As the interest in RNAi for ag-bio applications continues its rapid growth, the US Department of Agriculture has earmarked $500,000 to fund a three-year research project designed to create a framework for assessing the risk of crops that have been modified using the gene-silencing technology, Gene Silencing News has learned.
While the effort focuses on arthropods consuming corn that has been modified with double-stranded RNA, it is expected to help establish a method to assess the broader impact of RNAi, whether it is used to modify plants directly or added to pesticides, USDA research entomologist and project leader Jon Lundgren said.
While the application of RNAi to human health has received much attention, companies in the ag-bio space have also been increasingly looking to the technology to improve crop plants.
In September, for instance, Monsanto paid $29.2 million to pick up the exclusive rights to use Alnylam Pharmaceuticals' RNAi intellectual property for agricultural applications (GSN 9/6/2012). A Monsanto spokesperson said at the time that the company anticipates using RNAi for “a broad range of applications,” including pest control.
Monsanto is currently developing a strain of RNAi-modified corn that is resistant to corn rootworm, as well as a strain of soybeans that incorporates gene silencing to yield trans-fat-free and reduced-saturated fat oils. The firm is also developing an RNAi-based treatment for Israeli acute paralysis virus, which affects bees.
Other companies have gotten into the game too, including Syngenta, which recently bid $518 million to buy Devgen and its portfolio of RNAi-based crop-enhancement and -protection technologies (GSN 9/27/2012).
“Pest-control technology is changing so quickly, and there are new approaches for reducing agricultural pests all the time,” Lundgren said. “Understanding the long-term implications of these [technologies] is an important process to go through.
“With the rapid development of RNAi-based pest management [tools] … it became clear that this is something we're going to have to deal with,” he added. “The technology … is coming on very quickly and could be perfectly safe — we just want to be sure that it is.”
Lundgren stressed that while much has been learned about the effects of RNAi in the fields of functional genomics and human therapeutics, “we're operating at a little bit different scale.
“We're not talking about an individual cell or cell line, we're not talking about a sick person … we're talking about transforming a series of crop plants that are planted on 10 percent of the terrestrial land surface of the continental United States,” he said. From an environmental risk-assessment point of view, things such as off-target effects and non-target binding of small RNAs are “very important issues … that could have great effects on many species.”
The research project, which began on Sept. 1 and runs until the end of August 2015, is specifically geared toward determining “the likelihood of exposure to and toxicity of interference RNA to a core-based arthropod food web,” according to the USDA. It will “establish which species are at risk through consuming dsRNA-containing corn tissue under field conditions, and whether dsRNAs are transferred to higher trophic levels via consuming herbivorous prey.”
The first step involves using qPCR-based gut analysis to create a taxon-specific consumption index of corn tissue that is based on a frequency of consumption and the quantity of DNA consumed.
“By looking for corn-specific DNA sequences in the insects' stomachs, we can create a linkage between that particular organism and the corn crop,” Lundgren explained.
He noted that within just the USDA's corn fields in South Dakota, where the project is being run, “we could be talking about hundreds and hundreds of species. … We know the dominant species that occur within a corn-agro ecosystem, but we really don't know what the relative strengths of their trophic interactions are.”
The project will help address that question, making use gut-DNA extractions from 5,672 corn arthropods collected from corn fields of eastern South Dakota that have yielded primer sets useful in determining which arthropods have consumed corn tissue.
Once the key corn-consuming arthropods are identified, “we'll do some toxicity assays investigating a range of insecticidal small RNAs that are being developed either as a spray-on … or plugged into the crop plant itself to start looking at some fitness effects on these non-targeted organisms,” Lundgren said.
In this part of the project, insecticidal dsRNAs will be administered to three corn pests with different feeding ecologies: Coleomegilla maculata, Orius insidiosus, and Chrysoperla carnea.
“The presence of the interference RNA molecules in the various insects will be assessed using targeted PCR,” the USDA said. “If interference RNA is found to be present in the predators, then an exposure analysis of key herbivores and their predator communities will be conducted using PCR-based gut analysis.”
“Of those species that are exposed to the RNAi-containing corn or prey, specific indicator taxa will be selected from key functional groups and their genomes will be sequenced and placed into a searchable database,” the agency added.
Importantly, this database is expected to be useful for screening sequence homologies between current and future RNAi targets for use in non-target toxicity assays.
“Because we don't know the exact small RNAs that are going to be expressed in the plants or used as insecticides,” Lundgren said, “we want to build an adaptable system that we can have in place ... to assess not just what's coming down the pike right now, but for any new forms of RNAi-based technologies that may be coming in the future.”