Generation and genetic characterization of avian reovirus temperature-sensitive mutants

There currently is little known about the genetic and biological functions of avian reovirus (ARV), an atypical member of the family Reoviridae and the prototype of all nonenveloped viruses that induce syncytia formation. In this study, we created ARV temperature-sensitive (ts) mutants by chemical mutagenesis of ARV strain 138. We developed a novel efficiency of lysis (EOL) screening technique and used it and the classical efficiency of plating (EOP) assay to identify 17 ARV ts mutants. Pairwise mixed infections of these mutants and evaluation of recombinant progeny ts status led to their organization into seven recombination groups. This indicates that these new groups of mutants represent the majority of the ARV genome. To phenotypically characterize the ts mutants, progeny double-stranded RNA (dsRNA) produced at permissive and nonpermissive temperature was measured. Some mutants were capable of dsRNA synthesis at the restrictive temperature (RNA(+)), which indicates the effects of their ts lesions occur after RNA replication. Most mutants were RNA(-), which suggests their mutations affect stages in viral replication that precede progeny genome synthesis.