Plastidic ACCase Ile-1781-Leu is present in pinoxaden-resistant southern crabgrass (Digitaria ciliaris)

Abstract

Southern crabgrass [Digitaria ciliaris (Retz.) Koeler] is an annual grass weed that commonly infests turfgrass, roadsides, wastelands, and cropping systems throughout the southeastern United States. Two biotypes of D. ciliaris (R1 and R2) with known resistance to cyclohexanediones (DIMs) and aryloxyphenoxypropionates (FOPs) previously collected from sod production fields in Georgia were compared with a separate susceptible biotype (S) collected from Alabama for the responses to pinoxaden and to explore the possible mechanisms of resistance. Increasing rates of pinoxaden (0.1 to 23.5 kg ha−1) were evaluated for control of R1, R2, and S. The resistant biotypes, R1 and R2, were resistant to pinoxaden relative to S. The S biotype was completely controlled at rates of 11.8 and 23.5 kg ha−1, resulting in no aboveground biomass at 14 d after treatment. Pinoxaden rates at which tiller length and aboveground biomass would be reduced 50% (I50) and 90% (I90) for R1, R2, and S ranged from 7.2 to 13.2 kg ha−1, 6.9 to 8.6 kg ha−1, and 0.7 to 2.1 kg ha−1, respectively, for tiller length, and 7.7 to 10.2 kg ha−1, 7.2 to 7.9 kg ha−1, and 1.6 to 2.3 kg ha−1, respectively, for aboveground biomass. Prior selection pressure from DIM and FOP herbicides could result in the evolution of D. ciliaris cross-resistance to pinoxaden herbicides. Amplification of the carboxyl-transferase domain of the plastidic ACCase by standard PCR identified a point mutation resulting in an Ile-1781-Leu amino acid substitution only for the resistant biotype, R1. Further cloning of PCR product surrounding the 1781 region yielded two distinct ACCase gene sequences, Ile-1781 and Leu-1781. The amino acid substitution, Ile-1781-Leu in both resistant biotypes (R1 and R2), however, was revealed by next-generation sequencing of RNA using Illumina platform. A point mutation in the Ile-1781 codon leading to herbicide insensitivity in the ACCase enzyme has been previously reported in other grass species. Our research confirms that the Ile-1781-Leu substitution is present in pinoxaden-resistant D. ciliaris.

Diversity of α‐tubulin transcripts in Lolium rigidum

“…Both traditional Sanger sequencing from PCR and next generation sequencing are important for understanding the plant gene family. Previously, researchers have found that PCR‐based amplification may not be sufficient to reveal all target‐site resistance mutations 31, 32 . Here, we demonstrated that it is equally important to validate sequencing data from RNA‐seq by standard PCR, so that: (i) computational errors could be checked and corrected; and (ii) the nucleotide diversity of different individuals/populations, not limited to the few plants with transcriptome data, can be revealed.…”

Characterization of mutations conferring inherent resistance to acetyl coenzyme A carboxylase‐inhibiting herbicides in turfgrass and grassy weeds

“…Basak et al. (2020) also reported difficulty with PCR‐based sequencing of the ACCase CT domain coding region in southern crabgrass. The application of more advanced approaches such as next generation sequencing may be required for these species.…”

Detecting the effect of ACCase-targeting herbicides on ACCase activity utilizing a malachite green colorimetric functional assay

“…Previously collected seeds of two resistant biotypes (R1 and R2) of D. ciliaris with confirmed resistance to select ACCase-targeting herbicides and one susceptible (S) biotype were included in this study (Basak et al 2019;Yu et al 2017). To generate green plant material for enzyme extraction, seeds of resistant and susceptible D. ciliaris biotypes were sown in separate plastic flats containing commercial potting soil and peat moss (2:1 v/v).…”

“…This result can only be because of differential interaction with the ACCase substrate and the tested ACCase-targeting herbicide, as no absorption, translocation, or metabolism is at play as would be the case when screening whole plants. The ACCase carboxyl transferase domains were sequenced and reported for S, R1, and R2 previously (Basak et al 2019). No other amino acid substitutions except Ile-1781-Leu were observed between R1 or R2 that would explain the difference between these biotypes (Supplementary Data 2).…”

“…Koeler] were used as a model for evaluating the effectiveness of the malachite green assay in ACCase-resistance detection. Previous research quantified the TSR mechanism and rate response screen to these biotypes (Basak et al 2019;Yu et al 2017).…”