From midgut targets to genomic surveillance: Mechanism-informed diagnostics of resistance to Bt crops

Bacillus thuringiensis (Bt) maize and cotton have reduced pest pressure and insecticide use in major cropping systems, but field-evolved resistance in multiple pest species threatens their long-term durability. Managing resistance increasingly requires more than detecting reduced susceptibility: it also requires understanding which biological step has changed, because different mechanisms imply different risks of cross-resistance and different options for follow-up monitoring. In this review, we organize resistance to Cry and Vip traits around the major steps of intoxication, including toxin activation and processing, receptor- and transporter-related intoxication, and broader physiological or regulatory changes affecting midgut susceptibility. We then compare the main diagnostic approaches used in Bt resistance studies, from bioassays and toxin-panel tests to binding or processing assays, targeted genotyping, and population genomics, and we summarize what each approach can and cannot infer on its own. Building on this comparison, we propose an evidence-to-action workflow that links signal confirmation, breadth mapping, mechanism inference, and the threshold for marker deployment in surveillance. To improve comparability across studies, we also indicate a minimum set of reporting items to support comparability and introduce practical frameworks for grading evidence strength and judging marker readiness for surveillance, with particular attention to Vip traits, for which validated markers remain limited. Finally, we discuss how mechanistic evidence can support resistance monitoring and stewardship, and where emerging multi-omic and functional approaches may strengthen future diagnostics.