RNAi-mediated gene silencing reduces survival and Xylella fastidiosa load in the insect vector Philaenus spumarius

Abstract The spittlebug Philaenus spumarius (L.) (Hemiptera: Aphrophoridae) is the main epidemiologically relevant vector of the xylem-limited bacterium Xylella fastidiosa (Lysobacterales (Xanthomonadales): Xanthomonas ) in all the European outbreaks described so far. Current vector control strategies yield inconsistent results in terms of reduction of bacterial spread, and alternative control tools are urgently needed. RNA interference (RNAi) is a post-transcriptional gene silencing mechanism that can be triggered by administering double-stranded RNA (dsRNA) molecules to a target organism. Exogenous application of dsRNAs to crops has emerged as a powerful and sustainable tool in agriculture to control insect pests. Here, we evaluated survival probability and transcript reduction upon injection of spittlebug adults with dsRNAs targeting six candidate essential genes. Thereafter, we evaluated acquisition and inoculation efficiency as well as bacterial load following silencing of proton ATPase (ATPase) and Transient Receptor Potential mucolipin-3like isoform X1 (TRPML). The administration of dsATPase caused the highest mortality among the target genes tested, 83 and 96% at 5 and 10 days post injection, respectively. Micro-injection of dsATPase into insects prior to X. fastidiosa transmission experiments showed no change in transmission rates, but the insects harboured significantly lower bacterial populations than the control group, suggesting a possible alteration of feeding and fluid dynamics within the vector foregut affecting bacterial growth. Although delivering dsRNAs to sap-sucking insects is challenging, the development of RNAi-based tools paves the way for the control of vector insects and the pathogens they transmit.