Long dsRNAs promote an anti-viral response in Pacific oyster hampering ostreid herpesvirus 1 replication

Double stranded RNA-mediated genetic interference (RNAi) is a widely used reverse genetic tool for determining the loss-of-function phenotype of a gene. Here, the possible induction of an immune response by long dsRNA was tested in a marine bivalve, i.e. Crassostrea gigas, as well as the specific role of the subunit 2 of the nuclear factor κB inhibitor (IκB2). This gene is a candidate of particular interest for functional investigations in the context of massive mortality oyster events as Cg-IκB2 mRNA levels exhibited significant variation depending on the amount of ostreid herpesvirus 1 (OsHV-1) DNA detected. In the present study, dsRNAs targeting Cg-IκB2 and Green Fluorescence Protein genes were injected in vivo into oysters before being challenged by OsHV-1. Survival appeared close to 100% in both dsRNA injected conditions associated with a low detection of viral DNA and a low expression of a panel of 39 OsHV-1 genes as compared to infected control. Long dsRNA molecules, both Cg-IκB2- and GFP-dsRNA, may have induced an anti-viral state controlling the OsHV-1 replication and precluding the understanding of the Cg-IκB2 specific role. Immune-related genes including Cg-IκB1, Cg-Rel1, Cg-IFI44, Cg-PKR, and Cg-IAP appeared activated in dsRNA-injected condition potentially hampering viral replication and thus conferring a better resistance to OsHV-1 infection. We revealed that long dsRNA-mediated genetic interference triggered an anti-viral state in the oyster, emphasizing the need of new reverse genetics tools for assessing immune gene function and avoiding off-target effects in bivalves.