White spot syndrome virus VP28 specific double-stranded RNA provides protection through a highly focused siRNA population

Several studies have demonstrated that injection of double-stranded RNAs (dsRNA) homologous to mRNA for the white spot syndrome virus (WSSV) viral protein 28 (VP28) can induce protection in shrimp against WSSV through RNA interference (RNAi). In comparison to shrimp injected with either PBS or a green fluorescent protein (GFP) nonspecific dsRNA, we obtained nearly complete protection against WSSV infection in shrimp injected with VP28 dsRNA. Upregulation of host genes associated with small RNA silencing was measured 48 hours post treatment in groups injected with dsRNA, and although the VP28-treated group remained moderately upregulated after challenge with WSSV, many-fold higher induction was observed in both control groups reflecting the ongoing viral infection. RNA sequencing of VP28-treated shrimp demonstrated a siRNA population dominated by high levels of 22 nt long molecules narrowly targeting the VP28 mRNA both before and after challenge with WSSV. Conversely, while no siRNAs targeting WSSV were detected before challenge, a broad response of 22 nt siRNAs mapping across the entire WSSV genome were found in both control groups after challenge. These results give detailed insight to how dsRNA targeting VP28 function to induce protection against WSSV, by generating a highly focused population of 22 nt long siRNA molecules.