Following 15 yr of successful use, glyphosate failed to control a population of the widespread grass weed rigid ryegrass in Australia. This population proved to be resistant to glyphosate in pot dose-response experiments conducted outdoors, exhibiting 7- to 11-fold resistance when compared to a susceptible population. Some cross-resistance to diclofop-methyl (about 2.5-fold) was also observed. Similar levels of control of the resistant and susceptible populations were obtained following application of amitrole, chlorsulfuron, fluazifop-P-butyl, paraquat, sethoxydim, sirnazine, or tralkoxydim. The presence of glyphosate resistance in a major weed species indicates a need for changes in glyphosate use patterns.

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Glyphosate, the most commonly used herbicide in the world, controls a wide range of plant species, mainly because plants have little capacity to metabolize (detoxify) glyphosate. Massive glyphosate use has led to world-wide evolution of glyphosateresistant (GR) weed species, including the economically damaging grass weed Echinochloa colona. An Australian population of E. colona has evolved resistance to glyphosate with unknown mechanisms that do not involve the glyphosate target enzyme 5enolpyruvylshikimate-3-P synthase. GR and glyphosate-susceptible (S) lines were isolated from this population and used for resistance gene discovery. RNA sequencing analysis and phenotype/genotype validation experiments revealed that one aldoketo reductase (AKR) contig had higher expression and higher resultant AKR activity in GR than S plants. Two full-length AKR (EcAKR4-1 and EcAKR4-2) complementary DNA transcripts were cloned with identical sequences between the GR and S plants but were upregulated in the GR plants. Rice (Oryza sativa) calli and seedlings overexpressing EcAKR4-1 and displaying increased AKR activity were resistant to glyphosate. EcAKR4-1 expressed in Escherichia coli can metabolize glyphosate to produce aminomethylphosphonic acid and glyoxylate. Consistent with these results, GR E. colona plants exhibited enhanced capacity for detoxifying glyphosate into aminomethylphosphonic acid and glyoxylate. Structural modeling predicted that glyphosate binds to EcAKR4-1 for oxidation, and metabolomics analysis of EcAKR4-1 transgenic rice seedlings revealed possible redox pathways involved in glyphosate metabolism. Our study provides direct experimental evidence of the evolution of a plant AKR that metabolizes glyphosate and thereby confers glyphosate resistance.

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The development of herbicide multiple‐resistance in weed species represents a major threat to current agricultural practices. The mechanistic basis for herbicide multiple‐resistance has been investigated in a population of the annual grass weed Lolium rigidum Gaud. (annual ryegrass) resistant to herbicides affecting 6 target sites. A subset of the resistant population (R2 subset) has been isolated by germination on a medium containing the acetyl‐CoA carboxylase (ACCase, EC 6.4.1.2) inhibiting herbicide, sethoxydim ((2‐[1‐(ethoxyimino)butyl]‐5‐[2‐(ethylthio)propyl]‐3‐hydroxy‐2‐cyclohexen‐1‐one)). This 12% R2 subset of the population is 600 times more resistant to sethoxydim and between 30 to 200 times more resistant to other ACCase inhibitors than the bulk of the R population. The subset has a form of ACCase which is 6 to 55 times less sensitive to inhibition by these herbicides than the enzyme present in the bulk of the resistant or in the susceptible population. There was no difference in the uptake and metabolic degradation of [4‐14C]sethoxydim between the R2 subset and the unselected R population. These results show the accumulation of different resistance mechanisms in that single population. Furthermore we propose that this accumulation of multiple resistance mechanisms is the basis for herbicide multiple‐resistance in this biotype.

Thirteen biotypes of rigid ryegrass were screened for trifluralin resistance. From these, the two most resistant biotypes, SLR 31 and SLR 10, were chosen for further studies involving exposure to other dinitroanilines, mitosis-inhibiting herbicides and14C-trifluralin. SLR 31, and SLR 10 exhibited an approximate 10-fold reduced sensitivity to trifluralin in comparison to susceptible biotypes. Resistance to five other dinitroaniline herbicides was observed, with reduced sensitivity varying from 32-fold for ethalfluralin to 2.5-fold for isopropalin. The resistance in rigid ryegrass to other herbicides and drugs that affect mitosis were tested. Resistance comparable to that of trifluralin was recorded for the herbicides terbutol and DCPA, while low levels of cross-resistance to amiprophosmethyl was present. Trifluralin affected mitotic indices at a much lower level in the susceptible biotypes than in the resistant biotypes. No differences in the uptake and translocation of14C-trifluralin were observed between resistant and susceptible biotypes. Most of the14C detected in the plant material was in the root tissue. A small level of14C was detected in the seeds, and no substantial increases were noted in coleoptile tissue. The resistance spectra in SLR 31 and SLR 10 were phenotypically similar to those occurring in an intermediate trifluralin-resistant goosegrass and trifluralin-resistant green foxtail.

Rigid ryegrass population VLR69 has become resistant to nine classes of herbicides after 21 yr of exposure to five herbicides in five different chemical classes. The population was exposed to diuron in 17 seasons and is resistant to diuron (4 fold) and chlorotoluron (8 fold) when compared with a reference susceptible population (VLR1). VLR69 had six seasons of exposure to chlorsulfuron and exhibits a high level of resistance to chlorsulfuron (> 20 fold) and triasulfuron (> 25 fold) and a lesser change in sensitivity to sulfometuron (7 fold); however, 4% of the population has a high level of resistance to sulfometuron. Resistance to atrazine (5 fold), simazine (6 fold), and ametryn (10 fold) was observed after five seasons of exposure to atrazine. There is a high level of resistance to all aryloxyphenoxypropionate herbicides after only two exposures to diclofop eight generations prior to testing the population. The population was cross-resistant to tralkoxydim (> 9.5 fold) and sethoxydim (1.8 fold). There was a small change in sensitivity to paraquat (1.4 fold) after three generations of exposure. The population displayed cross-resistance to: imazaquin (7 fold), imazapyr (2.5 fold), metribuzin (8.7 fold), and metolachlor (2 fold) but was susceptible to oxyfluorfen and dinitroaniline herbicides. There was also a small shift in sensitivity to tridiphane (1.6 fold).

Abstract. The spectrum of herbicide resistance was determined in an annual ryegrass (Lolium rigidum Gaud.) biotype (SLR 3) that had been exposed to the grass herbicide sethoxydim, an inhibitor of the plastidic enzyme acetylcoenzyme A carboxylase (ACCase, EC 6.4.1.2), for three consecutive years. This biotype has an 18-fold resistance to sethoxydim and enhanced resistance to other cyclohexanedione herbicides compared with a susceptible biotype (VLR 1). The resistant biotype also has a 47-to > 300-fold cross-resistance to the aryloxyphenoxypropanoate herbicides which share ACCase as a target site. No resistance is evident to herbicide with a target site different from ACCase. The absorption of [4-14C]sethoxydim, the rate of metabolic degradation and the nature of the herbicide metabolites are similar in the resistant and susceptible biotypes. While the total activity of the herbicide target enzyme ACCase is similar in extracts from the two biotypes, the kinetics of herbicide inhibition differ. The concentrations of sethoxydim and tralkoxydim required to inhibit the activity of ACCase by 50% are 7.8 and > 9.5 times higher, respectively, in the resistant biotype. The activity of ACCase from the resistant biotype was also less sensitive to aryloxyphenoxypropanode herbicides than the susceptible biotype. The spectrum of resistance at the whole-plant level is correlated with resistance at the ACCase level and confirms that a less sensitive form of the target enzyme endows resistance in biotype SLR 3.