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Novel pesticides based on suppression of gene expression by Ribonucleic acid (RNA) -protein complexes - PS2147

Description
Insect pests of crops are a continuing problem in agriculture. Although chemical pesticides have been successfully used for many years, the environmental damage caused by non-specific pesticides has become a factor in limiting their use, or even in requiring their withdrawal from the marketplace. There is concern at present that existing pesticides, which are not harmful to humans or higher animals when correctly deployed, can have effects on beneficial insects - for example, neonicotinoid pesticides have been implicated in the decline in bee populations. An ideal insecticide would have no effects on any other organisms than those which it was designed to kill.

Research carried out previously by the applicants has sought to develop insecticides based on specific toxins produced by spiders and scorpions, and has developed a technology to allow these to be effective when delivered orally. While these toxins are insect-specific, and show selectivity between different orders of insect pests, they are not specific to individual pest species. To produce insecticides that are close to species-specific, a different approach is required.

The use of RNA interference (RNAi) to suppress gene expression is well-established as a research technique in insects. This relies on an artificial, designed double stranded RNA (dsRNA) being introduced in an insect; this results in the transcript of a specific gene being destroyed, so that the protein product of the gene is not produced. If the targetted gene is carefully selected, the resulting phenotype can be lethal - for example, if RNAi is used to suppress expression of a gene that produces a product vital for development. The effect is highly specific to a single species and close relatives, as it depends on the base sequences of the dsRNA and the target gene matching. We have shown that a specific dsRNA can result in RNAi effects that cause mortality to a crop pest, cabbage root fly, using genomic and DNA sequence information from the model dipteran species, the fruit fly Drosophila melanogaster (whose genetics have been extensively studied). In this case, house fly, Musca domestica, was used as a "working model" to develop methods and carry out initial assays to validate the effects produced by specific dsRNAs.

To turn this research into a crop protection method requires further technical development. The RNAi results obtained previously have been based on injection of dsRNA, which is not feasible for crop protection; oral delivery is not reproducibly effective. The proposed research programme therefore seeks to develop a method for oral delivery of dsRNA which will result in increased uptake, and effective suppression of gene expression, leading to mortality in the insect pest. This will be achieved by modifying the fusion protein technology used previously to confer oral toxicity onto spider and scorpion toxins, which has been shown to allow delivery of ingested materials to the insect circulatory system, as if they had been injected. The programme will seek to develop an orally-effective RNAi-based insecticide that can be produced by fermentation technologies, allowing scale-up for possible commercial manufacture.
Objective
1. To produce a recombinant RNA-binding carrier protein for transport of dsRNA across the gut epithelium of insects (9 months), and to demonstrate that the protein forms stable complexes with dsRNA (12 months).
2. To feed a carrier-dsRNA complex to the "working model" insect and show that RNA transport across the gut epithelium into the haemolymph occurs (18 months).
3. To show that carrier-dsRNA complexes produced with dsRNAs directed against the M. domestica and D. radicum homologues of the D. melanogaster thread gene can cause gene suppression and effects on phenotype, including mortality (24 months).
4. To investigate "scale-up" of production of carrier-dsRNA complexes for commercial production (24 months)
Project Documents
• FRP - Final Report : PS2147 Final Report   (100k)
Time-Scale and Cost
From: 2013

To: 2015

Cost: £150,254
Contractor / Funded Organisations
Durham University, F E R A (FERA)
Keywords
Application              
Insecticide use              
Fields of Study
Pesticide Safety