CRISPR‐mediated gene knockout reveals nicotinic acetylcholine receptor (nAChR) subunit α6 as a target of spinosyns in Helicoverpa armigera

BACKGROUND: The spinosyn insecticides (spinosad and spinetoram) have been intensively used to control a wide range of agricultural pests. However, resistance to spinosyns has been evolved in several agricultural pests. Disruption of the nicotinic acetylcholine receptor subunit α6 (nAChRα6) has been associated with high levels of resistance to spinosyns in both field and laboratory-selected strains of several insect pests. Among the twelve nAChR subunits of Helicoverpa armigera, Haα6 has the closest sequence similarity (66.02%) to Haα7. Here we used CRISPR-mediated knockouts to evaluate the role of two nAChR subunits (Haα6 and Haα7) of H. armigera in toxicity of spinosyns. RESULTS: Individual knockouts of Haα6 and Haα7 were created utilizing CRISPR/Cas9 system in H. armigera. The Haα6 knockout strain (Haα6-KO) exhibited high levels of resistance to spinosad (531-fold) and spinetoram (1105-fold) compared with the wild-type parent SCD strain, whereas the Haα7 knockout strain (Haα7-KO) showed no significant susceptibility changes to both spinosyns. Genetic analyses demonstrated that resistance to spinosad conferred by knockout of Haα6 was autosomal, incompletely recessive and tightly linked to the disruption mutation of Haα6. Both Haα6-KO and Haα7-KO strains had no significant effects on susceptibility to other four insecticides including emamectin benzoate, beta-cypermethrin, chlorantraniliprole and indoxacarb. CONCLUSION: Our results provide in vivo functional evidence for Haα6 as a target of spinosyns in H. armigera, and little or no role of Haα7 in mediating toxicity of spinosyns. The results are valuable to the development of resistance monitoring and management methods for spinosyn resistance in H. armigera. This article is protected by copyright. All rights reserved.