A novel ACCase Trp-1999-Gly mutation confers resistance to ACCase-inhibiting herbicides in Chinese sprangletop (Leptochloa chinensis)

Chinese sprangletop ( Leptochloa chinensis (L.) Nees) is among the most economically damaging and disruptive weeds found in rice fields across China, and it exhibits a high degree of cyhalofop-butyl resistance for the past few years. Here, a suspected cyhalofop-butyl-resistant population HFCH5 (R) was collected from a rice field and analyzed to explore the mechanistic basis for its resistance activity. Compared with the susceptible HFLY1 (S) population, this R population exhibited differing levels of resistance to four ACCase-inhibiting herbicides (cyhalofop-butyl (RI = 11.52), fenoxaprop- P -ethyl (RI = 11.05), metamifop (RI = 6.61), and clethodim (RI = 4.06)), although it did exhibit susceptibility to florpyrauxifen-benzyl, tripyrasulfone, pyraclonil, and anilofos. This cyhalofop-butyl resistance in R plants was not effectively mitigated by pre-treatment with the cytochrome P450 inhibitor piperonyl butoxide or the glutathione S-transferase inhibitor 4-chloro-7-nitrobenzoxadiazole. Further analyses of these R plants revealed the substitution of Trp-1999 (TGG) for a Gly residue (GGG) within the ACCase carboxyltransferase domain. When the wild-type and mutated forms of this ACCase protein were subjected to structural modeling while complexed with a range of herbicides with ACCase-inhibiting activity, this Trp-1999-Gly substitution was found to be associated with a slight decrease in the affinity of this protein for a series of ACCase-inhibiting herbicides. A dCAPS marker was developed to aid in the rapid detection of this Trp (TGG)-1999-Gly (GGG) mutation within L. chinensis . Notably, before or after four ACCase-inhibiting herbicides (cyhalofop-butyl, fenoxaprop- P -ethyl, metamifop and clethodim) treatment, ACCase gene expression level of the R plants was not significantly higher (fold change <2, P  > 0.05) than that of the S plants. This study is the first report that in grass species, the newly identified ACCase Trp-1999-Gly mutation is associated with ACCase-inhibiting herbicide resistance.