Optimizing Efficient RNAi-Mediated Control of Hemipteran Pests (Psyllids, Leafhoppers, Whitefly): Modified Pyrimidines in dsRNA Triggers.

Wayne Brian Hunter,William M Wintermantel

doi:10.3390/plants10091782

Abstract

The advantages from exogenously applied RNAi biopesticides have yet to be realized in through commercialization due to inconsistent activity of the dsRNA trigger, and the activity level of RNAi suppression. This has prompted research on improving delivery methods for applying exogenous dsRNA into plants and insects for the management of pests and pathogens. Another aspect to improve RNAi activity is the incorporation of modified 2′-F pyrimidine nucleotides into the dsRNA trigger. Modified dsRNA incorporating 32–55% of the 2′-F- nucleotides produced improved RNAi activity that increased insect mortality by 12–35% greater than non-modified dsRNA triggers of the same sequence. These results were repeatable across multiple Hemiptera: the Asian citrus psyllid (Diaphorina citri, Liviidae); whitefly (Bemisia tabaci, Aleyroididae); and the glassy-winged sharpshooter (Homalodisca vitripennis, Cicadellidae). Studies using siRNA with modified 2′-F- pyrimidines in mammalian cells show they improved resistance to degradation from nucleases, plus result in greater RNAi activity, due to increase concentrations and improved binding affinity to the mRNA target. Successful RNAi biopesticides of the future will be able to increase RNAi repeatability in the field, by incorporating modifications of the dsRNA, such as 2′-F- pyrimidines, that will improve delivery after applied to fruit trees or crop plants, with increased activity after ingestion by insects. Costs of RNA modification have decreased significantly over the past few years such that biopesticides can now compete on pricing with commercial chemical products.

Highlights

In the hemipterans we identify multiple variants of Syntaxin 1A (1–4) supporting the findings of other studies in dipterans, Drosophila [179] that reported the Syntaxin 1A subfamily to consists of only two genes that function as the t-SNARE in synaptic vesicle fusion
The dsRNA triggers were made to the soluble Trehalase mRNA in D. citri and applied as exogenous topical sprays to citrus seedlings at 70 μg dsRNA per plant and provided for a 15-d feeding access period
Results showed that in the modified Syntaxin-dsRNA treatment there was a survival of 17.7% of whitefly nymphs that developed into adults at 9 days post-treatment

Summary

Introduction

Safer® Insecticidal Soap Concentrate (Woodstream Corporation, Lititz, PA, USA). M-PedeTM or Safer SoapTM, at concentrations of 2% v/v in water on citrus trees infested with psyllid adults and nymphs was only effective with repeated (>3) applications to produce. Direct sprays of soap solutions (0.8–2% in water) of either M-Pede or Safer Insecticidal Soap were acutely toxic to psyllid adults and nymphs (regardless of gender). The insecticidal soaps were not toxic to eggs at rates of up to 2% [199]. Residues of the soaps were not effective at reducing adult psyllids, even when the concentrations were increased to 4% [199]. Since the soaps caused some mortality effects of early nymphs upon hatching, we chose to compare Safer Insecticidal

Methods

Results

Conclusion