A biotype of Eleusine indica L. Gaertn. from Malaysia, which had a field history of two or three applications of fluazifop-p-butyl per year over 4-5 years, was previously found to show 100-fold resistance to the herbicide under field conditions compared with a susceptible population. This biotype is cross-resistant to other aryloxyphenoxypropanoate and cyclohexanedione herbicides. Previous research has shown no evidence for differential uptake, translocation, or metabolism of fluazifop-butyl between the R and S biotypes. Fluazifop acid reduced [14C]acetate incorporation into the lipid fraction of leaf disks of the S biotype but not of the R biotype. Acetyl coenzyme A carboxylase (ACCase) from the S biotype was sensitive to fluazifop, fenoxaprop, sethoxydim, and clethodim, with I50 values ranging from 1.0 to 5.6 μM. In contrast, ACCase from the R biotype was much less sensitive to fluazifop, fenoxaprop, and sethoxydim, with I50 values of >500, 25, and 77 μM, respectively. ACCase from the R biotype showed only a low level of resistance to clethodim, with an I50 of 6.6 μM. The close correlation between the whole plant and ACCase sensitivities to various ACCase inhibitors suggests that herbicide resistance in this biotype is conferred by a mutation to the herbicide target site, ACCase.