Insecticides help combat infectious diseases carried by mosquitoes and other insects, but overuse of these products has resulted in insecticide resistance. In certain insecticides, the active compounds bind to voltage-gated sodium ion channels of the nervous system. However, a common point mutation in this ion channel changes a normal leucine to a phenylalanine, making the insects resistant to these compounds.1 “It’s like a soft spot in the protein,” explained Ethan Bier, a geneticist at the University of California, San Diego.
Ethan Bier and his team developed a gene drive that altered the genotype of a population before eliminating itself from the genome.
University of California, San Diego
Bier’s group previously exploited this mutation to develop a gene editing tool, called a gene drive, that reverted the mutated ion channel in a fruit fly model back to the wild type version.2 Gene drives eliminate or modify undesired genes in an animal with CRISPR, and proliferate through a population because the drive is designed to always copy itself back into the genome.
“The biggest problem of a gene drive—that's also its biggest strength—[is that] it can spread automatically through the whole population,” said Philipp Messer, a population geneticist at Cornell University who was not involved in the study. Once introduced into a population, existing gene drives cannot be eliminated and are hard to confine. “Can we really justify to genetically change the whole species?” Messer added.
Bier recognized this concern as well. To address this issue, he and his team developed a modified gene drive system that did not copy the Cas9 and guide RNA. In a study published in Nature Communications, the researchers showed that this modified drive, which they called an allele drive, reverted the majority of an insect population to the target genotype while gradually eliminating the gene editing elements.3 The system offers an opportunity to revert mutated genes in organisms back to wild type form, enabling researchers to study insecticide resistance without permanently inserting gene editing machinery.
In the first step to improving the gene drive system, Bier and his team inserted a guide RNA that recognized the allele mutant that they wanted to target into a pigmentation locus. In this location, the pigment gene is interrupted, which causes a mating disadvantage to male flies and helps to eliminate the allele drive over time. When flies inherit this system, the guide RNA recognizes the mutant allele in the ion channel gene, if it is present, and cuts it. If the DNA is repaired, then the wild type allele from the drive system is used as a reference.
The researchers determined that this system increased the presence of the wild type allele by directly converting the mutant amino acid to the wild type or by preventing the successful passage of mutation-carrying chromosomes. Since the CRISPR system was also not independently replicated, the researchers assessed how long the allele drive remained in the population and its efficiency in reverting the insects to the wild type genotype.
When the team introduced the allele drive at a one to three ratio into a population with normal pigmentation, they observed that this system reverted 80 percent of the population to be insecticide-sensitive after eight generations. Simultaneously, in this model, the researchers determined that the allele drive was gradually eliminated from the genome over eight to nine generations by screening for a fluorescence marker tagged to the allele drive.
Bier and his group want to explore transferring this system into mosquitoes and are interested in determining if it could be adapted to aid against other pests as well. “The hope is then that very soft touch would convince people that the whole approach is worth trying,” Bier said.
Due to the need to release many insects with the allele drive to be effective, Messer was concerned about the approach’s feasibility in a real-world setting. However, he does think it has potential in another context. “What's fantastic about this system is [that] this could actually work in field studies,” he said. Researchers could trial gene editing in natural populations and study its effects without the concern of permanently altering these organisms.
- Clarkson CS, et al. The genetic architecture of target-site resistance to pyrethroid insecticides in the African malaria vectors Anopheles gambiae and Anopheles coluzzii. Mol Ecol. 2021;30(21):5303-5317.
- Kaduskar B, et al. Reversing insecticide resistance with allelic-drive in Drosophila melanogaster. Nat Commun. 2022;13(1):291.
- Auradkar A, et al. A self-eliminating allelic-drive reverses insecticide resistance in Drosophila leaving no transgene in the population. Nat Commun. 2024;15(1):9961.
