Expanding, Characterizing, and Repurposing the Invertebrate Virosphere

Abstract

Modern medicine demands the capacity to deliver genetic or biological cargo to specific cell types.  Past efforts to achieve this goal have relied on the retooling and re-engineering of a small subset of vertebrate viruses with limited success.  Remaining challenges with regards to in vivo delivery include finding novel viral vectors that can achieve different target specificities in addition to those that are more amenable to synthesize de novo.  In an attempt to address these remaining limitations, we collected and sampled diverse invertebrate species to isolate and identify RNA viruses associated with them.  As the invertebrate virosphere remains largely unknown, we hypothesized that we would identify novel viruses whose components could be characterized and repurposed to build a new suite of viral-based tools.  To this end, we isolated and sequenced RNA from a diverse library of invertebrates (including 42 insects) by next-generation sequencing and subsequently performed de novo genome assembly on the reads obtained. Captured reads were analyzed for signatures of RNA dependent RNA polymerases (RdRps) – a necessary component of all RNA viruses. The two putative novel virus genome assemblies discovered were named Castor and Pollux, and were characterized and independently confirmed by quantitative PCR.  These small RNA viruses or their RdRps (less than 5kB) will, in the future, be synthesized and artificially launched in mammalian cells to ascertain whether they can be selected via guided evolution to function and deliver a desired genetic or biological cargo.