Functional characterization of the E75 Gene in the Diamondback moth, Plutella xylostella and its role in insect pest management

Diamondback moth, Plutella xylostella, is a severe pest of Brassica crops, with a global impact of USD 5 billion/year. It rapidly developed resistance to insecticides and Bt formulations, necessitating sustainable alternatives like RNA interference (RNAi)-based biopesticides. Sprayable RNAi with encapsulated double-stranded RNA broadcast onto plants, offers integrated pest and resistance management options. Successful RNAi requires effective targets and efficient dsRNA delivery methods. To address this, AgroSpheres developed AgriCell technology, a proprietary sprayable insecticidal RNAi platform effective against numerous insect pests. We demonstrated that AgriCell-mediated sprayable RNAi effectively reduced the expression of target genes in a Bt-resistant strain of P. xylostella, causing significant mortality and global gene expression changes. Our RNA-sequencing analysis identified a potential target gene, Ecdysone-Induced Protein 75 (E75), upregulated in response to the AgriCell technology’s target gene suppression. E75 is a crucial regulator in the ecdysone-signaling pathway, controlling insect development and metamorphosis. E75 comprises three isoforms: E75A, E75B, and E75C, each playing distinct roles in molting, larval-pupal metamorphosis, and egg production in Drosophila melanogaster. However, the role of E75 in P. xylostella has remained elusive. Therefore, we characterized its role in P. xylostella through phylogenetic and RNAi-mediated knockdown studies. Our phylogenetic analysis clustered the identified E75 with the E75A isoform of holometabolous insects. Additionally, RNAi-mediated knockdown of E75 in P. xylostella blocked the larval-pupal transition, as they remained in the larval stage and eventually died. Our findings enhance the understanding of the developmental role of E75 and highlight its potential as a target for sustainable RNAi-mediated pest management.