Trade-offs constrain the success of glyphosate-free farming.

Glyphosate, the most widely used herbicide, is linked with environmental harm and there is a drive to replace it in agricultural systems. We model the impacts of discontinuing glyphosate use and replacing it with cultural control methods. We simulate winter wheat arable systems reliant on glyphosate and typical in northwest Europe. Removing glyphosate was projected to increase weed abundance, herbicide risk to the environment, and arable plant diversity and decrease food production. Weed communities with evolved resistance to non-glyphosate herbicides were not projected to be disproportionately affected by removing glyphosate, despite the lack of alternative herbicidal control options. Crop rotations with more spring cereals or grass leys for weed control increased arable plant diversity. Stale seedbed techniques such as delayed drilling and choosing ploughing instead of minimum tillage had varying effects on weed abundance, food production, and profitability. Ploughing was the most effective alternative to glyphosate for long-term weed control while maintaining production and profit. Our findings emphasize the need for careful consideration of trade-offs arising in scenarios where glyphosate is removed. Integrated Weed Management (IWM) with more use of cultural control methods offers the potential to reduce chemical use but is sensitive to seasonal variability and can incur negative environmental and economic impacts.

The Cys-2088-Arg mutation in the ACCase gene and enhanced metabolism confer cyhalofop-butyl resistance in Chinese sprangletop (Leptochloa chinensis).

Acetyl-CoA carboxylase (ACCase)-inhibiting herbicides are among the most commonly used herbicides to control grassy weeds, especially Leptochloa chinensis, in rice fields across China. Herein, we collected a suspected resistant (R) population of L. chinensis (HFLJ16) from Lujiang county in Anhui Province. Whole plant dose response tests showed that, compared with the susceptible (S) population, the R population showed high resistance to cyhalofop-butyl (22-fold) and displayed cross-resistance to metamifop (9.7-fold), fenoxaprop-P-ethyl (18.7-fold), quizalofop-P-ethyl (7.6-fold), clodinafop-propargyl (12-fold) and clethodim (8.4-fold). We detected an amino acid substitution (Cys-2088-Arg) in the ACCase of resistant L. chinensis. However, ACCase gene expression levels were not significantly different (P > 0.05) between R plants and S plants, without or with cyhalofop-butyl treatment. Furthermore, pretreatment with piperonyl butoxide (PBO, a cytochrome P450 monooxygenase (CYP450) inhibitor) or 4-chloro-7-nitrobenzoxadiazole (NBD-Cl, a glutathione-S-transferase (GST) inhibitor), inhibited the resistance of the R population to cyhalofop-butyl significantly (by approximately 60% and 26%, respectively). Liquid chromatography tandem mass spectrometry analysis showed that R plants metabolized cyhalofop-butyl and cyhalofop acid (its metabolite) significantly faster than S plants. Three CYP450 genes, one GST gene, and two ABC transporter genes were induced by cyhalofop-butyl and were overexpressed in the R population. Overall, GST-associated detoxification, CYP450 enhancement, and target-site gene mutation are responsible for the resistance of L. chinensis to cyhalofop-butyl.

Metabolism of the 4-Hydroxyphenylpyruvate Dioxygenase Inhibitor, Mesotrione, in Multiple-Herbicide-Resistant Palmer amaranth (Amaranthus palmeri).

Metabolic resistance to the maize-selective, HPPD-inhibiting herbicide, mesotrione, occurs via Phase I ring hydroxylation in resistant waterhemp and Palmer amaranth; however, mesotrione detoxification pathways post-Phase I are unknown. This research aims to (1) evaluate Palmer amaranth populations for mesotrione resistance via survivorship, foliar injury, and aboveground biomass, (2) determine mesotrione metabolism rates in Palmer amaranth populations during a time course, and (3) identify mesotrione metabolites including and beyond Phase I oxidation. The Palmer amaranth populations, SYNR1 and SYNR2, exhibited higher survival rates (100%), aboveground biomass (c.a. 50%), and lower injury (25-30%) following mesotrione treatment than other populations studied. These two populations also metabolized mesotrione 2-fold faster than sensitive populations, PPI1 and PPI2, and rapidly formed 4-OH-mesotrione. Additionally, SYNR1 and SYNR2 formed 5-OH-mesotrione, which is not produced in high abundance in waterhemp or naturally tolerant maize. Metabolite features derived from 4/5-OH-mesotrione and potential Phase II mesotrione-conjugates were detected and characterized by liquid chromatography-mass spectrometry (LCMS).

A Critical Review of Conventional and Modern Approaches to Develop Herbicide-Resistance in Rice.

Together with rice, weeds strive for nutrients and space in farmland, resulting in reduced rice yield and quality. Planting herbicide-resistant rice varieties is one of the effective ways to control weeds. In recent years, a series of breakthroughs have been made to generate herbicide-resistant germplasm, especially the emergence of biotechnological tools such as gene editing, which provides an inherent advantage for the knock-out or knock-in of the desired genes. In order to develop herbicide-resistant rice germplasm resources, gene manipulation has been conducted to enhance the herbicide tolerance of rice varieties through the utilization of techniques such as physical and chemical mutagenesis, as well as genome editing. Based on the current research and persisting problems in rice paddy fields, research on the generation of herbicide-resistant rice still needs to explore genetic mechanisms, stacking multiple resistant genes in a single genotype, and transgene-free genome editing using the CRISPR system. Current rapidly developing gene editing technologies can be used to mutate herbicide target genes, enabling targeted genes to maintain their biological functions, and reducing the binding ability of target gene encoded proteins to corresponding herbicides, ultimately resulting in herbicide-resistant crops. In this review article, we have summarized the utilization of conventional and modern approaches to develop herbicide-resistant cultivars in rice as an effective strategy for weed control in paddy fields, and discussed the technology and research directions for creating herbicide-resistant rice in the future.

Role of glutathione S-transferases in the mode of action of herbicides that inhibit amino acid synthesis in Amaranthus palmeri.

Acetolactate synthase inhibitors (ALS inhibitors) and glyphosate are two classes of herbicides that act by inhibiting an enzyme in the biosynthetic pathway of branched-chain or aromatic amino acids, respectively. Besides amino acid synthesis inhibition, both herbicides trigger similar physiological effects in plants. The main aim of this study was to evaluate the role of glutathione metabolism, with special emphasis on glutathione S-transferases (GSTs), in the mode of action of glyphosate and ALS inhibitors in Amaranthus palmeri. For that purpose, plants belonging to a glyphosate-sensitive (GLS) and a glyphosate-resistant (GLR) population were treated with different doses of glyphosate, and plants belonging to an ALS-inhibitor sensitive (AIS) and an ALS-inhibitor resistant (AIR) population were treated with different doses of the ALS inhibitor nicosulfuron. Glutathione-related contents, GST activity, and related gene expressions (glutamate-cysteine ligase, glutathione reductase, Phi GST and Tau GST) were analysed in leaves. According to the results of the analytical determinations, there were virtually no basal differences between GLS and GLR plants or between AIS and AIR plants. Glutathione synthesis and turnover did not follow a clear pattern in response to herbicides, but GST activity and gene expression (especially Phi GSTs) increased with both herbicides in treated sensitive plants, possibly related to the rocketing H2O2 accumulation. As GSTs offered the clearest results, these were further investigated with a multiple resistant (MR) population, compressing target-site resistance to both glyphosate and the ALS inhibitor pyrithiobac. As in single-resistant plants, measured parameters in the MR population were unaffected by herbicides, meaning that the increase in GST activity and expression occurs due to herbicide interactions with the target enzymes.

Acting pre-emptively reduces the long-term costs of managing herbicide resistance.

Globally, pesticides improve crop yields but at great environmental cost, and their overuse has caused resistance. This incurs large financial and production losses but, despite this, very diversified farm management that might delay or prevent resistance is uncommon in intensive farming. We asked farmers to design more diversified cropping strategies aimed at controlling herbicide resistance, and estimated resulting weed densities, profits, and yields compared to prevailing practice. Where resistance is low, it is financially viable to diversify pre-emptively; however, once resistance is high, there are financial and production disincentives to adopting diverse rotations. It is therefore as important to manage resistance before it becomes widespread as it is to control it once present. The diverse rotations targeting high resistance used increased herbicide application frequency and volume, contributing to these rotations' lack of financial viability, and raising concerns about glyphosate resistance. Governments should encourage adoption of diverse rotations in areas without resistance. Where resistance is present, governments may wish to incentivise crop diversification despite the drop in wheat production as it is likely to bring environmental co-benefits. Our research suggests we need long-term, proactive, food security planning and more integrated policy-making across farming, environment, and health arenas.

A first survey for herbicide resistant weeds across major maize growing areas in the North Island of New Zealand.

Weeds are increasingly documented with evolved resistance to herbicides globally. Three species have been reported as resistant in maize crops in New Zealand: Chenopodium album to atrazine and dicamba, Persicaria maculosa to atrazine and Digitaria sanguinalis to nicosulfuron. Despite knowledge of these cases, the distribution of these resistant biotypes is unknown. This study aimed to determine the prevalence of known resistant weeds in major maize growing areas in New Zealand, and to pro-actively screen other species for resistance. Weed seeds of broadleaf and grass species were collected from 70 randomly selected maize growing farms in the North Island in 2021-2022. Seeds were grown and treated with herbicides at recommended field rates. Atrazine-resistant C. album were recorded in a third of surveyed farms and nicosulfuron-resistant D. sanguinalis in a sixth. Half of Waikato farms and a quarter of Bay of Plenty farms (no Hawkes Bay or Wellington farms) had atrazine-resistant C. album. Dicamba-resistant C. album were not detected, nor were atrazine-resistant P. maculosa. Nicosulfuron resistant D. sanguinalis was recorded in 19% of Waikato farms, 6% of Bay of Plenty farms and 9% of Hawkes Bay farms (no Wellington farms). Amaranthus spp., Fallopia convolvulus, Persicaria spp., Solanum spp., Echinochloa crus-galli, Panicum spp. and Setaria spp. were not resistant to any of the herbicides tested. Twenty-nine to 52% of maize farms in the North Island are estimated to have herbicide resistant weeds. Resistance is common in maize farms in Waikato and western Bay of Plenty. Resistance is rare in southern regions, with only one instance of nicosulfuron-resistant D. sanguinalis and no resistant C. album. Most annual weeds in maize are not resistant to herbicides; although atrazine resistant C. album is widespread, it is currently controlled with alternative herbicides. Resistant D. sanguinalis appears to be an emerging problem.

Diverse ALS mutations and cross-and multiple-resistance to ALS and EPSPS inhibitors in flucarbazone­sodium-resistant Bromus japonicus populations from Hebei province, China.

Japanese brome (Bromus japonicus) has become one of the main weeds in wheat fields in Hebei province of China and causes a large decrease of wheat production. A total of 44 putative resistant and 2 susceptible Japanese brome populations were collected in the 2021/2022 crop season from Hebei province of China to determine resistance levels to flucarbazone­sodium and to investigate the diversity of acetolactate synthase (ALS) mutations, as well as to confirm the cross-and multiple-resistance levels to ALS and EPSPS (5-enolpyruvate shikimate-3-phosphate synthetase) inhibitors. Whole plant bioassay results showed that 15 out of 44 populations tested or 34% were resistant to flucarbazone­sodium. The resistance indices of Japanese brome to flucarbazone­sodium ranged from 43 to 1977. The resistant populations were mainly distributed in Baoding and Shijiazhuang districts, and there was only one resistant population in Langfang district. Resistant Japanese brome had diverse ALS mutations, including Pro-197-Ser, -Thr, -Arg and Asp-376-Glu. The incidence of Pro-197-Ser mutation was the highest at 68%. Application of the CYP450 inhibitor malathion suggested that CYP450 was involved in metabolic resistance in a population without an ALS mutation. The population with Pro-197-Thr mutation evolved weak cross-resistance to mesosulfuron-methyl and pyroxsulam, and it is in the process of evolving multiple-resistance to glyphosate.

Pyroxasulfone Metabolism in Resistant Lolium rigidum: Is It All Down to GST Activity?

Resistance to the herbicide pyroxasulfone has slowly but steadily increased in agricultural weeds. The evolved resistance of one Lolium rigidum population has been attributed to the conjugation of pyroxasulfone to reduced glutathione, mediated by glutathione transferase (GST) activity. To determine if GST-based metabolism is a widespread mechanism of pyroxasulfone resistance in L. rigidum, a number of putative-resistant populations were screened for GST activity toward pyroxasulfone, the presence of GSTF13-like isoforms (previously implicated in pyroxasulfone conjugation in this species), tissue glutathione concentrations, and response to inhibitors of GSTs and oxygenases. Although there were no direct correlations between pyroxasulfone resistance levels and these individual parameters, a random forest analysis indicated that GST activity was of primary importance for L. rigidum resistance to this herbicide.

CRISPR/Cas9-mediated homology donor repair base editing system to confer herbicide resistance in maize (Zea mays L.).

Weed infestation is a significant concern to crop yield loss, globally. The potent broad-spectrum glyphosate (N-phosphomethyl-glycine) has a widely utilized herbicide, acting on the shikimic acid pathway within chloroplast by inhibiting 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). This crucial enzyme plays a vital role in aromatic amino acid synthesis. Repurposing of CRISPR/Cas9-mediated gene-editing was the inflection point for generating novel crop germplasm with diverse genetic variations in essential agronomic traits, achieved through the introduction of nucleotide substitutions at target sites within the native genes, and subsequent induction of indels through error-prone non-homologous end-joining DNA repair mechanisms. Here, we describe the development of efficient herbicide-resistant maize lines by using CRISPR/Cas9 mediated site-specific native ZmEPSPS gene fragment replacement via knock-out of conserved region followed by knock-in of desired homologous donor repair (HDR-GATIPS-mZmEPSPS) with triple amino acid substitution. The novel triple substitution conferred high herbicide tolerance in edited maize plants. Transgene-free progeny harbouring the triple amino acid substitutions revealed agronomic performances similar to that of wild-type plants, suggesting that the GATIPS-mZmEPSPS allele substitutions are crucial for developing elite maize varieties with significantly enhanced glyphosate resistance. Furthermore, the aromatic amino acid contents in edited maize lines were significantly higher than in wild-type plants. The present study describing the introduction of site-specific CRISPR/Cas9- GATIPS mutations in the ZmEPSPS gene via genome editing has immense potential for higher tolerance to glyphosate with no yield penalty in maize.

Controlled release herbicide formulation for effective weed control efficacy.

Controlled release formulation (CRF) of herbicide is an effective weed management technique with less eco-toxicity than other available commercial formulations. To maximise the effectiveness of CRFs however, it is crucial to understand the herbicide-releasing behaviour at play, which predominately depends on the interaction mechanisms between active ingredients and carrier materials during adsorption. In this study, we investigated and modelled the adsorption characteristics of model herbicide 2,4-D onto two organo-montmorillonites (octadecylamine- and aminopropyltriethoxysilane-modified) to synthesise polymer-based CRFs. Herbicide-releasing behaviour of the synthesised CRF microbeads was then analysed under various experimental conditions, and weed control efficacy determined under glasshouse conditions. Results revealed that adsorption of 2,4-D onto both organo-montmorillonites follows the pseudo-second-order kinetics model and is predominately controlled by the chemisorption process. However, multi-step mechanisms were detected in the adsorption on both organoclays, hence intra-particle diffusion is not the sole rate-limiting step for the adsorption process. Both organoclays followed the Elovich model, suggesting they have energetically heterogeneous surfaces. Herbicide-releasing behaviours of synthesised beads were investigated at various pH temperatures and ionic strengths under laboratory and glasshouse conditions. Furthermore, weed control efficacy of synthesised beads were investigated using pot studies under glasshouse condition. Desorption studies revealed that both synthesised microbeads have slow releasing behaviour at a wide range of pHs (5-9), temperatures (25-45 °C), and ionic strengths. The results also revealed that synthesised microbeads have excellent weed control efficacy on different broad-leaf weed species under glasshouse conditions.

Investigating the impacts of airborne dust on herbicide performance on Amaranthus retroflexus.

Dust pollution poses environmental hazards, affecting agriculture through reduced sunlight exposure, photosynthesis, crop yields, and food security. This study explores the interference of dust pollution on herbicide efficacy to control weeds in a semi-arid region. In a factorial experiment conducted in 2019 and replicated in 2020, the interaction of dust and various herbicide applications, including bentazon, sulfosulfuron, tribenuron-methyl, aminopyralid + florasulam, foramsulfuron + iodosulfuron + thiencarbazone, 2,4-D + MCPA, and acetochlor, in controlling Amaranthus retroflexus L. were assessed. Dust induced a 9.2% reduction in the total chlorophyll content of A. retroflexus, while herbicide application independently led to a 67.5% decrease. Contrary to expectations, herbicides performed better in dust, except bentazon, which caused a 28% drop in plant height and a 29% decrease in total biomass compared to non-dust conditions. Both herbicides and dust exerted suppressive effects on A. retroflexus's leaf and stem weights and overall biomass. Despite dust presence, tribenuron-methyl (95.8%), aminopyralid + florasulam (95.7%), sulfosulfuron (96.5%), and foramsulfuron + iodosulfuron + thiencarbazone (97.8%) effectively controlled A. retroflexus. These findings indicate that dust's effect on herbicide efficacy is herbicide-dependent but except bentazon, dust generally increased herbicide efficacy and amplified the control of A. retroflexus.

Metabolic Resistance to Protoporphyrinogen Oxidase-Inhibitor Herbicides in a Palmer amaranth Population from Kansas.

Palmer amaranth has evolved target and nontarget site resistance to protoporphyrinogen oxidase-inhibitor herbicides in the United States. Recently, a population (KCTR) from a long-term conservation tillage study in Kansas was found to be resistant to herbicides from six sites of action, including to PPO-inhibitors, even with this herbicide group being minimally used in this field. This research investigated the level of resistance to postemergence PPO-inhibitors, target- and nontarget-site resistance mechanism(s), and efficacy of pre-emergence chemistries. The greenhouse experiments confirmed 6.1- to 78.9-fold resistance to lactofen in KCTR, with the level of resistance increasing when KCTR was purified for the resistance trait. PPO2 sequences alignment revealed the absence of known mutations conferring resistance to PPO-inhibitors in KCTR Palmer amaranth, and differential expression of the PPO2 gene did not occur. KCTR metabolized fomesafen faster than the susceptible population, indicating that herbicide detoxification is the mechanism conferring resistance in this population. Further, treatment with the cytochrome P450-inhibitor malathion followed by lactofen restored the sensitivity of KCTR to this herbicide. Despite being resistant to POST applied PPO-inhibitors, KCTR Palmer amaranth was completely controlled by the labeled rate of the PRE applied PPO-inhibitors fomesafen, flumioxazin, saflufenacil, sulfentrazone, and oxadiazon. The overall results suggest that P450-mediated metabolism confers resistance to PPO-inhibitors in KCTR, rather than alterations in the PPO2, which were more commonly found in other Palmer amaranth populations. Future work will focus on identifying the fomesafen metabolites and on unravelling the genetic basis of metabolic resistance to PPO-inhibitor herbicides in KCTR Palmer amaranth.

Glyphosate hormesis effects on the vegetative and reproductive development of glyphosate-susceptible and -resistant Conyza sumatrensis biotypes.

Low glyphosate doses that produce hormesis may alter the susceptibility to herbicides of weeds or enhance their propagation and dispersal. The objective of this work was to evaluate the hormetic effects of glyphosate on the vegetative, phenological and reproductive development in resistant (R) and susceptible (S) Conyza sumatrensis biotypes. The glyphosate resistance level of biotype R was 11.2-fold compared to the S biotype. Glyphosate doses <11.25 g ae ha-1 induced temporary and permanent hormetic effects for the number of leaves, plant height and dry mass accumulation up to 28 d after application in both R and S biotypes. The S biotype required 15-19% fewer thermal units at 1.4 and 2.8 g ae ha-1 glyphosate than untreated plants to reach the bolting stage. Also, this biotype had less thermal units associated with the appearance (1225 vs 1408 units) and opening (1520 vs 1765 units) of the first capitulum than the R biotype. In addition, glyphosate affected reproductive traits of both biotypes compared to their controls, increasing the number of capitulum's and seeds per plant up to 37 and 41% (at 2.8 and 0.7 g ae h-1, respectively) in the S biotype, and by 48 and 114% (both at 5.6 g ae ha-1) in the R biotype. Depending on environmental parameters, glyphosate may or may not cause hormetic effects on the vegetative and phenological development of C. sumatrenis biotypes; however, this herbicide increases the speed and fecundity of reproduction, regardless of the glyphosate susceptibility level, which can alter the population dynamics and glyphosate susceptibility of future generations.

Trp-574-Leu mutation and metabolic resistance by cytochrome P450 gene conferred high resistance to ALS-inhibiting herbicides in Descurainia sophia.

Descurainia sophia (flixweed) is a troublesome weed in winter wheat fields in North China. Resistant D. sophia populations with different acetolactate synthetase (ALS) mutations have been reported in recent years. In addition, metabolic resistance to ALS-inhibiting herbicides has also been identified. In this study, we collected and purified two resistant D. sophia populations (R1 and R2), which were collected from winter wheat fields where tribenuron-methyl provided no control of D. sophia at 30 g a.i. ha-1. Whole plant bioassay and ALS activity assay results showed the R1 and R2 populations had evolved high-level resistance to tribenuron-methyl and florasulam and cross-resistance to imazethapyr and pyrithiobac­sodium. The two ALS genes were cloned from the leaves of R1 and R2 populations, ALS1 (2004 bp) and ALS2 (1998 bp). A mutation of Trp 574 to Leu in ALS1 was present in both R1 and R2. ALS1 and ALS2 were cloned from R1 and R2 populations respectively and transferred into Arabidopsis thaliana. Homozygous T3 transgenic seedlings with ALS1 of R1 or R2 were resistant to ALS-inhibiting herbicides and the resistant levels were the same. Transgenic seedlings with ALS2 from R1 or R2 were susceptible to ALS-inhibiting herbicides. Treatment with cytochrome P450 inhibitor malathion decreased the resistant levels to tribenuron-methyl in R1 and R2. RNA-Seq was used to identify target cytochrome P450 genes possibly involved in resistance to ALS-inhibiting herbicides. There were five up-regulated differentially expressed cytochrome P450 genes: CYP72A15, CYP83B1, CYP81D8, CYP72A13 and CYP71A12. Among of them, CYP72A15 had the highest expression level in R1 and R2 populations. The R1 and R2 populations of D. sophia have evolved resistance to ALS-inhibiting herbicides due to Trp 574 Leu mutation in ALS1 and possibly other mechanisms. The resistant function of CYP72A15 needs further research.

Fenoxaprop-P-ethyl, mesosulfuron-ethyl, and isoproturon resistance status in Beckmannia syzigachne from wheat fields across Anhui Province, China.

Severe infestations of American sloughgrass (Beckmannia syzigachne (Steud.) Fernald) in wheat fields throughout Anhui Province, China, pose a significant threat to local agricultural production. This study aims to evaluate the susceptibility of 37 B. syzigachne populations collected from diverse wheat fields in Anhui Province to three commonly used herbicides: fenoxaprop-P-ethyl, mesosulfuron-ethyl, and isoproturon. Single-dose testing revealed that out of the 37 populations, 31, 26, and 11 populations had either evolved or were evolving resistance to fenoxaprop-P-ethyl, mesosulfuron-ethyl, and isoproturon, respectively. Among them, 25 populations displayed concurrent resistance to both fenoxaprop-P-ethyl and mesosulfuron-ethyl, while eight exhibited resistance to all three tested herbicides. Whole-plant bioassays confirmed that approximately 84% of the fenoxaprop-P-ethyl-resistant populations manifested high-level resistance (resistance index (RI) ≥10); 62% of the mesosulfuron-ethyl-resistant populations and 82% of the isoproturon-resistant populations exhibited low- to moderate-level resistance (2 ≤ RI <10). Three distinct target-site mutations were identified in 27% of fenoxaprop-P-ethyl-resistant populations, with no known resistance mutations detected in the remaining herbicide-resistant populations. The inhibition of cytochrome P450s (P450s) and/or glutathione S-transferases (GSTs) substantially increased susceptibility in the majority of resistant populations lacking mutations at the herbicide target site. In conclusion, resistance to fenoxaprop-P-ethyl and mesosulfuron-ethyl was widespread in B. syzigachne within Anhui Province's wheat fields, while resistance to isoproturon was rapidly evolving due to its escalating usage. Target-site mutations were present in approximately one-third of fenoxaprop-P-ethyl-resistant populations, and alternative mechanisms involving P450s and/or GSTs could explain the resistance observed in most of the remaining populations.

Response characterization and target site mechanism study in glyphosate-resistant populations of Lolium multiflorum L. from Brazil.

Italian ryegrass (Lolium multiflorum L.) is an invasive species widely spread in croplands worldwide. The intensive use of glyphosate has resulted in the selection of resistance to this herbicide in Italian ryegrass. This work characterized the response to glyphosate of Italian ryegrass populations from the South and Southwest regions of Paraná, Brazil. A total of 44 Italian ryegrass populations were collected in farming areas, and were classified for glyphosate resistance with 75% of populations resistant to gloyphosate. Of these, 3 resistant (VT05AR, MR20AR and RN01AR) and three susceptible (VT07AS, MR05AS and RN01AS) of these populations were selected to determine the resistance level and the involvement of the target site mechanisms for glyphosate resistance. Susceptible populations GR50 ranged from 165.66 to 218.17 g.e.a. ha-1 and resistant populations from 569.37 to 925.94, providing RI ranging from 2.88 and 4.70. No mutation in EPSPS was observed in the populations, however, in two (MR20AR and RN02AR) of the three resistant populations, an increase in the number of copies of the EPSPs gene (11 to 57×) was detected. The number of copies showed a positive correlation with the gene expression (R2 = 0.86) and with the GR50 of the populations (R2 = 0.81). The increase in EPSPS gene copies contributes to glyphosate resistance in Italian ryegrass populations from Brazil.

Inheritance and stacking effect of mutant ALS genes in Schoenoplectiella juncoides (Roxb.) Lye (Cyperaceae).

Schoenoplectiella juncoides, a noxious sedge weed in Japanese rice paddy, has two ALS genes, and ALS-inhibitor-resistant plants have a mutation in one of the ALS genes. The authors aimed (a) to quantitate the effect of the number of mutant alleles of ALS genes on whole-plant resistance of S. juncoides and (b) to clarify a mode of inheritance of the resistance by investigating resistance levels of the progenies of a hybrid between two S. juncoides plants with Trp574Leu substitution in different ALS. A dose-response analysis on the parental lines and the F1 population suggested that the two ALS genes contribute equally to whole-plant resistant levels. A dose-response study on the F2 population indicated that it could be classified into five groups based on the sensitivities to metsulfuron-methyl. The five groups (in ascending order of resistance levels) were considered to have zero, one, two, three, and four mutant alleles. The stacking effect of mutant alleles on resistance enhancement was more significant when the number of mutant alleles was low than when it was high; in other words, each additional mutant allele stacking increases plant resistance, but the effect saturates as the number of mutant alleles increases. A chi-square test supported that the segregation ratio of the five groups corresponds to 1:4:6:4:1 of Mendelian independence for the two ALS loci.

Gene flow and spontaneous mutations are responsible for imidazolinone herbicide-resistant weedy rice (Oryza sativa L.).

For more than two decades, weedy rice (Oryza sativa L.) has been controlled in rice fields by using imidazolinone (IMI) herbicide-resistant rice technology (Clearfield®). Outcrossing in weedy rice populations and spontaneous mutations are potential problems with herbicide-resistant crop management technologies, such as the IMI-resistant rice. The aim of this study was to characterize the mechanism of IMI herbicide resistance in weedy rice through dose-response bioassay study and evaluating amino acid substitutions in acetolactate synthase (ALS) protein. A total of 118 suspected IMI-resistant weedy rice samples, which survived in the field after an IMI herbicide application, were collected at harvest time from Türkiye in 2020 and 2021. Single-dose imazamox application experiment revealed that 38 plants survived herbicide treatment. The imazamox resistance of the surviving plants was confirmed by dose-response experiment. ALS gene region underwent a sanger DNA partial sequencing. No substitution was found in 10 samples, however, amino acid substitutions were found in 26 samples with S563N, one sample with S653T, and one sample with E630D. The S653N point is the same substitution point that serves as the origin of resistance for the Clearfield® rice varieties that are commonly cultivated in the region. It has been hypothesized that the gene flow from IMI-resistant rice may be the cause of resistance in the IMI resistant weedy rice samples with S653N. The other substitution, S653T, were considered spontaneous mutation to IMI resistance. Interestingly, the S653T mutation was detected for the first time in weedy rice. The mechanism of resistance of 10 resistant weedy rice was not confirmed in this study, however, it may be a non-target resistance or another mutation point in target site, but evidently, they did not acquire resistance by gene flow from IMI-resistant rice. It has been concluded that the effectiveness of IMI-resistant rice technology in controlling weedy rice has drastically decreased due to possible gene flow, spontaneous mutation and non-target resistance. In addition to cultural controls like clean seed, clean machinery and crop rotation, other herbicide-tolerant rice systems such as Provisia® and Roxy-RPS® rice are needed to create a diverse weedy rice management ensemble available for rice production and move towards sustainable rice farming.