Current status of community resources and priorities for weed genomics research.

Weeds are attractive models for basic and applied research due to their impacts on agricultural systems and capacity to swiftly adapt in response to anthropogenic selection pressures. Currently, a lack of genomic information precludes research to elucidate the genetic basis of rapid adaptation for important traits like herbicide resistance and stress tolerance and the effect of evolutionary mechanisms on wild populations. The International Weed Genomics Consortium is a collaborative group of scientists focused on developing genomic resources to impact research into sustainable, effective weed control methods and to provide insights about stress tolerance and adaptation to assist crop breeding.

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.

RNA and protein biomarkers for detecting enhanced metabolic resistance to herbicides mesosulfuron-methyl and fenoxaprop-ethyl in black-grass (Alopecurus myosuroides).

BACKGROUND: The evolution of non-target site resistance (NTSR) to herbicides leads to a significant reduction in herbicide control of agricultural weed species. Detecting NTSR in weed populations prior to herbicide treatment would provide valuable information for effective weed control. While not all NTSR mechanisms have been fully identified, enhanced metabolic resistance (EMR) is one of the better studied, conferring tolerance through increased herbicide detoxification. Confirming EMR towards specific herbicides conventionally involves detecting metabolites of the active herbicide molecule in planta, but this approach is time-consuming and requires access to well-equipped laboratories. RESULTS: In this study, we explored the potential of using molecular biomarkers to detect EMR before herbicide treatment in black-grass (Alopecurus myosuroides). We tested the reliability of selected biomarkers to predict EMR and survival after herbicide treatments in both reference and 27 field-derived black-grass populations collected from sites across the UK. The combined analysis of the constitutive expression of biomarkers and metabolism studies confirmed three proteins, namely, AmGSTF1, AmGSTU2 and AmOPR1, as differential biomarkers of EMR toward the herbicides fenoxaprop-ethyl and mesosulfuron in black-grass. CONCLUSION: Our findings demonstrate that there is potential to use molecular biomarkers to detect EMR toward specific herbicides in black-grass without reference to metabolism analysis. However, biomarker development must include testing at both transcript and protein levels in order to be reliable indicators of resistance. This work is a first step towards more robust resistance biomarker development, which could be expanded into other herbicide chemistries for on-farm testing and monitoring EMR in uncharacterised black-grass populations. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The role of interspecific variability and herbicide pre-adaptation in the cinmethylin response of Alopecurus myosuroides.

BACKGROUND: Cinmethylin is an inhibitor of plant fatty acid biosynthesis, with in-plant activity caused by its binding to fatty acid thioesterases (FATs). The recent registration of cinmethylin for pre-emergence herbicidal use in the UK represents a new mode-of-action (MOA) for control of the grassweed blackgrass (Alopecurus myosuroides). To date there is little published information on the extent of blackgrass' inter-population variability in sensitivity to cinmethylin, nor on any potential effect of existing non-target-site resistance (NTSR) mechanisms on cinmethylin efficacy. RESULTS: Here we present a study of variability in cinmethylin sensitivity amongst 97 UK blackgrass populations. We demonstrate that under controlled conditions, a UK field-rate dose of 500 g ha-1 provides effective control of the tested populations. Nevertheless, we reveal significant inter-population variability at doses below this rate, with populations previously characterised as strongly NTSR displaying the lowest sensitivity to cinmethylin. Assessment of paired resistant 'R' and sensitive 'S' lines from standardised genetic backgrounds confirms that selection for NTSR to the acetyl-CoA-carboxylase inhibitor fenoxaprop, and the microtubule assembly inhibitor pendimethalin, simultaneously results in reduced sensitivity to cinmethylin at doses below 500 g ha-1. Whilst we find no resistance to the field-rate dose, we reveal that cinmethylin sensitivity can be further reduced through experimental selection with cinmethylin. CONCLUSION: Cinmethylin therefore represents a much-needed further MOA for blackgrass control, but needs to be carefully managed within a resistance monitoring and integrated weed management (IWM) framework to maximise the effective longevity of this compound. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

The blackgrass genome reveals patterns of non-parallel evolution of polygenic herbicide resistance.

Globally, weedy plants are a major constraint to sustainable crop production. Much of the success of weeds rests with their ability to rapidly adapt in the face of human-mediated management of agroecosystems. Alopecurus myosuroides (blackgrass) is a widespread and impactful weed affecting agriculture in Europe. Here we report a chromosome-scale genome assembly of blackgrass and use this reference genome to explore the genomic/genetic basis of non-target site herbicide resistance (NTSR). Based on our analysis of F2 seed families derived from two distinct blackgrass populations with the same NTSR phenotype, we demonstrate that the trait is polygenic and evolves from standing genetic variation. We present evidence that selection for NTSR has signatures of both parallel and non-parallel evolution. There are parallel and non-parallel changes at the transcriptional level of several stress- and defence-responsive gene families. At the genomic level, however, the genetic loci underpinning NTSR are different (non-parallel) between seed families. We speculate that variation in the number, regulation and function of stress- and defence-related gene families enable weedy species to rapidly evolve NTSR via exaptation of genes within large multi-functional gene families. These results provide novel insights into the potential for, and nature of plant adaptation in rapidly changing environments.

Dissecting weed adaptation: Fitness and trait correlations in herbicide-resistant Alopecurus myosuroides.

BACKGROUND: Unravelling the genetic architecture of non-target-site resistance (NTSR) traits in weed populations can inform questions about the inheritance, trade-offs and fitness costs associated with these traits. Classical quantitative genetics approaches allow study of the genetic architecture of polygenic traits even where the genetic basis of adaptation remains unknown. These approaches have the potential to overcome some of the limitations of previous studies into the genetics and fitness of NTSR. RESULTS: Using a quantitative genetic analysis of 400 pedigreed Alopecurus myosuroides seed families from nine field-collected populations, we found strong heritability for resistance to the acetolactate synthase and acetyl CoA carboxylase inhibitors (h2  = 0.731 and 0.938, respectively), and evidence for shared additive genetic variance for resistance to these two different herbicide modes of action, rg  = 0.34 (survival), 0.38 (biomass). We find no evidence for genetic correlations between life-history traits and herbicide resistance, indicating that resistance to these two modes of action is not associated with large fitness costs in blackgrass. We do, however, demonstrate that phenotypic variation in plant flowering characteristics is heritable, h2  = 0.213 (flower height), 0.529 (flower head number), 0.449 (time to flowering) and 0.372 (time to seed shed), demonstrating the potential for adaptation to other nonchemical management practices (e.g. mowing of flowering heads) now being adopted for blackgrass control. CONCLUSION: These results highlight that quantitative genetics can provide important insight into the inheritance and genetic architecture of NTSR, and can be used alongside emerging molecular techniques to better understand the evolutionary and fitness landscape of herbicide resistance. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Investigating the origins and evolution of a glyphosate-resistant weed invasion in South America.

The global invasion, and subsequent spread and evolution of weeds provides unique opportunities to address fundamental questions in evolutionary and invasion ecology. Amaranthus palmeri is a widespread glyphosate-resistant (GR) weed in the USA. Since 2015, GR populations of A. palmeri have been confirmed in South America, raising questions about introduction pathways and the importance of pre- vs. post-invasion evolution of GR traits. We used RAD-sequencing genotyping to characterize genetic structure of populations from Brazil, Argentina, Uruguay and the USA. We also quantified gene copy number of the glyphosate target, 5-enolpyruvyl-3-shikimate phosphate synthase (EPSPS), and the presence of an extrachromosomal circular DNA (eccDNA) replicon known to confer glyphosate resistance in USA populations. Populations in Brazil, Argentina and Uruguay were only weakly differentiated (pairwise FST  ≤0.043) in comparison to USA populations (mean pairwise FST  =0.161, range =0.068-0.258), suggesting a single major invasion event. However, elevated EPSPS copy number and the EPSPS replicon were identified in all populations from Brazil and Uruguay, but only in a single Argentinean population. These observations are consistent with independent in situ evolution of glyphosate resistance in Argentina, followed by some limited recent migration of the eccDNA-based mechanism from Brazil to Argentina. Taken together, our results are consistent with an initial introduction of A. palmeri into South America sometime before the 1980s, and local evolution of GR in Argentina, followed by a secondary invasion of GR A. palmeri with the unique eccDNA-based mechanism from the USA into Brazil and Uruguay during the 2010s.

Genomic-based epidemiology reveals independent origins and gene flow of glyphosate resistance in Bassia scoparia populations across North America.

Genomic-based epidemiology can provide insight into the origins and spread of herbicide resistance mechanisms in weeds. We used kochia (Bassia scoparia) populations resistant to the herbicide glyphosate from across western North America to test the alternative hypotheses that (i) a single EPSPS gene duplication event occurred initially in the Central Great Plains and then subsequently spread to all other geographical areas now exhibiting glyphosate-resistant kochia populations or that (ii) gene duplication occurred multiple times in independent events in a case of parallel evolution. We used qPCR markers previously developed for measuring the structure of the EPSPS tandem duplication to investigate whether all glyphosate-resistant individuals had the same EPSPS repeat structure. We also investigated population structure using simple sequence repeat markers to determine the relatedness of kochia populations from across the Central Great Plains, Northern Plains and the Pacific Northwest. We found that the original EPSPS duplication genotype was predominant in the Central Great Plains where glyphosate resistance was first reported. We identified two additional EPSPS duplication genotypes, one having geographical associations with the Northern Plains and the other with the Pacific Northwest. The EPSPS duplication genotype from the Pacific Northwest seems likely to represent a second, independent evolutionary origin of a resistance allele. We found evidence of gene flow across populations and a general lack of population structure. The results support at least two independent evolutionary origins of glyphosate resistance in kochia, followed by substantial and mostly geographically localized gene flow to spread the resistance alleles into diverse genetic backgrounds.

Fitness Cost Associated With Enhanced EPSPS Gene Copy Number and Glyphosate Resistance in an Amaranthus tuberculatus Population.

The evolution of resistance to pesticides in agricultural systems provides an opportunity to study the fitness costs and benefits of novel adaptive traits. Here, we studied a population of Amaranthus tuberculatus (common waterhemp), which has evolved resistance to glyphosate. The growth and fitness of seed families with contrasting levels of glyphosate resistance was assessed in the absence of glyphosate to determine their ability to compete for resources under intra- and interspecific competition. We identified a positive correlation between the level of glyphosate resistance and gene copy number for the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) glyphosate target, thus identifying gene amplification as the mechanism of resistance within the population. Resistant A. tuberculatus plants were found to have a lower competitive response when compared to the susceptible phenotypes with 2.76 glyphosate resistant plants being required to have an equal competitive effect as a single susceptible plant. A growth trade-off was associated with the gene amplification mechanism under intra-phenotypic competition where 20 extra gene copies were associated with a 26.5 % reduction in dry biomass. Interestingly, this growth trade-off was mitigated when assessed under interspecific competition from maize.

Adaptation at different points along antibiotic concentration gradients.

Antibiotic concentrations vary dramatically in the body and the environment. Hence, understanding the dynamics of resistance evolution along antibiotic concentration gradients is critical for predicting and slowing the emergence and spread of resistance. While it has been shown that increasing the concentration of an antibiotic slows resistance evolution, how adaptation to one antibiotic concentration correlates with fitness at other points along the gradient has not received much attention. Here, we selected populations of Escherichia coli at several points along a concentration gradient for three different antibiotics, asking how rapidly resistance evolved and whether populations became specialized to the antibiotic concentration they were selected on. Populations selected at higher concentrations evolved resistance more slowly but exhibited equal or higher fitness across the whole gradient. Populations selected at lower concentrations evolved resistance rapidly, but overall fitness in the presence of antibiotics was lower. However, these populations readily adapted to higher concentrations upon subsequent selection. Our results indicate that resistance management strategies must account not only for the rates of resistance evolution but also for the fitness of evolved strains.

Review: Allelochemicals as multi-kingdom plant defence compounds: towards an integrated approach.

The capability of synthetic pesticides to manage weeds, insect pests and pathogens in crops has diminished due to evolved resistance. Sustainable management is thus becoming more challenging. Novel solutions are needed and, given the ubiquity of biologically active secondary metabolites in nature, such compounds require further exploration as leads for novel crop protection chemistry. Despite improving understanding of allelochemicals, particularly in terms of their potential for use in weed control, their interactions with multiple biotic kingdoms have to date largely been examined in individual compounds and not as a recurrent phenomenon. Here, multi-kingdom effects in allelochemicals are introduced by defining effects on various organisms, before exploring current understanding of the inducibility and possible ecological roles of these compounds with regard to the evolutionary arms race and dose-response relationships. Allelochemicals with functional benefits in multiple aspects of plant defence are described. Gathering these isolated areas of science under the unified umbrella of multi-kingdom allelopathy encourages the development of naturally-derived chemistries conferring defence to multiple discrete biotic stresses simultaneously, maximizing benefits in weed, insect and pathogen control, while potentially circumventing resistance. © 2020 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Population genomics of selectively neutral genetic structure and herbicide resistance in UK populations of Alopecurus myosuroides.

BACKGROUND: Alopecurus myosuroides (blackgrass) is a major weed in Europe with known resistance to multiple herbicide modes of action. In the UK, there is evidence that blackgrass has undergone a range expansion. In this paper, genotyping-by-sequencing and population-level herbicide resistance phenotypes are used to explore spatial patterns of selectively neutral genetic variation and resistance. We also perform a preliminary genome-wide association study (GWAS) and genomic prediction analysis to evaluate the potential of these approaches for investigating nontarget site herbicide resistance. RESULTS: Blackgrass was collected from 47 fields across the British Isles and up to eight plants per field population (n = 369) were genotyped by Restriction site-associated DNA (RAD)-sequencing. A total of 20 426 polymorphic loci were identified and used for population genetic analyses. Phenotypic assays revealed significant variation in herbicide resistance between populations. Population structure was weak (FST = 0.024-0.048), but spatial patterns were consistent with an ongoing westward and northward range expansion. We detected strong and consistent Wahlund effects (FIS = 0.30). There were no spatial patterns of herbicide resistance or evidence for confounding with population structure. Using a combination of population-level GWAS and genomic prediction we found that the top 20, 200, and 2000 GWAS loci had higher predictive abilities for fenoxaprop resistance compared to all markers. CONCLUSION: There is likely extensive human-mediated gene flow between field populations of the weed blackgrass at a national scale. The lack of confounding of adaptive and neutral genetic variation can enable future, more extensive GWAS analyses to identify the genetic architecture of evolved herbicide resistance. © 2020 Society of Chemical Industry.

Publisher Correction: Evolution of generalist resistance to herbicide mixtures reveals a trade-off in resistance management.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Evolution of generalist resistance to herbicide mixtures reveals a trade-off in resistance management.

Intense selection by pesticides and antibiotics has resulted in a global epidemic of evolved resistance. In agriculture and medicine, using mixtures of compounds from different classes is widely accepted as optimal resistance management. However, this strategy may promote the evolution of more generalist resistance mechanisms. Here we test this hypothesis at a national scale in an economically important agricultural weed: blackgrass (Alopecurus myosuroides), for which herbicide resistance is a major economic issue. Our results reveal that greater use of herbicide mixtures is associated with lower levels of specialist resistance mechanisms, but higher levels of a generalist mechanism implicated in enhanced metabolism of herbicides with diverse modes of action. Our results indicate a potential evolutionary trade-off in resistance management, whereby attempts to reduce selection for specialist resistance traits may promote the evolution of generalist resistance. We contend that where specialist and generalist resistance mechanisms co-occur, similar trade-offs will be evident, calling into question the ubiquity of resistance management based on mixtures and combination therapies.

The costs of human-induced evolution in an agricultural system.

Pesticides have underpinned significant improvements in global food security, albeit with associated environmental costs. Currently, the yield benefits of pesticides are threatened as overuse has led to wide-scale evolution of resistance. Yet despite this threat, there are no large-scale estimates of crop yield losses or economic costs due to resistance. Here, we combine national-scale density and resistance data for the weed Alopecurus myosuroides (black-grass) with crop yield maps and a new economic model to estimate that the annual cost of resistance in England is £0.4bn in lost gross profit (2014 prices), and annual wheat yield loss due to resistance is 0.8 million tonnes. A total loss of herbicide control against black-grass would cost £1bn and 3.4 million tonnes of lost wheat yield annually. Worldwide, there are 253 herbicide-resistant weeds, so the global impact of resistance could be enormous. Our research provides an urgent case for national-scale planning to combat further evolution of resistance, and an incentive for policies focused on increasing yields through more sustainable food-production systems rather than relying so heavily on herbicides.

Alterations in Life-History Associated With Non-target-site Herbicide Resistance in Alopecurus myosuroides.

The evolution of resistance to herbicides is a classic example of rapid contemporary adaptation in the face of a novel environmental stress. Evolutionary theory predicts that selection for resistance will be accompanied by fitness trade-offs in environments where the stress is absent. Alopecurus myosuroides, an autumn-germinating grass weed of cereal crops in North-West Europe, has evolved resistance to seven herbicide modes-of-action, making this an ideal species to examine the presence and magnitudes of such fitness costs. Here, we use two contrasting A. myosuroides phenotypes derived from a common genetic background, one with enhanced metabolism resistance to a commercial formulation of the sulfonylurea (ALS) actives mesosulfuron and iodosulfuron, and the other with susceptibility to these actives (S). Comparisons of plant establishment, growth, and reproductive potential were made under conditions of intraspecific competition, interspecific competition with wheat, and over a gradient of nitrogen deprivation. Herbicide dose response assays confirmed that the two lines had contrasting resistance phenotypes, with a 20-fold difference in resistance between them. Pleiotropic effects of resistance were observed during plant development, with R plants having a greater intraspecific competitive effect and longer tiller lengths than S plants during vegetative growth, but with S plants allocating proportionally more biomass to reproductive tissues during flowering. Direct evidence of a reproductive cost of resistance was evident in the nitrogen deprivation experiment with R plants producing 27% fewer seed heads per plant, and a corresponding 23% reduction in total seed head length. However, these direct effects of resistance on fecundity were not consistent across experiments. Our results demonstrate that a resistance phenotype based on enhanced herbicide metabolism has pleiotropic impacts on plant growth, development and resource partitioning but does not support the hypothesis that resistance is associated with a consistent reproductive fitness cost in this species. Given the continued difficulties associated with unequivocally detecting costs of herbicide resistance, we advocate future studies that adopt classical evolutionary quantitative genetics approaches to determine genetic correlations between resistance and fitness-related plant life history traits.

Molecular mechanisms of adaptive evolution revealed by global selection for glyphosate resistance.

The human-directed, global selection for glyphosate resistance in weeds has revealed a fascinating diversity of evolved resistance mechanisms, including herbicide sequestration in the vacuole, a rapid cell death response, nucleotide polymorphisms in the herbicide target (5-enolpyruvylshikimate-3-phosphate synthase, EPSPS) and increased gene copy number of EPSPS. For this latter mechanism, two distinct molecular genetic mechanisms have been observed, a tandem duplication mechanism and a large extrachromosomal circular DNA (eccDNA) that is tethered to the chromosomes and passed to gametes at meiosis. These divergent mechanisms have a range of consequences for the spread, fitness, and inheritance of resistance traits, and, particularly in the case of the eccDNA, demonstrate how evolved herbicide resistance can generate new insights into plant adaptation to contemporary environmental stress.

Evolutionary epidemiology predicts the emergence of glyphosate resistance in a major agricultural weed.

The evolution of resistance to herbicides is a striking example of rapid, human-directed adaptation with major consequences for food production. Most studies of herbicide resistance are performed reactively and focus on post hoc determination of resistance mechanisms following the evolution of field resistance. If the evolution of resistance can be anticipated, however, pro-active management to slow or prevent resistance traits evolving can be advocated. We report a national-scale study that combines population monitoring, glyphosate sensitivity assays, quantitative genetics and epidemiological analyses to pro-actively identify the prerequisites for adaptive evolution (directional selection and heritable genetic variation) to the world's most widely used herbicide (glyphosate) in a major, economically damaging weed species, Alopecurus myosuroides. Results highlighted pronounced, heritable variability in glyphosate sensitivity amongst UK A. myosuroides populations. We demonstrated a direct epidemiological link between historical glyphosate selection and current population-level sensitivity, and show that current field populations respond to further glyphosate selection. This study provides a novel, pro-active assessment of adaptive potential for herbicide resistance, and provides compelling evidence of directional selection for glyphosate insensitivity in advance of reports of field resistance. The epidemiological approach developed can provide a basis for further pro-active study of resistance evolution across pesticide resistance disciplines.

The evolutionary origins of pesticide resistance.

Durable crop protection is an essential component of current and future food security. However, the effectiveness of pesticides is threatened by the evolution of resistant pathogens, weeds and insect pests. Pesticides are mostly novel synthetic compounds, and yet target species are often able to evolve resistance soon after a new compound is introduced. Therefore, pesticide resistance provides an interesting case of rapid evolution under strong selective pressures, which can be used to address fundamental questions concerning the evolutionary origins of adaptations to novel conditions. We ask: (i) whether this adaptive potential originates mainly from de novo mutations or from standing variation; (ii) which pre-existing traits could form the basis of resistance adaptations; and (iii) whether recurrence of resistance mechanisms among species results from interbreeding and horizontal gene transfer or from independent parallel evolution. We compare and contrast the three major pesticide groups: insecticides, herbicides and fungicides. Whilst resistance to these three agrochemical classes is to some extent united by the common evolutionary forces at play, there are also important differences. Fungicide resistance appears to evolve, in most cases, by de novo point mutations in the target-site encoding genes; herbicide resistance often evolves through selection of polygenic metabolic resistance from standing variation; and insecticide resistance evolves through a combination of standing variation and de novo mutations in the target site or major metabolic resistance genes. This has practical implications for resistance risk assessment and management, and lessons learnt from pesticide resistance should be applied in the deployment of novel, non-chemical pest-control methods.

Gene drive systems: do they have a place in agricultural weed management?

There is a pressing need for novel control techniques in agricultural weed management. Direct genetic control of agricultural pests encompasses a range of techniques to introduce and spread novel, fitness-reducing genetic modifications through pest populations. Recently, the development of CRISPR-Cas9 gene editing has brought these approaches into sharper focus. Proof of concept for CRISPR-Cas9-based gene drives has been demonstrated for the control of disease-vectoring insects. This article considers whether and how gene drives may be applied in agricultural weed management, focusing on CRISPR-Cas9-based systems. Population-suppression drives might be employed to introduce and proliferate deleterious mutations that directly impact fitness and weediness, whereas population-sensitizing drives would seek to edit weed genomes so that populations are rendered more sensitive to subsequent management interventions. Technical challenges relating to plant transformation and gene editing in planta are considered, and the implementation of gene drives for timely and sustainable weed management is reviewed in the light of weed population biology. The technical, biological, practical and regulatory challenges remain significant. Modelling-based studies can inform how and if gene drives could be employed in weed populations. These studies are an essential first step towards determining the utility of gene drives for weed management. © 2018 The Author. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.