Length-dependent innate antiviral effects of double-stranded RNA in the rainbow trout (Oncorhynchus mykiss) cell line, RTG-2

• dsRNA induced type I IFN mediated antiviral responses in a length dependent manner, longer dsRNA induced a stronger response.
• Length dependence was observed for both poly I:C and in vitro transcribed dsRNA molecules.
• In vitro transcribed dsRNA molecules induced a different transcript expression profile compared to poly I:C.
• The length dependent effects were not as a result of entry efficiency differences.

Effectively all viruses produce long dsRNA during their replicative cycle. In mammals long dsRNA molecules induce a robust response through the production of type 1 interferon, interferon-stimulated genes (ISGs) and an antiviral state. This response is less well understood in fish. We investigated the ability of a rainbow trout cell line, RTG-2, to respond to two different lengths of in vitro transcribed dsRNA (200 bp and 1264 bp) based on the viral hemorrhagic septicemia virus genomic sequence, and high and low molecular weight poly I:C (synthetic dsRNA). To explore the innate immune response we used qRT-PCR to measure immune gene transcript levels, an ISG-promoter reporter assay, and an antiviral protection assay. We saw a significantly greater immune response in all assays in response to the longer dsRNA molecule compared to their shorter counterpart. We saw significantly more interferon and ISG transcripts, stronger induction of a protective antiviral state, and more robust activation of the ISG-promoter. This response was not found to be due to a better uptake of the longer dsRNA molecules as a cellular uptake assay showed no differences between lengths. These data suggest that dsRNA-mediated innate immune responses are length-dependent and longer molecules induce a more robust response. There were also some differences in the cells response to in vitro transcribed dsRNA compared to poly I:C. This provides important information for potential dsRNA-based antiviral therapies and vaccine adjuvants.