Resistance to cyhalofop-butyl and metamifop in Echinochloa crus galli var. mitis conferred by the Cys-2088-Arg mutation in ACCase gene and GST-mediated enhanced metabolism

Echinochloa crus-galli var. mitis (ECM) is one of the major Echinochloa species in the rice field, with increasing reports of herbicide resistance. This study identified an ECM population (ECM_2312) exhibiting cross-resistance to four acetyl-CoA carboxylase (ACCase)-inhibiting herbicides, including cyhalofop-butyl (8.7-fold), metamifop (15.8-fold), clodinafop-propargyl (16.1-fold) and fenoxaprop-P-ethyl (25.0-fold)，compared with the susceptible population ECM_2201. A Cys-2088-Arg mutation was detected in ACCase2C gene copy in ECM_2312, confirming the involvement of that target-site resistance. Liquid chromatography–tandem mass spectrometry (HPLC-MS) analysis revealed that the degradation of the herbicides active forms (cyhalofop-acid and metamifop) were faster in the resistant population than in the susceptible population. Pretreatment with the glutathione S-transferase (GST) inhibitor, 4-chloro-7-nitrobenzoxadiazole (NBD-Cl) reduced the resistance level from 8.7-fold to 4.9 fold for cyhalofop-butyl and from 15.8-fold to 3.0-fold for metamifop. Moreover, GST activity was consistently higher in the resistant population than the susceptible population, both in the absence and presence of herbicide treatment. Furthermore, five GST genes exhibited sustained overexpression for 96 h in the resistant plants following exposure to either cyhalofop-butyl or metamifop. This study demonstrated an ECM population possesses both a target-site mutation (Cys-2088-Arg) and GST-mediated enhanced metabolic resistance to ACCase-inhibiting herbicides.