Abstract
Deltaviruses were recently discovered across a broad range of metazoan species. Their genome encodes a single protein and thus requires host factors for replication, which are likely evolutionarily conserved given their broad host range. Here we describe a genetic tool to unambiguously determine whether an RNA virus replicates in model yeast species. Our system involves transcriptional induction of viral RNA from an integrated cDNA template followed by its genomic excision. Testing Hepatitis D and Rodent delta viruses revealed that neither viral RNAs can replicate in yeasts, suggesting that Ascomycetes lack factors essential for their RNA-dependent amplification.