Glufosinate resistance in Palmer amaranth (Amaranthus palmeri S. Watson) was recently detected in three accessions from Arkansas, USA. Amaranthus palmeri is the first and only broadleaf weed species resistant to this herbicide, and the resistance mechanism is still unclear. A previous study characterized the glufosinate resistance level in the accessions from Arkansas. A highly glufosinate-resistant accession was further used to investigate the mechanism conferring glufosinate resistance in A. palmeri. Experiments were designed to sequence the herbicide target enzyme cytosolic and chloroplastic glutamine synthetase isoforms (GS1 and GS2, respectively) and quantify copy number and expression. Absorption, translocation, and metabolism of glufosinate using the ¹⁴C-labeled herbicide were also evaluated in the resistant and susceptible accessions. The glufosinate-resistant accession had an increase in copy number and expression of GS2 compared with susceptible plants. All accessions showed only one GS1 copy and no differences in expression. No mutations were identified in GS1 or GS2. Absorption (54% to 60%) and metabolism (13% to 21%) were not different between the glufosinate-resistant and glufosinate-susceptible accessions. Most residues of glufosinate (94% to 98%) were present in the treated leaf. Glufosinate translocation to tissues above the treated leaf and in the roots was not different among accessions. However, glufosinate translocation to tissues below the treated leaf (not including roots) was greater in the resistant A. palmeri (2%) compared with the susceptible (less than 1%) accessions. The findings of this paper strongly indicate that gene amplification and increased expression of the chloroplastic glutamine synthetase enzyme are the mechanisms conferring glufosinate resistance in the A. palmeri accession investigated. Thus far, no additional resistance mechanism was observed, but further investigations are ongoing.

Some herbicide-resistant weeds become resistant by generating additional copies of specific loci. For example, amplification of the locus encoding chloroplastic glutamine synthetase (GS2) produces herbicide resistance in the glufosinate-resistant Palmer amaranth (Amaranthus palmeri) accession MSR2. Previously, overamplification of the glyphosate-resistant gene encoding 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) in Palmer amaranth was determined to be driven by an extrachromosomal circular DNA (eccDNA). Here, we describe a rearranged eccDNA that confers resistance to both glyphosate and glufosinate ammonium due to the coduplication of the native chromosomal regions that contain the genes that encode for these herbicides target proteins. In addition to EPSPS, the replicon carries 2 GS2 isoforms (GS2.1 and GS2.2) and other genes. MSR2 samples harbored eccDNA carrying only EPSPS coexisting with eccDNAs harboring both EPSPS and GS2. A second glufosinate-resistant Palmer amaranth accession (MSR1) showed distinct GS2.1 and GS2.2 amplification patterns from MSR2, suggesting the existence of diverse replicons in Palmer amaranth. EPSPS copy number was correlated with both GS2 isoforms copy number in MSR2, further supporting the coexistence of these genes in the same replicon. These findings shed light on the complexity of eccDNA formation in plant systems, with the collection and accumulation of extra pieces of DNA.

In Arkansas, resistance to protoporphyrinogen IX oxidase (PPO)-inhibiting herbicides in Amaranthus palmeri S. Wats. is mainly due to target site mutations. Although A. palmeri PPO-mutations are well investigated, the cross-resistance that each ppo mutant endows to weed populations is not yet well understood. We aimed to evaluate the response of PPO-resistant A. palmeri accessions, harboring the ppo2 mutations ΔG210 and G399A, to multiple PPO-inhibiting herbicides. Six resistant and one susceptible field accessions were subjected to a dose–response assay with fomesafen, and selected survivors from different fomesafen doses were genotyped to characterize the mutation profile. The level of resistance to fomesafen was determined and a cross-resistance assay was conducted with 1 and 2 times the labeled doses of selected PPO herbicides. The accession with higher predicted dose to control 50% of the population (ED50) had a higher frequency of ΔG210-homozygous survivors. Survivors harboring both mutations, and those that were ΔG210-homozygous, incurred less injury at the highest fomesafen rate tested (1120 g ai ha⁻¹). The populations with a high frequency of ΔG210-homozygous survivors, and those with individuals harboring ΔG210 + G399A mutations, exhibited high potential for cross-resistance to other PPO herbicides. The new PPO–herbicide chemistries (saflufenacil, trifludimoxazin) generally controlled the PPO-resistant populations.

Echinochloa colona (junglerice) is a problematic global weed for many crops, primarily controlled with herbicides. Drought stress alters the overall plant physiology and reduces herbicide efficacy. This research aimed to study the joint effect of drought stress (DS) and recurrent selection with sublethal dose of herbicide on adaptive gene expression and herbicide efficacy on E. colona. Three factors were evaluated: (A) E. colona generation (G0, original population from susceptible standard; G1 and G2, progenies of recurrent selection); (B) herbicide treatment (florpyrauxifen-benzyl, 0.25×; glyphosate, 0.125×; quinclorac, 0.125× the recommended dose; and nontreated check); (C) DS (50% and 100% field capacity). Recurrent exposure to sublethal herbicide dose, combined with drought stress, favors the selection of plants less susceptible to the herbicide. Upregulation of defense (antioxidant) genes (APX: ascorbate peroxidase), herbicide detoxification genes (CYP450 family: cytochrome P450), stress acclimation genes (HSP: heat-shock protein, TPP: trehalose phosphate phosphatase, and TPS: trehalose phosphate synthase), and genes related to herbicide conjugation (UGT: UDP glucosyltransferase) in the G2 population was significant. Recurrent exposure to sublethal herbicide dose under drought stress reduces junglerice sensitivity to herbicide, seemingly due to “imprinted” upregulation of metabolic and protection genes in response to these stresses.

Global climate change, specifically rising temperature, can alter the molecular physiology of weedy plants. These changes affect herbicide efficacy and weed management. This research aimed to investigate the combined effect of heat stress (HS) and sublethal doses of herbicides (four active ingredients) on adaptive gene expression and efficacy of herbicide on Echinochloa colona (L.) Link (junglerice). Three factors were evaluated; factor A was E. colona generation (G0-original population from susceptible standard; G1 and G2 were progenies of recurrent selection), factor B was herbicide treatment (florpyrauxifen-benzyl, glufosinate-ammonium, imazethapyr, quinclorac and nontreated check) and factor C was HS (30 and 45 °C). The herbicides were applied at 0.125× the recommended dose. Recurrent exposure to HS, combined with sublethal doses of herbicides, favors the selection of plants less susceptible to the herbicide. Upregulation of defense (antioxidant) genes (APX: Ascorbate peroxidase), herbicide detoxification genes (CYP450 family: Cytochrome P450), stress acclimation genes (HSP: Heat shock protein, TPP: Trehalose phosphate phosphatase and TPS: Trehalose phosphate synthase) and genes related to herbicide conjugation (UGT: UDP Glucosyltransferase) was significant. The positive regulation of these genes may promote increased tolerance of E. colona to these herbicides.