Alejandra Flores, Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA

Alejandra Flores 1, Ryan D. Xavier 1, Kristen M. Ogden 1,2 1 Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA 2 Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tennessee, USA

Virus Replication: Entry, Exit and Everything in Between

Recombination facilitates the generation of defective viral genomes (DVGs), truncated derivatives of the parental genome that require a helper virus to replicate. Although recombination mechanisms that generate DVGs are described for positive-sense and negative-sense RNA viruses, such mechanisms are poorly understood for viruses with double-stranded RNA genomes. To investigate the contributions of the polymerase complex, which mediates recombination, to patterns of DVGs and recombination junctions, we exchanged the polymerase complex between two strains of mammalian orthoreovirus (reovirus) that are known to differ in DVG pattern. We identified recombination junctions using ClickSeq, which avoids artifactual recombination, and ViReMa, an algorithm that annotates recombinant species in virus populations. The polymerase complex did not obviously affect viral recombination junction patterns. Instead, these patterns correlated with the identity of a viral RNA gene segment, even in the presence of a non-native polymerase complex or virus background. We also found that specific sites within the sequence of a given viral gene segment are preferred for recombination. Junction start and stop sites often occur in regions of sequence microhomology. While we detect many instances of these short stretches of identical nucleotides within a given viral gene segment, only a select few positions are associated with recombination junctions. Overall, these data suggest that for reovirus, recombination events that can mediate DVG formation are highly selective, and properties of viral RNA segments, rather than those of the polymerase complex, primarily dictate where recombination occurs. Insights from these observations suggest a rudimentary model for double-stranded RNA virus recombination.