Amarish K. Yadav, Cole Butler, Akihiko Yamamoto, +2 , and Maxwell J. Scott

PNAS June 12, 2023; 120 (25) e2301525120

https://doi.org/10.1073/pnas.2301525120

Significance

Spotted wing drosophila (Drosophila suzukii) is a vinegar fly with a worldwide distribution that does significant economic damage to soft-skinned fruits. We made and evaluated split CRISPR homing gene drives that target the conserved female-specific exon of the doublesex gene, which is essential for sexual development in Drosophila. Our results suggest that homing gene drives could provide a cost-effective approach for suppression of D. suzukii populations.

Abstract

Genetic-based methods offer environmentally friendly species-specific approaches for control of insect pests. One method, CRISPR homing gene drive that target genes essential for development, could provide very efficient and cost-effective control. While significant progress has been made in developing homing gene drives for mosquito disease vectors, little progress has been made with agricultural insect pests. Here, we report the development and evaluation of split homing drives that target the doublesex (dsx) gene in Drosophila suzukii, an invasive pest of soft-skinned fruits. The drive component, consisting of dsx single guide RNA and DsRed genes, was introduced into the female-specific exon of dsx, which is essential for function in females but not males. However, in most strains, hemizygous females were sterile and produced the male dsx transcript. With a modified homing drive that included an optimal splice acceptor site, hemizygous females from each of the four independent lines were fertile. High transmission rates of the DsRed gene (94 to 99%) were observed with a line that expressed Cas9 with two nuclear localization sequences from the D. suzukii nanos promoter. Mutant alleles of dsx with small in-frame deletions near the Cas9 cut site were not functional and thus would not provide resistance to drive. Finally, mathematical modeling showed that the strains could be used for suppression of lab cage populations of D. suzukii with repeated releases at relatively low release ratios (1:4). Our results indicate that the split CRISPR homing gene drive strains could potentially provide an effective means for control of D. suzukii populations.

See https://www.pnas.org/doi/10.1073/pnas.2301525120