Silencing of Double-Stranded Ribonuclease Improves Oral RNAi Efficacy in Southern Green Stinkbug Nezara viridula

Abstract

Variability in RNA-interference (RNAi) efficacy among different insect orders poses a big hurdle in the development of RNAi-based pest control strategies. The activity of double-stranded ribonucleases (dsRNases) in the digestive canal of insects can be one of the critical factors affecting oral RNAi efficacy. Here, the involvement of these dsRNases in the southern green stinkbug Nezaraviridula was investigated. First, the full sequence of the only dsRNase (NvdsRNase) in the transcriptome of N. viridula was obtained, followed by an oral feeding bioassay to evaluate the effect of NvdsRNase-silencing on oral RNAi efficacy. The NvdsRNase was first silenced in nymphs by NvdsRNase-dsRNA injections, followed by exposure to an artificial diet containing a lethal αCop-specific dsRNA. A significantly higher mortality was observed in the NvdsRNase-silenced nymphs when placed on the dsαCop-containing diet (65%) than in the dsGFP injected and dsαCop fed control (46.67%). Additionally, an ex vivo dsRNA degradation assay showed a higher stability of dsRNA in the saliva and midgut juice of NvdsRNase-silenced adults. These results provide evidence for the involvement of NvdsRNase in the reduction of oral RNAi efficacy in N. viridula. This information will be useful in further improving potential RNAi-based strategies to control this pest.

A Protective dsRNA is Crucial for Optimum RNAi Gene Silencing in Chilo partellus

“…The protection of dsRNA from nuclease will be a better way of achieving optimum knockdown in C. partellus and facilitating RNAi-mediated control strategy. Efforts to protect dsRNA from nucleases degradation by silencing nuclease have been demonstrated to enhance dsRNA uptake in agricultural pests and subsequently improve RNAi efficiency (Giesbrecht et al 2020;Wang et al 2020;Sharma et al 2021). Research evidence has shown that nanoparticle-mediated delivery overcomes poor cellular internalization and nucleolytic degradation of dsRNAs for effective RNAi response in insect cells (Christiaens et al 2018;Dhandapani et al 2019;Wang et al 2019;Baddar et al 2020;Yan et al 2021).…”

RNA interference in Pardosa pseudoannulata, an important predatory enemy against several insect pests, through ingestion of dsRNA‐expressing Escherichia coli

“…A double‐stranded RNA nuclease was associated with rapid degradation of dsRNA in the saliva of the southern green stinkbug ( Nezara viridula ). And the blockage of dsRNA nuclease synthesis significantly enhances the RNAi efficacy via orally delivered dsRNA in N. viridula (Sharma et al ., 2021). To overcome this barrier, formulation of dsRNA with polymer or liposomic nanoparticles was a strategy, which can prevent degradation of dsRNA in the digestive system of insects (Christiaens et al ., 2018; 2020).…”

Lab-to-Field Transition of RNA Spray Applications – How Far Are We?

“…Given these challenges, a recent breakthrough was achieved, demonstrating that chitosan nanoparticles-mediated dsRNA delivery in Helicoverpa armigera protects from degradation by nucleases and insect gut pH (Kolge et al, 2021). Beyond shielding dsRNA from RNase degradation, the silencing of a dsRNA ribonuclease improved oral RNAi efficacy in the southern green stinkbug (Sharma et al, 2021). This finding indicates that the simultaneous application of protective agents such as RNase inhibitors or the silencing of dsRNases might encourage targeted detoxification mediated by P450s, ABC transporters and others (Wang et al, 2021).…”