Identification of broadly expressed nervous system-related genes as effective RNAi targets in the brown marmorated stink bug, Halyomorpha halys.

Abstract

Background: The brown marmorated stink bug, Halyomorpha halys (BMSB), is an invasive polyphagous pest causing major losses in fruit and field crops in Europe and the Americas. With insecticide options increasingly limited by resistance and regulatory restrictions, RNA interference (RNAi) offers a species-specific alternative. As RNAi is fully functional in BMSB following hemolymph injection, we evaluated neuronal and neuron-associated genes as systemic RNAi targets using dsRNA microinjection.

Results: A literature screen of >80 insect RNAi targets identified 12 neuronal/synaptic genes, from which five were selected in BMSB: acetylcholinesterase (HhAChE), alpha-soluble NSF attachment protein (HhAsnap), Shaker (HhSh), tyrosine hydroxylase (HhTh) and Ras opposite protein (HhRop). Previously published transcriptomic data showed HhAChE, HhSh and HhTh were enriched in brain/central nervous system, whereas HhAsnap and HhRop were broadly expressed at higher abundance. Adults injected with dsRNA (334-364 bp; 2 μL, 1000 ng/μL) targeting HhAsnap or HhRop showed strong mortality: 96.8% (30/31) and 100% (34/34) by Day (D)13, respectively (log-rank, P < 0.0001). By contrast, there was no significant difference between insects injected with dsRNA targeting HhAche, HhSh and HhTh and the negative controls. Feeding assays showed >60% reductions in salivary sheath formation from D4 to D8 after dsAsnap or dsRop treatment. Quantitative real-time (qRT)-PCR at 72 h confirmed ~58% knockdown of HhRop, whereas HhAChE and HhAsnap showed no significant whole-body depletion.

Conclusion: Broadly and highly expressed targets (HhAsnap, HhRop) were associated with strong mortality and feeding suppression, whereas brain-enriched targets did not. Expression breadth and transcript abundance appear critical for RNAi responsiveness in BMSB, identifying HhAsnap and HhRop as promising candidates for oral or spray-based RNAi control. © 2026 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.