OBP11-mediated sequestration reduces chlorpyrifos bioavailability and toxicity in Spodoptera exigua

Odorant-binding proteins (OBPs) are traditionally known for olfactory functions, but emerging evidence suggests their involvement in insecticide transport and resistance. Here, we systematically analyzed 36 OBP genes in the global agricultural pest Spodoptera exigua, examining their tissue-specific expression, differential regulation in a multi-resistant field strain, and binding affinities to 34 insecticides. Multiple OBPs showed prominent expression in hemolymph, fat body, and midgut, indicating non-olfactory functions. Notably, SeOBP11 was upregulated >20-fold in resistant larvae despite modest baseline expression. Fluorescence competitive binding assays revealed that six recombinant SeOBPs exhibited highly selective and unpredictable interaction patterns with insecticides-chlorpyrifos bound all six SeOBPs with high affinity (Ki = 5.0-15.2 μM), while 20 of 34 insecticides showed no detectable binding to any tested OBP. Molecular docking confirmed that chlorpyrifos, thiodicarb, and dimethoate compete for the same OBP11 binding cavity, with hydrophobic interactions dominating high-affinity complexes. Crucially, co-injection of SeOBP11 with chlorpyrifos significantly reduced in vivo toxicity (mortality decreased from ∼80% to ∼40%), demonstrating that OBP binding sequesters insecticide, reducing bioavailability. This sequestration-based resistance mechanism is distinct from canonical metabolic, target-site, or penetration resistance. Our findings establish OBPs as multifunctional carriers contributing directly to insecticide resistance in S. exigua, provide a mechanistic basis for field-observed multi-resistance, and identify SeOBP11 as a potential molecular marker for resistance monitoring.