Evolution of target-based resistance mechanism to acetyl-coenzyme A carboxylase herbicides in Digitaria ciliaris

Abstract

Digitaria ciliaris (Retz.) Koeler (southern crabgrass) is an annual grass weed of the southern United States, commonly infests all crops, non-crops, and even in turf. This grass is historically controlled in both crop and turf systems for decades by employing postemergence ACCase-targeting herbicides. Resistance to these herbicides is common and continues to develop. D. ciliaris has developed resistance to ACCase targeting herbicides. ACCase resistant D. ciliaris was recently identified in sod production fields in Georgia. To date, no study has been reported on the possible resistant mechanism for D. ciliaris to the ACCase targeting herbicides. The primary goal of this dissertation research is the determination of the ACCase resistance mechanism in D. ciliaris. The dissertation, therefore, starts with a short literature review concerning the motivation for this research. In chapter 2 we report on the cross-resistance to pinoxaden herbicide in D. ciliaris from the resistant biotypes to sethoxydim and select aryloxyphenoxypropionate (FOPs) herbicides using pinoxaden response evaluation in the greenhouse. Prior selection pressure with other ACCase herbicides could evolve cross-resistance to pinoxaden of D. ciliaris in the United States. In addition, the target-based resistance, Ile-1781-Leu amino acid substitution in the ACCase gene is one of the causal mechanisms of resistance in D. ciliaris determined by amplification of the plastidic ACCase gene using a standard PCR and Next-Generation Sequencing profile. Chapter 3 we report on the detection of ACCase enzyme activity in the resistant and susceptible biotypes for the validation of target-based resistance to ACCase herbicides from the malachite green colorimetric functional assay. Greater ACCase enzyme activity in the resistant biotypes to ACCase-targeting herbicides, sethoxydim, clethodim, fluazifop-p-butyl, and pinoxaden conferred the target-based resistance to the resistant biotypes. In addition, the malachite green functional assay can be used for the measuring of the target-site activity in suspected resistance to ACCase herbicides instead of the 14C-based radiometric assay. Chapter 4 presents the elucidation of the differential ACCase enzyme inhibition mechanisms in the two resistant biotypes by using the gene expression profile. ACCase gene expression in the R2 biotype was the responsible factor for the inhibition differentiation of the ACCase enzyme from the R1 and S biotypes. Finally, Chapter 5 presents a study on the evaluation of the three different bioassay methods assessing the detection of ACCase-targeting herbicides resistance in D. ciliaris. Resistant biotypes were separated from the susceptible with different parameters from the three different bioassays such as agar-based gel box assay, leaf flotation assay, and electrical conductivity assay. Herbicide sensitivity for the suspected resistance population can be primarily screened using these rapid bioassays. The results derived from our research would facilitate increasing our understanding of resistance mechanisms along with genetic variation in the D. ciliaris genome.