Molecular insights into bifenazate resistance in two-spotted spider mite populations on hops

Two-Spotted Spider Mites (TSSM) are cosmopolitan pests that feed on more than 1,100 different plant species, including hops—an essential crop in beer production. Hops (Humulus lupulus) provide bitterness, aroma, and preservative qualities to beer. In the Pacific Northwest (PNW), which accounts for approximately 99% of U.S. hop production, TSSM has emerged as a major threat to hop production. To date, acaricide application remains the primary method for managing TSSM infestations. However, repeated and prolonged use of acaricides has led to the rapid development of resistance and tolerance in TSSM populations. We performed RNA sequencing to investigate the molecular mechanisms underlying bifenazate resistance in TSSM affecting hops. A susceptible TSSM population was collected from a hops field in Prosser, Washington, and subjected to selection pressure with bifenazate to generate resistant populations in the lab. Genes that are involved in several biological pathways related to metabolic processes, including genes encoding cytochrome P450s, carboxylesterases, lipocalin, and peptidases, were identified by gene ontology enrichment analysis. Although no differential expression was observed for glutathione S-transferases, UDP-glycosyltransferases, or ABC transporter genes, mutations identified in their active sites in resistant populations suggest that altered enzyme function may contribute to acaricide resistance through modified substrate metabolism. Additionally, RNA sequencing reads were used to study mitochondrial gene expression and identify mutations potentially associated with bifenazate resistance in TSSM. These findings help us understand the molecular mechanisms underlying bifenazate resistance and support the development of molecular tools to detect and monitor acaricide resistance in hop fields.