A New Pro-197-Ile Mutation in Amaranthus palmeri Associated with Acetolactate Synthase-Inhibiting Herbicide Resistance

Abstract

Palmer amaranth (Amaranthus palmeri S. Watson), native to North America, is one of the most prominent invasive weed species on agricultural land. Acetolactate synthase (ALS)-resistant A. palmeri (Amaranthus palmeri) is widespread, while the research focus on resistance pattern and molecular basis of A. palmeri to imazethapyr is seldom documented in China. An A. palmeri population that survived the recommended rate of imazethapyr was collected in Shandong Province, China. The resistant mechanism and pattern of A. palmeri to imazethapyr was investigated. Dose-response assay showed that the resistant (R) population displayed a high resistance level (292.5-fold) to imazethapyr compared with the susceptible (S) population. Sequence analysis of the ALS gene revealed that nucleotide mutations resulted in three resistance-conferring amino acid substitutions, Pro-197-Ile, Trp-574-Leu, and Ser-653-Asp, in the individual plants of the R population. An in vitro enzyme assay indicated that the ALS was relatively unsusceptible to imazethapyr in the R population, showing a resistance index of 88.6-fold. ALS gene expression and copy number did not confer resistance to imazethapyr in the R population. Pro-197-Ile is the first reported amino acid substitution conferring ALS resistance to A. palmeri. This is the first case of an imazethapyr-resistant A. palmeri biotype in China.

Keywords: ALS gene copy number; ALS gene expression; ALS gene mutation; Amaranthus palmeri; imazethapyr; resistance.

Figure 1

Dry shoot mass reduction in the susceptible (S) and the resistant Amaranthus palmeri population (R) at 21 d after treatment with imazethapyr in two growth chamber experiments. Results were pooled over experimental runs. Vertical bars represent the standard error of the mean (n = 6).

Figure 2

Target-site mutations in the ALS gene conferring herbicide resistance in the R population: (a) Pro-197-Ile (CCC mutated to ATC), (b) Trp-574-Leu (TGG mutated to TTG), and (c) Ser-653-Asp (AGC mutated to AAC) in the ALS gene sequence.

Figure 3

In vitro ALS activity assays in response to imazethapyr were performed using protein extracts of plants derived from the resistant A. palmeri (Amaranthus palmeri) population (●) and the susceptible population (○). Results were pooled over experimental runs. Vertical bars represent the standard error of the mean (n = 3).

Figure 4

ALS copy number relative to A36 for the S and R populations; ALS gene expression relative to A36 for the S and R populations not treated with imazethapyr. Vertical bars represent the mean ± standard deviation; ns represents non-significant.