Identification of insecticidal double-stranded RNAs targeting the proteasome pathway in the hop flea beetle Psylliodes attenuata using dsRIP

The hop flea beetle, Psylliodes attenuata , is a major pest of hop ( Humulus lupulus ), an important crop for the brewing industry. Restrictions on chemical insecticides and insecticide resistance create a need for alternative control strategies. RNA interference (RNAi) uses double-stranded RNA (dsRNA) to silence essential pest genes and cause insecticidal effects. In this study, we aimed to establish P. attenuata as an RNAi-based pest control target. We generated a transcriptome assembly using a public RNA sequencing dataset and used the dsRIP web platform to identify essential gene targets and design effective dsRNAs. Three dsRNAs targeting proteasome subunits were tested in feeding bioassays by applying them to hop leaf disks. The treatments caused significant mortality of up to 64% and feeding inhibition of up to 45%. Sequencing of small RNAs bound to the RNA-induced silencing complex identified predominantly 21-nucleotide small interfering RNAs produced from the orally delivered dsRNAs, supporting activation of the RNAi pathway. Additional RNA sequencing of adults enabled assembly of a higher-quality transcriptome with 89% BUSCO completeness for future RNAi studies. Overall, the feeding bioassays and molecular resources establish P. attenuata as a promising target for RNAi-based pest control and demonstrate the usefulness of dsRIP for RNAi research in insects previously unstudied at the molecular level.