Enantioselective Bioactivity and Molecular Basis of Fluralaner Enantiomers in <i>Tetranychus cinnabarinus</i> (Boisduval)

Fluralaner, a chiral isoxazoline compound, exhibits pronounced enantioselective toxicity against <i>Tetranychus cinnabarinus</i>, a significant agricultural mite. In this study, the <i>S</i>-(+)-fluralaner enantiomer exhibited over 75-fold higher adulticidal activity and more than 225-fold stronger inhibition of egg hatching compared to the <i>R</i>-(-)-enantiomer. Enzymatic degradation assays indicated that the <i>R</i>-form was preferentially metabolized by mite enzymes. Transcriptomic profiling and biochemical validation identified cytochrome P450 monooxygenases as key contributors to this enantioselective detoxification. Electrophysiological recordings in <i>Xenopus laevis</i> oocytes expressing the GABA receptor subunit <i>TcRDL2</i> revealed enantioselective inhibition of GABA induced currents, with <i>S</i>-(+)-fluralaner exhibiting the greatest antagonistic effect. Structural modeling further demonstrated distinct hydrogen bonding patterns between each enantiomer and <i>TcRDL2</i>, providing a mechanistic explanation for their differential activity. These results suggest that both metabolic and target selectivity contribute to the superior bioactivity of <i>S</i>-(+)-fluralaner, supporting its potential as a enantiomer-specific acaricide and informing the rational design of next-generation chiral pesticides.