dsRNA synergist targeting ABCB1 transporters sensitizes resistant Varroa destructor mites to amitraz
The ectoparasitic mite Varroa destructor is the primary biotic threat to managed honey bees, and its control relies on a limited number of chemical miticides. Amitraz remains widely used because of its efficacy and relatively low bee toxicity. However, resistance is increasingly reported, threatening its long-term utility. While mutations in the octopaminergic signaling pathway are strongly associated with amitraz resistance, additional mechanisms influencing toxicant uptake and efflux are likely to contribute. ATP-binding cassette (ABC) transporters, including ABCB1/P-glycoproteins, are well-established mediators of xenobiotic efflux and pesticide tolerance across arthropods. We evaluated a double-stranded RNA (dsRNA) synergist targeting V. destructor ABCB1 to test whether RNA interference (RNAi) mediated disruption of efflux pathways could enhance amitraz susceptibility. The full-length ABCB1 coding sequence was cloned, dsRNAs were screened, and mites were treated prior to amitraz exposure in laboratory bioassays. Mite transcriptomic responses to the dsRNA were monitored by RT-qPCR and RNA sequencing. Honey bee toxicity assays were conducted, including conservative tests feeding the dsRNA together with a known ABC transporter substrate.Mites exposed to ABCB1 dsRNA showed greater amitraz-induced mortality than those treated with non-specific dsRNA. This effect was observed across multiple trials and amitraz concentrations. RT-qPCR assays revealed increased ABCB1 transcript abundance following dsRNA treatment but transcriptomic analyses showed significant reduction in expression in the ABCB1 transcript, along with transcripts annotated as similar to a zinc finger protein and a chondroitin sulfate synthase. Chronic dietary exposure to ABCB1 dsRNA did not impact honey bee survival and toxicity assays incorporating verapamil (ABC transporter inhibitor) and acetamiprid (ABC transporter substrate) as a high-risk interaction control showed minimal adverse effects on bees. RNAi mediated perturbation of ABCB1 sensitizes V. destructor to amitraz and identifies transporter-mediated efflux as a modifiable component of amitraz tolerance. By increasing mite susceptibility, this approach may reduce selection pressure, slow resistance evolution, and extend the functional lifespan of amitraz. dsRNA synergists can provide a targeted resistance-management strategy that complements existing miticides while avoiding the nonselective toxicity associated with broad-spectrum transporter inhibitors.