecting TGB1 and CP genes, for which the average effects were deleterious. Moreover, significant epistatic interactions were observed between nonsynonymous mutations and the genetic background, meaning that the effect of a given nucleotide substitution on a particular genomic context cannot be predicted by knowing its effect in a different one.
Conclusions: Our results indicated that PepMV genome has a surprisingly high robustness against mutations. We also found that fitness consequences of a given mutation differ between the two strains analyzed. This discovery suggests that the strength of selection, and thus the rates of evolution, vary among PepMV strains.
Keywords: Epistasis; Mutational fitness effect; Pepino mosaic virus; Real-time quantitative PCR; Site-directed mutagenesis; Virus accumulation; Virus evolution.