Selection for high levels of resistance to double-stranded RNA (dsRNA) in Colorado potato beetle (Leptinotarsa decemlineata Say) using non-transgenic foliar delivery

Swati Mishra,James Dee,Sandy Menasha,Gary Felton,James Baum,Tim Waters,Ken Ostlie,Justin Clements,William Moar,Mark Clough,Naymã Pinto Dias,Juan Luis Jurat-Fuentes,Aaron Buzza,William E Snyder,Jodi Dufner-Beattie,Andrei Alyokhin,Silvia I Rondon,Russell Groves

doi:10.1038/s41598-021-85876-1

Abstract

Insecticidal double-stranded RNAs (dsRNAs) silence expression of vital genes by activating the RNA interference (RNAi) mechanism in insect cells. Despite high commercial interest in insecticidal dsRNA, information on resistance to dsRNA is scarce, particularly for dsRNA products with non-transgenic delivery (ex. foliar/topical application) nearing regulatory review. We report the development of the CEAS 300 population of Colorado potato beetle (Leptinotarsa decemlineata Say) (Coleoptera: Chrysomelidae) with > 11,100-fold resistance to a dsRNA targeting the V-ATPase subunit A gene after nine episodes of selection using non-transgenic delivery by foliar coating. Resistance was associated with lack of target gene down-regulation in CEAS 300 larvae and cross-resistance to another dsRNA target (COPI β; Coatomer subunit beta). In contrast, CEAS 300 larvae showed very low (~ 4-fold) reduced susceptibility to the Cry3Aa insecticidal protein from Bacillus thuringiensis. Resistance to dsRNA in CEAS 300 is transmitted as an autosomal recessive trait and is polygenic. These data represent the first documented case of resistance in an insect pest with high pesticide resistance potential using dsRNA delivered through non-transgenic techniques. Information on the genetics of resistance and availability of dsRNA-resistant L. decemlineata guide the design of resistance management tools and allow research to identify resistance alleles and estimate resistance risks.

Highlights

Publication of the first study providing experimental evidence for the use of insecticidal double-stranded RNA as a plant-incorporated protectant[1] led to increasing interest in the commercial development of this technology for pest c ontrol[2]
The sustainable use and long-term efficacy of double-stranded RNA (dsRNA)-based insecticidal products should rely heavily on effective insect resistance management (IRM) strategies guided by information obtained from resistance studies in model pests
Using L. decemlineata, a target pest for development of dsRNA-based insecticidal products and a species with a history of developing resistance to nearly every class of insecticide[15], we describe the first case of insect resistance against non-transgenic foliar delivery of insecticidal dsRNA

Summary

Introduction

Publication of the first study providing experimental evidence for the use of insecticidal double-stranded RNA (dsRNA) as a plant-incorporated protectant[1] led to increasing interest in the commercial development of this technology for pest c ontrol[2]. Interest in commercialization of this technology has fueled progress towards evaluating the insecticidal activity of dsRNA against several insect pests, and studying the underlying mechanisms that determine RNAi efficiency, including different methods of dsRNA delivery[9]. In contrast to transgenic delivery of dsRNA, there is currently no available information for any insect on mechanisms of resistance to insecticidal dsRNA delivered using non-transgenic methods. It is plausible that different mechanisms of resistance to dsRNA may be selected in different target species, as suggested by the differences in susceptibility to dsRNA observed among insects[13,14] These observations highlight the importance of considering different delivery methods and target pests in dsRNA resistance studies

Methods

Results

Conclusion