Multiple Herbicide Resistance Research Articles

Weed control in wheat with pyroxasulfone and its combinations with other herbicides

AbstractField and pot investigations were conducted to determine the effectiveness of pyroxasulfone alone and its combinations with other herbicides against diverse weed flora of wheat (Triticum aestivum L.) including multiple herbicide‐resistant (MHR) littleseed canarygrass (Phalaris minor Retz.). Applications of pyroxasulfone 100–127.5 g/ha as pre‐emergence (PE) or early post‐emergence (EPOE) @ 63.75 g/ha at 21–23 days after sowing (1 day before irrigation) were highly effective for control of grass weeds namely P. minor and wild oat (Avena ludoviciana Dur.). It was poor for control of broad‐leaved weeds (Medicago denticulata Willd. and Rumex dentatus L.). However, pyroxasulfone in tank‐mix combination with metsulfuron 4 g/ha, triasulfuron 20 g/ha, and pyroxsulam 18 g/ha effectively controlled (96.5%–99.8%) the diverse weed flora and improved the wheat grain yield (69.5%–285.9%) over untreated weedy control. Also, the pre‐mix of pyroxasulfone + pendimethalin applied as PE was superior to either of these applied alone for weed control and grain yield. Pyroxasulfone 100–127.5 g/ha had yield gain of 119.6%–125.4% and 10.1%–26% over untreated control and pendimethalin 800–1250 g/ha, respectively. In pot studies, straw burnt ash drastically reduced the pyroxasulfone efficacy against P. minor and A. ludoviciana. Pyroxasulfone was also effective in pot studies for control of MHR P. minor having resistance against acetyl‐coA carboxylase (ACCase), acetolactate synthase (ALS), and photosynthesis at the photosystem‐II site‐A (PS‐II) inhibitor herbicides (clodinafop, sulfosulfuron, and isoproturon, respectively). The studies indicate that pyroxasulfone as PE or EPOE can be an alternative grass weed control herbicide in wheat in particular for the control of MHR P. minor.

Convergent Adaptation of Multiple Herbicide Resistance to Auxin Mimics and ALS- and EPSPS-Inhibitors in Brassica rapa from North and South America.

Herbicide-resistant weeds have been identified and recorded on every continent where croplands are available. Despite the diversity of weed communities, it is of interest how selection has led to the same consequences in distant regions. Brassica rapa is a widespread naturalized weed that is found throughout temperate North and South America, and it is a frequent weed among winter cereal crops in Argentina and in Mexico. Broadleaf weed control is based on glyphosate that is used prior to sowing and sulfonylureas or mimic auxin herbicides that are used once the weeds have already emerged. This study was aimed at determining whether a convergent phenotypic adaptation to multiple herbicides had occurred in B. rapa populations from Mexico and Argentina by comparing the herbicide sensitivity to inhibitors of the acetolactate synthase (ALS), 5-enolpyruvylshikimate-3-phosphate (EPSPS), and auxin mimics. Five B. rapa populations were analyzed from seeds collected in wheat fields in Argentina (Ar1 and Ar2) and barley fields in Mexico (Mx1, Mx2 and MxS). Mx1, Mx2, and Ar1 populations presented multiple resistance to ALS- and EPSPS-inhibitors and to auxin mimics (2,4-D, MCPA, and fluroxypyr), while the Ar2 population showed resistance only to ALS-inhibitors and glyphosate. Resistance factors ranged from 947 to 4069 for tribenuron-methyl, from 1.5 to 9.4 for 2,4-D, and from 2.7 to 42 for glyphosate. These were consistent with ALS activity, ethylene production, and shikimate accumulation analyses in response to tribenuron-methyl, 2,4-D, and glyphosate, respectively. These results fully support the evolution of the multiple- and cross-herbicide resistance to glyphosate, ALS-inhibitors, and auxinic herbicides in B. rapa populations from Mexico and Argentina.

The Impact of the Addition of Urea Ammonium Nitrate to 2,4-D on the Control of Multiple-Herbicide-Resistant Canada Fleabane

Limited information exists on the impact of the addition of 28% urea ammonium nitrate (UAN) to 2,4-D, applied preplant (PP) to soybean, on the control of multiple-herbicide-resistant (MHR) Canada fleabane. A total of five field experiments were conducted over a two-year period (2020, 2021) in southwestern Ontario to determine if MHR Canada fleabane control with 2,4-D ester applied PP can be improved by adding UAN to the spray solution. Glyphosate + 2,4-D ester applied PP with various rates of UAN caused no soybean injury. The control of MHR Canada fleabane decreased when UAN was added to the spray solution, especially at higher rates evaluated. At 4 weeks after treatment (WAT), the predicted percent UAN concentration in the spray solution that caused a 5, 10, 20, or 50% decrease in MHR Canada fleabane control was 12, 17, 25, and 52%, respectively. At 8 WAT, the predicted percent UAN concentration that caused a 5, 10, 20, or 50% decrease in MHR Canada fleabane control was 10, 13, 18, and 33%, respectively. At 8 WAT, the predicted percent UAN concentration that caused a 5 and 10% increase in MHR Canada fleabane density was 36 and 73%, respectively. At 8 WAT, the predicted percent UAN concentration that caused a 5, 10, 20, or 50% increase in MHR Canada fleabane biomass was 2, 4, 8, and 19%, respectively. The predicted percent UAN concentration that caused a 5, 10, and 20% decrease in soybean yield was 1, 3, and 12%, respectively. This study concludes that the addition of 28% UAN, especially at the higher concentration, to 2,4-D ester reduces control and increases the density and biomass of MHR Canada fleabane.

Multiple herbicide-resistant waterhemp control with Group 15 herbicides

AbstractWaterhemp has evolved resistance to seven herbicide modes of action in the United States and to five in Canada, which limits weed control options for producers. The objective of this research was to quantify the level and duration of residual control of multiple herbicide-resistant (MHR) waterhemp with five Group 15 herbicides (acetochlor, dimethenamid-p, flufenacet, pyroxasulfone, andS-metolachlor) applied preemergence in a non-crop area. Four field trials were conducted over a 2-yr period (2021, 2022) in southwestern Ontario, Canada. By 4 wk after application (WAA) 91% of waterhemp had emerged in the nontreated control area. The numerical control of waterhemp with all Group 15 herbicides, with the exception of pyroxasulfone, was greatest at 4 WAA, then control declined. Flufenacet provided the lowest waterhemp control; dimethenamid-p andS-metolachlor provided intermediate control, and acetochlor and pyroxasulfone provided the highest control. Waterhemp control with pyroxasulfone peaked at 6 WAA with 99% and declined to 77% at 12 WAA. Flufenacet (low and high rates) was predicted to reduce waterhemp emergence by 50% for 42 to 44 d after application (DAA). Dimethenamid-p,S-metolachlor, and acetochlor (both formulations and three rates) were predicted to reduce waterhemp emergence by 80% for 36, 43, and 33 to 51 DAA, respectively; in contrast, pyroxasulfone was predicted to reduce waterhemp emergence by 80% for 82 DAA. This study concludes that of the Group 15 herbicides evaluated, flufenacet provides the lowest and shortest residual control of waterhemp, and pyroxasulfone provides the highest and longest residual control of waterhemp.

Control of Multiple-Herbicide-Resistant Canada Fleabane With Fall, Spring, and Sequential Herbicide Applications in Winter Wheat

Limited information exists on the efficacy of pyrasulfotole/bromoxynil, fluroxypyr/halauxifen-methyl + MCPA EHE, and clopyralid applied in the fall, spring, or sequentially [fall followed by (fb) spring] for the control of multiple-herbicide-resistant (MHR) Canada fleabane in winter wheat under Ontario environmental conditions. Three field experiments were initiated in the autumn of 2020 and 2021 for a total of 6 site-years to evaluate fall- and spring-applied herbicides and their sequential applications for the control of MHR Canada fleabane in winter wheat in Ontario. Pyrasulfotole/bromoxynil applied in the fall, spring, or sequentially controlled MHR Canada fleabane 83, 99, and 100%, respectively at 8 weeks after the spring application (WAB); the spring and sequential applications provided better control than the fall application. Fluroxypyr/halauxifen + MCPA and clopyralid applied in the fall, spring, or sequentially controlled MHR Canada fleabane 97 to 100% and 99 to 100%, respectively at 8 WAB. Based on orthogonal contrasts the spring and sequential herbicide applications provided greater control than the fall application (8 WAB). MHR Canada fleabane interference reduced winter wheat yield up to 27% in this study. Based on orthogonal contrasts reduced MHR Canada fleabane interference with the fall application resulted in 17% higher winter wheat yield than when herbicide application was delayed to the spring. Although MHR Canada fleabane was controlled very effectively with clopyralid winter wheat yield was lower, presumably due to crop injury; this observation will have to be explored further in future research. Results from this study indicate that pyrasulfotole/bromoxynil and fluroxypyr/halauxifen + MCPA applied in the fall can be used to effectively control MHR Canada fleabane and minimize winter wheat yield loss due to weed interference.

ABC transporters linked to multiple herbicide resistance in blackgrass (Alopecurus myosuroides).

Enhanced detoxification is a prominent mechanism protecting plants from toxic xenobiotics and endows resistance to diverse herbicide chemistries in grass weeds such as blackgrass (Alopecurus myosuroides). The roles of enzyme families which impart enhanced metabolic resistance (EMR) to herbicides through hydroxylation (phase 1 metabolism) and/or conjugation with glutathione or sugars (phase 2) have been well established. However, the functional importance of herbicide metabolite compartmentalisation into the vacuole as promoted by active transport (phase 3), has received little attention as an EMR mechanism. ATP-binding cassette (ABC) transporters are known to be important in drug detoxification in fungi and mammals. In this study, we identified three distinct C-class ABCCs transporters namely AmABCC1, AmABCC2 and AmABCC3 in populations of blackgrass exhibiting EMR and resistance to multiple herbicides. Uptake studies with monochlorobimane in root cells, showed that the EMR blackgrass had an enhanced capacity to compartmentalize fluorescent glutathione-bimane conjugated metabolites in an energy-dependent manner. Subcellular localisation analysis using transient expression of GFP-tagged AmABCC2 assays in Nicotiana demonstrated that the transporter was a membrane bound protein associated with the tonoplast. At the transcript level, as compared with herbicide sensitive plants, AmABCC1 and AmABCC2 were positively correlated with EMR in herbicide resistant blackgrass being co-expressed with AmGSTU2a, a glutathione transferase (GST) involved in herbicide detoxification linked to resistance. As the glutathione conjugates generated by GSTs are classic ligands for ABC proteins, this co-expression suggested AmGSTU2a and the two ABCC transporters delivered the coupled rapid phase 2/3 detoxification observed in EMR. A role for the transporters in resistance was further confirmed in transgenic yeast by demonstrating that the expression of either AmABCC1 or AmABCC2, promoted enhanced tolerance to the sulfonylurea herbicide, mesosulfuron-methyl. Our results link the expression of ABCC transporters to enhanced metabolic resistance in blackgrass through their ability to transport herbicides, and their metabolites, into the vacuole.

Herbicide‐resistant barnyardgrass (Echinochloa crus‐galli) in global rice production

AbstractBarnyardgrass (Echinochloa crus‐galli (L.) Beauv.), an annual species of the family Poaceae, is a major weed problem in rice‐producing countries throughout the globe. Synthetic herbicides can effectively control this grass in rice paddies, but the development of resistant biotypes after the continuous use of the same active ingredients has led to low herbicide efficacy and yield losses. In this review, a summary of resistant‐barnyardgrass cases in global rice production is reported based on data from the International Herbicide‐Resistant Weed Database. The first case of resistant barnyardgrass in rice paddies was to the photosystem‐II inhibitor propanil in the late 1980s. Eighty‐five (85) out of 116 cases in the period from 1986 to 2022 refer to resistant barnyardgrass (E. crus‐galli var. crus‐galli, E. crus‐galli var. formosensis and E. crus‐galli var. zelayensis) in 16 countries. Barnyardgrass has been found resistant to acetolactate synthase (ALS) inhibitors (34 cases), acetyl‐CoA carboxylase (ACCase) inhibitors (23 cases), photosystem‐II inhibitors (11 cases), auxin mimics/cellulose biosynthesis inhibitors (9 cases), very long chain fatty acid inhibitors (6 cases), and microtubule assembly inhibitors (1 case). The majority of all resistance cases reported to the active ingredients penoxsulam, bispyribac‐sodium, and imazamox (ALS inhibitors), cyhalofop‐butyl and fenoxaprop‐ethyl (ACCase inhibitors), propanil (photosystem‐II inhibitors), and quinclorac (auxin mimics/cellulose biosynthesis inhibitors). Although target‐site resistance with specific mutations has been identified, non‐target site resistance mainly through herbicide detoxification is also of great concern increasing the chance of multiple herbicide resistance evolution. Rotation of herbicides should be adopted concerning the modes of action used as well as the application methods to mitigate resistance evolution of this weed in rice paddies.

Integrated weed management strategies for the depletion of multiple herbicide-resistant waterhemp (Amaranthus tuberculatus) seed in the soil seedbank

AbstractThe development of an integrated weed management (IWM) strategy for control of multiple herbicide-resistant (MHR) waterhemp can provide field crop producers with a strategy to deplete the number of waterhemp seeds in the soil seedbank. Field experiments were established on two commercial farms in Ontario, Canada, with MHR waterhemp in 2017. The number of waterhemp seeds in the seedbank at the Cottam and Walpole Island sites prior to establishing the experiments was 413 and 40 million seeds ha−1, respectively. The goal of this 9-yr study is to document the depletion in the number of waterhemp seeds in the seedbank after Years 3, 6, and 9 (spring 2020, 2023, and 2026) and to identify management practices that can reduce the number of waterhemp seeds by 95% or more. Relative to the number of seeds in the soil seedbank when the experiment was initiated, at the Cottam site after 3 yr of this experiment, in the “control” treatment (continuous soybean seeded in rows spaced 75 apart, and sprayed with glyphosate) there was a numeric 31% increase in the number of waterhemp seeds in the seedbank; in contrast, in the three-crop rotation of corn/soybean/winter wheat (with or without a cover crop after winter wheat harvest), soybean seeded in rows spaced 37.5 cm apart, with herbicide applications using a total of eight different herbicide modes of action resulted in a 65% to 66% decrease in the number of waterhemp seeds in the soil seedbank. At the Walpole Island site after 3 yr of this experiment, the number of waterhemp seeds in the seedbank was not affected by the IWM programs evaluated. Results indicate that a diversified integrated waterhemp management program dramatically decreased the number of waterhemp seeds in the seedbank at one of two sites.

Multiple-herbicide-resistant waterhemp control in glyphosate/glufosinate/2,4-D-resistant soybean with one- and two-pass weed control programs

AbstractWaterhemp control in Ontario has increased in complexity due to the evolution of biotypes that are resistant to five herbicide modes of action (Groups 2, 5, 9, 14, and 27 as categorized by the Weed Science Society of America). Four field trials were carried out over a 2-yr period in 2021 and 2022 to assess the control of multiple-herbicide-resistant (MHR) waterhemp biotypes in glyphosate/glufosinate/2,4-D-resistant (GG2R) soybean using one- and two-pass herbicide programs. S-metolachlor/metribuzin, pyroxasulfone/sulfentrazone, pyroxasulfone/flumioxazin, and pyroxasulfone + metribuzin applied preemergence (PRE) controlled MHR waterhemp similarly by 46%, 63%, 60%, and 69%, respectively, at 8 wk after postemergence (POST) application (WAA-B). A one-pass application of 2,4-D choline/glyphosate DMA POST provided greater control of MHR waterhemp than glufosinate. Two-pass herbicide programs of a PRE herbicide followed by (fb) a POST-applied herbicide resulted in greater MHR waterhemp control compared to a single PRE or POST herbicide application. PRE herbicides fb glufosinate or 2,4-D choline/glyphosate DMA POST controlled MHR waterhemp by 74% to 91% and by 84% to 96%, respectively, at 8 WAA-B. Two-pass herbicide applications of an effective PRE residual herbicide fb 2,4-D choline/glyphosate DMA POST in GG2R soybean can effectively manage waterhemp that is resistant to herbicides in Groups 2, 5, 9, 14, and 27.

Impact of Acetochlor Rate and Application Timing on Multiple-Herbicide-Resistant Waterhemp Control in Corn and Soybean

Documented 6-way (Groups 2, 4, 5, 9, 14, and 27) and 5-way (Groups 2, 5, 9, 14, and 27) multiple-herbicide-resistant (MHR) waterhemp have been confirmed in the US and Canada, respectively causing corn and soybean yield losses > 70%. The objective of this study was to determine the effect of acetochlor application timing and rate on non-emerged MHR waterhemp control in corn and soybean. Acetochlor is not yet registered in Canada, but it could be useful component of an integrated MHR waterhemp control program. Two studies, one in corn and one in soybean, were conducted in southwestern Ontario, Canada from 2020 to 2022. Three rates of acetochlor were applied preplant (PP), preemergence (PRE) and postemergence (POST) to non-emerged waterhemp. In corn, acetochlor [Emulsifiable Concentrate (EC)] applied at 1,225, 2,100 and 2,950 g ai ha-1 controlled MHR waterhemp 81, 85, and 90%, respectively, at 8 weeks after POST application (WAC). Acetochlor EC applied POST or PRE provided better control than when applied PP at 4, 8, and 12 WAC. In soybean, acetochlor [Capsule suspension (CS)] applied at 1,050, 1,375, and 1,700 g ai ha-1 controlled MHR waterhemp 63, 70, and 74%, respectively, at 8 WAC. The timing of acetochlor CS application did not affect MHR waterhemp control. Acetochlor applied at the low, medium, and high rate reduced waterhemp density by 87, 89, and 92% in corn, and by 82, 84, and 87% in soybean, respectively. The high rate of acetochlor provides acceptable control of MHR waterhemp in corn; control in soybean was inadequate.

Multiple Resistance to Three Modes of Action of Herbicides in a Single Italian Ryegrass (Lolium multiflorum L.) Population in China

Italian ryegrass (Lolium multiflorum L.), a cross-pollinated grass, is gradually becoming a predominant weed in wheat fields in China and is evolving resistance to many groups of herbicides. The aim of this study is to determine the resistance levels of a single L. multiflorum population from a wheat field in Henan Province China, to three modes of action (MoAs) of herbicides and to further characterize the potential resistance mechanisms. This L. multiflorum population evolved multiple herbicide resistances to pyroxsulam [acetolactate synthase (ALS)], pinoxaden [acetyl-CoA carboxylase (ACCase)] and isoproturon [photosystem II (PSII)]. Target-site resistance (TSR) mutations (Pro-197-Gln, Pro-197-Thr, and Trp-574-Leu) and non-target-site resistance (NTSR) mediated by cytochrome P450 monooxygenase (CYP450) genes were associated with pyroxsulam resistance. Pinoxaden resistance was conferred by two TSR mutations, which referred to a rare Ile-2041-Val mutation and a common Ile-1781-Leu mutation but with two different nucleotide substitutions (CTA/TTA). CYP450- and glutathione-S-transferase (GST)-mediated resistances were the main resistance mechanisms for this multiple herbicide-resistant (MHR) population to the PSII inhibitor isoproturon. This is the first case of a single L. multiflorum population evolving multiple resistance to three herbicide MoAs (ALS, ACCase and PSII) in China. Diverse resistance mechanisms including TSR and NTSR mean L. multiflorum exhibits a high degree of resistance plasticity.

Confirmation of 4-Hydroxyphenylpyruvate Dioxygenase Inhibitor-Resistant and 5-Way Multiple-Herbicide-Resistant Waterhemp in Ontario, Canada

This is the first confirmation of 4-hydroxyphenylpyruvate dioxygenase-(HPPD)-inhibitor resistant waterhemp and the first report of 5-way multiple-herbicide-resistant (MHR) waterhemp in Ontario to Group 2, 5, 9, 14, and 27 herbicides. Seed was collected across southern Ontario in 2021 and tested for resistance to imazethapyr, atrazine, metribuzin, glyphosate, lactofen, mesotrione, and mesotrione + atrazine representing Group 2, 5, 5, 9, 14, 27, and 27 + 5 herbicides, respectively. All of the waterhemp populations collected in 2021 had 4-way resistance to imazethapyr, atrazine, glyphosate, and mesotrione. In seven Ontario counties, 5-way resistant waterhemp was confirmed which includes resistance to the HPPD-inhibiting herbicides.

Target gene mutation and enhanced metabolism confer fomesafen resistance in an Amaranthus retroflexus L. population from China

Amaranthus retroflexus L., a troublesome annual dicotyledonous weed species, is highly competitive with soybean (Glycine max L.). A single-dose herbicide-resistance screening assay identified an A. retroflexus population with suspected resistance to fomesafen. Whole-plant dose–response assays demonstrated that the resistant population (2492) was resistant to protoporphyrinogen oxidase (PPO)-inhibiting herbicides (50.6-fold fomesafen resistance and > 8.1-fold lactofen resistance) compared to a susceptible (S) population. PPX2 gene sequence analysis showed an Arg128Gly amino acid substitution in the 2492 population. Moreover, pretreatment of malathion and the fomesafen metabolic assays through HPLC–MS demonstrated enhanced fomesafen metabolism in the 2492 population. Additionally, the 2492 population was 10.4-fold more resistant to the ALS-inhibiting herbicide imazethapyr and 16.8-fold more resistant to thifensulfuron-methyl than the S population. ALS gene sequence analysis showed an Ala205Val amino acid substitution in the 2492 population. This population of A. retroflexus has coexisting target-site resistance and non-target-site mechanisms for resistance to fomesafen. Multiple herbicide resistance may mean it is necessary to adjust weed management strategies to better control the resistant population.

Control of Multiple-Herbicide-Resistant Green Pigweed (Amaranthus powellii) With Preemergence and Postemergence Herbicides in Ontario Corn Production

Green pigweed [Amaranthus powellii S.Wats.] is a prolific annual dicot weed that is a prominent weed of crop production in northeastern North America. Green pigweed interference has been documented to reduce corn yields up to 54% in the absence of control strategies. In 2021, a green pigweed biotype from a field near Dresden, Ontario, Canada was determined to be resistant to MCPA, mecoprop, dichlorprop-p, aminocyclopyrachlor (synthetic auxins), and imazethapyr (acetolactate synthase-(ALS)-inhibitor), further impacting control of this weed biotype. Two field studies, with herbicides applied preemergence (PRE) or postemergence (POST), were conducted in 2020 and repeated in 2021. The objective of the research was to determine the most effective PRE and POST herbicides for the control of multiple-herbicide-resistant (MHR) green pigweed in Ontario corn production. 18 PRE and 18 POST herbicide treatments were evaluated in separate studies. Visible crop injury, visible green pigweed control at specified timepoints after herbicide application, green pigweed density, green pigweed biomass, and corn yield at harvest maturity were collected. In the PRE study, rimsulfuron + mesotrione was identified as the most efficacious treatment providing 88% control at 8 WAA. In the POST study, atrazine was identified as the most efficacious treatment providing 94% control at 8 WAA. Control with all PRE herbicide treatments was impacted by rainfall following application. MCPA ester applied POST controlled green pigweed 30% at 8 WAA; reduced control is attributed to herbicide resistance in this biotype. When compared to similar studies, control of green pigweed was reduced with some of the POST herbicides tested. While MHR green pigweed represents an additional challenge for growers, there are efficacious herbicide treatments that would allow it to be managed in corn production.

Tribenuron-methyl metabolism and the rare Pro197Phe double mutation together with 2,4-D metabolism and reduced absorption can evolve in Papaver rhoeas with multiple and cross herbicide resistance to ALS inhibitors and auxin mimics

Multiple resistance mechanisms to ALS inhibitors and auxin mimics in two Papaver rhoeas populations were investigated in wheat fields from Portugal. Dose-response trials, also with malathion (a cytochrome P450 inhibitor), cross-resistance patterns for ALS inhibitors and auxin mimics, alternative herbicides tests, 2,4-D and tribenuron-methyl absorption, translocation and metabolism experiments, together with ALS activity, gene sequencing and enzyme modelling and ligand docking were carried out. Results revealed two different resistant profiles: one population (R1) multiple resistant to tribenuron-methyl and 2,4-D, the second (R2) only resistant to 2,4-D. In R1, several target-site mutations in Pro197 and enhanced metabolism (cytochrome P450-mediated) were responsible of tribenuron-methyl resistance. For 2,4-D, reduced transport was observed in both populations, while cytochrome P450-mediated metabolism was also present in R1 population. Moreover, this is the first P. rhoeas population with enhanced tribenuron-methyl metabolism. This study reports the first case for P. rhoeas of the amino acid substitution Pro197Phe due to a double nucleotide change. This double mutation could cause reduced enzyme sensitivity to most ALS inhibitors according to protein modelling and ligand docking. In addition, this study reports a P. rhoeas population resistant to 2,4-D, apparently, with reduced transport as the sole resistance mechanism.

Control of Multiple-Herbicide-Resistant Green Pigweed (Amaranthus powellii) with Preemergence and Postemergence Herbicides in Ontario Soybean Production

Green pigweed [Amaranthus powellii S. Wats.] is a competitive, annual, broadleaf weed that can significantly reduce soybean yield due to rapid early growth and biomass production. As a consequence of green pigweed’s high competitiveness with crops, the weed species is generally managed with the use of herbicides; this may, however, lead to the selection of resistance. A green pigweed biotype with resistance to MCPA, mecoprop, dichlorprop-p, aminocyclopyrachlor (synthetic auxins), and imazethapyr (acetolactate synthase-(ALS)-inhibitor) was recently confirmed in Ontario, Canada. Research was conducted to identify alternative effective preemergence (PRE) and postemergence (POST) herbicides for the control of multiple-herbicide-resistant (MHR) green pigweed in Ontario soybean production. Four field trials, two with PRE and two with POST herbicides were conducted near Dresden, Ontario in 2019 and 2020. Visible soybean injury, green pigweed control, density, aboveground biomass, and soybean yield were evaluated following application with 19 PRE herbicide treatments and 12 POST herbicide treatments in separate studies. At 8 wk after application (WAA), pyroxasulfone/flumioxazin applied PRE controlled green pigweed 95% and was the most effective soil-applied herbicide. In the POST study, glyphosate was the most effective herbicide, controlling green pigweed 94% at 8 WAA. Imazethapyr, applied PRE or POST did not control this green pigweed biotype as it is also resistant to ALS-inhibitors. Control with other ALS-inhibiting herbicides as well as with PPO-inhibiting herbicides was variable, and further research is needed to determine the reason for variable control with these herbicides.

Target-site mutation and cytochrome P450s confer resistance to multiple herbicides in Italian ryegrass (Lolium multiflorum Lam.) from China

Italian ryegrass (Lolium multiflorum Lam.) is a highly competitive grass weed capable of severely infesting wheat fields worldwide. In recent years, Chinese farmers are finding it increasingly difficult to control L. multiflorum when using acetyl-CoA carboxylase (ACCase)-inhibiting herbicides. Accordingly, clarifying the resistance pattern of putative resistant populations of L. multiflorum is important for selecting alternative herbicides for effective weed control. In this study, a putative resistant L. multiflorum population, HN-03, which apparently has survived clodinafop-propargyl treatment under the recommended field application rate, was collected from a wheat field in Zhumadian, Henan, China. Whole-plant bioassay revealed that the HN-03 population had evolved high-level resistance to clodinafop-propargyl. Sequencing of the ACCase gene revealed a point mutation, of TGG to TGT, at codon position 1999, causing a Trp-to-Cys substitution in the resistant plants. The HN-03 population also exhibited cross-resistance to fenoxaprop-P-ethyl, clethodim, and pinoxaden and multiple-resistance to mesosulfuron-methyl and bixlozone, but it remained susceptible to cypyrafluone. Pre-treatment with piperonyl butoxide and malathion considerably increased the susceptibility of the HN-03 population to fenoxaprop-P-ethyl and mesosulfuron-methyl, by 25.3% and 36.6%, respectively. This study shows the L. multiflorum population HN-03 has evolved multiple herbicide resistance due to both a target-site mutation and P450s-mediated enhanced metabolism. Cypyrafluone can be used as an alternative herbicide to control resistant L. multiflorum, and it might be potentially helpful for improving the management of herbicide resistance in other aggressive weeds.

Cross‐ and multiple herbicide resistance in Lolium multiflorum across Uruguay

AbstractHerbicide resistance is currently one of the most important problems in agricultural systems, threatening their sustainability. In Uruguay, there have been increasing reports of herbicide failures to control LOLMU (Lolium multiflorum Lam.) populations. However, methodical characterization of herbicide resistance cases is lacking. Seeds from 66 LOLMU populations were collected from cropping fields and screened in greenhouse conditions with the recommended (1×) and half the recommended (1/2×) label dose of glyphosate, clethodim, pinoxaden and a formulation with iodosulfuron‐methyl‐sodium and mesosulfuron‐methyl (IodoMeso). Populations were classified as resistant when mortality was equal to or lower than 80% in response to 1× doses, assessed 28 days after treatment (DAT). Results show that 80% of LOLMU populations were resistant to glyphosate, followed by 49%, 36% and 24% resistance to IodoMeso, pinoxaden and clethodim respectively. Moreover, 52% of the tested populations were resistant to more than one herbicide, with an important frequency of populations resistant to glyphosate and IodoMeso (17%) and to all tested herbicides (16%). In addition, a high correlation of resistance levels was found between pinoxaden and clethodim resistant populations (R2 = 0.84). Geographic distribution of tested populations suggests that multiple herbicide resistance is concentrated in fields with a longer history of herbicide use. Overall, these results confirm that herbicide resistance in LOLMU is widespread in agricultural regions of Uruguay and that significant number of multiple resistance cases already exist and are expected to increase rapidly if management practices are not adjusted quickly and substantially. This survey represents a key input to improve herbicide‐resistant LOLMU management strategies.

Directed Evolution of Phi Class Glutathione Transferases Involved in Multiple-Herbicide Resistance of Grass Weeds and Crops.

The extensive application of herbicides in crop cultivation has indisputably led to the emergence of weed populations characterized by multiple herbicide resistance (MHR). This phenomenon is associated with the enhanced metabolism and detoxifying ability of endogenous enzymes, such as phi class glutathione transferases (GSTFs). In the present work, a library of mutant GSTFs was created by in vitro directed evolution via DNA shuffling. Selected gstf genes from the weeds Alopecurus myosuroides and Lolium rigidum, and the cereal crops Triticum durum and Hordeum vulgare were recombined to forge a library of novel chimeric GSTFs. The library was activity screened and the best-performing enzyme variants were purified and characterized. The work allowed the identification of enzyme variants that exhibit an eight-fold improvement in their catalytic efficiency, higher thermal stability (8.3 °C) and three-times higher inhibition sensitivity towards the herbicide butachlor. The crystal structures of the best-performing enzyme variants were determined by X-ray crystallography. Structural analysis allowed the identification of specific structural elements that are responsible for kcat regulation, thermal stability and inhibition potency. These improved novel enzymes hold the potential for utilization in biocatalysis and green biotechnology applications. The results of the present work contribute significantly to our knowledge of the structure and function of phi class plant GSTs and shed light on their involvement in the mechanisms of MHR.

Full chloroplast sequencing using genome skimming for novel plant DNA barcode discovery in Amaranths

DNA barcoding has been established as an efficient, sensitive and reliable methodology for plant identification. However, in spite of efforts to find a universal DNA plant barcode, some taxa are not sufficiently resolved by typical plant barcoding genes like matK or rbcL. We have used a technique known as genome skimming, which relies on the empiric low coverage sequencing of a full plant genome, resulting in high coverage of the high copy genome fractions such as chloroplast and rDNA. Phylogenetic studies show that these regions are a reservoir of variability which could be further exploited for DNA barcode discovery. We sequenced eight amaranth species using Illumina Next Generation Sequencing technology to test the feasibility of this technique. Amaranths were chosen due to their increasing impact as invasive species bearing multiple herbicide resistance mechanisms. Our results showed that complete chloroplast genomes could be assembled for all of the eight species tested. We obtained an average of 47 million reads for each one of the amaranth nuclear genomes, which range in size between 400-700Mb approximately. These reads provide an average theoretical coverage of 10-15X for each nuclear genome, but resulted in an average chloroplast genome coverage in the range of 500-8000X due to multiple chloroplast genome copies per cell. Alignment of the eight chloroplast genomes shows variability in the single copy regions (Fig. 1), especially on intergenic sections (Fig. 2). Additional preliminary analyses also show variation among different populations of the same species, demonstrating the importance of studying both inter and intraspecific diversity to design reliable and accurate DNA barcodes that can be used in species identification.