The hidden diversity: Replication-strand asymmetry shapes short-term nucleotide and structural variation in an RNA virus

The Orsay virus (OrV), a positive-sense RNA virus that infects the microscopic worm Caenorhabditis elegans, establishes latent infections without significantly affecting the host's fitness. Researchers employed strand-specific RNA-seq across twelve time points corresponding to the worm's larval development to examine the OrV replication dynamics and short-term evolutionary patterns.

The viral accumulation was observed to follow four distinct phases and displayed a strong strand asymmetry, which is indicative of the predominant stamping-machine replication mechanism.

When conducting diversity analyses, scientists discovered that the negative-strand genomes contained a higher number of mutations compared to the positive strands. These mutations were particularly concentrated in two regions: the replicase C-terminal tail and the structural protein regions. Interestingly, many negative-strand variants were unable to successfully transfer to the positive strands, suggesting that there are constraints or purifying selection occurring at the RNA level within cells.

In addition to the previously mentioned findings, the research also revealed the existence of non-standard viral genomes in OrV for the first time. These genomes were dominated by deletions in RNA2, including a recurrent large deletion that may serve as a subgenomic RNA encoding a shorter version of the {delta} protein. The findings presented in this study emphasize replication asymmetry, strand-specific mutational filtering, and the prevalence of non-standard genomes as crucial factors influencing OrV's evolution during the initial stages of infection.