Rescue of the 1947 Zika Virus Prototype Strain with a Cytomegalovirus Promoter-Driven cDNA Clone

Schwarz, Megan C.; Sourisseau, Marion; Espino, Michael M.; Gray, Essanna S.; Chambers, Matthew T.; Tortorella, Domenico; Evans, Matthew J.

For most positive-strand RNA viruses, where the genome directly encodes the proteins required for its replication, transfection of a permissive cell with viral RNA is sufficient to initiate infection that eventually results in the release of viral progeny. These genomes can be cloned into plasmids with elements required for in vitro transcription of the long viral RNA for transfection. Such an approach has only recently been used to rescue a 2010 Cambodia ZIKV isolate (5) . Since flaviviruses have capped RNA genomes, the transcription step can be circumvented by transfecting a plasmid carrying the viral genome under the control of an RNA polymerase II promoter, with the correct 3= end of the viral RNA being generated by a hepatitis D virus ribozyme (HDVr) (6, 7) . The plasmid DNA is required only to initiate transcription of the first round of viral RNA, which can serve as both a translation and a replication template to initiate "infection" of transfected cells. Indeed, this is the approach that Tsetsarkin et al. recently used to rescue a 2015 Brazil ZIKV isolate (8) .