TOLERANCE INTERVALS FOR GENE FLOW RATES FROM TRANSGENIC TO NON-TRANSGENIC WHEAT AND CORN USING A LOGISTIC REGRESSION MODEL WITH RANDOM LOCATION EFFECTS

Crop scientists and government regulators are interested in mediating pollen flow from transgenic crops to other crops and weed species. To this end, a multi-year, multilocation series of experiments was conducted in eastern Colorado by the Department of Soil and Crop Sciences at Colorado State University. These experiments were done to estimate the distance required between plots of transgenic corn and wheat and plots of the respective non-transgenic crop to obtain at most a regulated limit of cross-pollination. The experiments involved planting a rectangle of transgenic crop in the middle of a non-transgenic field and measuring the proportion of cross-pollinated crop at various distances along transects radiating in multiple directions. Gene flow to the non-transgenic crop was evaluated in wheat using herbicide tolerance and in corn using kernel color. An initial Generalized Linear Mixed Model with binomial response and logit link was estimated with independent variables: a square root transformation of distance, an additional covariate, and a random location effect. For corn, the additional covariate was transect orientation; for wheat, it was the relative heading time of the recipient variety. An enhanced model that included additional sources of variation was also examined. The analysis for both of these assumed models addresses two problems: 1) an Upper Tolerance Limit on the binomial probability of cross-pollination, which includes 100c% of the locations with 100d% confidence, at set values of the independent variables; and 2) an Upper Tolerance Limit on the distance at which 100c% of the locations will have binomial probability of cross-pollination less than a specified value, with 100d% confidence, at set values of the other independent variables. The problems are addressed using Frequentist and Bayesian methods

Synthetic auxin herbicides : finding the lock and key to weed resistance

Synthetic auxin herbicides are designed to mimic indole-3-acetic acid (IAA), an integral plant hormone affecting cell growth, development, and tropism. In this review, we explore target site genes in the auxin signaling pathway including SCFTIR1/AFB, Aux/IAA, and ARFs that are confirmed or proposed mechanisms for weed resistance to synthetic auxin herbicides. Resistance to auxin herbicides by metabolism, either by enhanced cytochrome P450 detoxification or by loss of pro-herbicide activation, is a major non-target-site resistance pathway. We speculate about potential fitness costs of resistance due to effects of resistance-conferring mutations, provide insight into the role of polyploidy in synthetic auxin resistance evolution, and address the genetic resources available for weeds. This knowledge will be the key to unlock the long-standing questions as to which components of the auxin signaling pathway are most likely to have a role in resistance evolution. We propose that an ambitious research effort into synthetic auxin herbicide/target site interactions is needed to 1) explain why some synthetic auxin chemical families have activity on certain dicot plant families but not others and 2) fully elucidate target-site cross-resistance patterns among synthetic auxin chemical families to guide best practices for resistance management

Global patterns of herbicide resistance evolution in Amaranthus spp.: an analysis comparing species, cropping regions and herbicides

Herbicide resistance in weeds is an evolutionary process. Although there is a great global diversity of weeds, independent origins of herbicide resistance evolution have been shown to converge into similar molecular and physiological resistance mechanisms in geographically distant weed populations. Amaranthus species have shown an extraordinary ability to evolve herbicide resistance and invade new environments at a global scale, which represents an opportunity for identifying adaptive evolutionary patterns. The most frequent cases of herbicide-resistant Amaranthus species have been identified in North America, where A. hybridus, A. palmeri, A. tuberculatus and A. retroflexus comprise more than 90% of them. Meanwhile, A. retroflexus, A. hybridus and A. palmeri have been the most reported species in South America. Around 70% of the cases of herbicide-resistant Amaranthus species have been identified in global soybean and corn crops. The higher fecundity and adaptability of plants to a broad range of environments would make populations more likely to persist and be selected for herbicide resistance. Co-evolution of multiple herbicide resistance mechanisms at the plant and/or population level is evident in weed species. For Amaranthus spp., resistance cases highlight evolutionary responses to herbicide use with clear patterns of selection for multiple herbicide resistance in particular regions and spread to new areas within and between global cropping systems. Seed-mediated gene flow is an important component to the spread of herbicide resistant Amaranthus spp. populations. Reduction of the intensity of herbicide selection by combining diverse and integrated weed control practices should be a common goal in weed management programs.Fil: Yanniccari, Marcos Ezequiel. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gaines, Todd. University of Colorado; Estados UnidosFil: Scursoni, Julio Alejandro. Universidad de Buenos Aires. Facultad de Agronomía; ArgentinaFil: De Prado, Rafael. Universidad de Córdoba; EspañaFil: Vila Aiub, Martin Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentin

Response of \u3ci\u3eAmaranthus\u3c/i\u3e spp. following exposure to sublethal herbicide rates via spray particle drift

The adverse consequences of herbicide drift towards sensitive crops have been extensively reported in the literature. However, little to no information is available on the consequences of herbicide drift onto weed species inhabiting boundaries of agricultural fields. Exposure to herbicide drift could be detrimental to long-term weed management as several weed species have evolved herbicide-resistance after recurrent selection with sublethal herbicide rates This study investigated the deposition of glyphosate, 2,4-D, and dicamba spray particle drift from applications with two different nozzles in a low speed wind tunnel, and their impact on growth and development of Amaranthus spp. Herbicide drift resulted in biomass reduction or complete plant mortality. Inflection points (distance to 50% biomass reduction) for Amaranthus tuberculatus were 7.7, 4.0, and 4.1 m downwind distance for glyphosate, 2,4-D, and dicamba applications with the flat-fan nozzle, respectively, whereas these values corresponded to 2.8, 2.5, and 1.9 m for applications with the air-inclusion nozzle. Inflection points for Amaranthus palmeri biomass reduction were 16.3, 10.9, and 11.5 m for glyphosate, 2,4-D, and dicamba applications with the flat-fan nozzle, respectively, whereas these values corresponded to 7.6, 5.4, and 5.4 m for applications with the air-inclusion nozzle. Plants were more sensitive to glyphosate at higher exposure rates than other herbicides, whereas plants were more sensitive to 2,4-D and dicamba at lower exposure rates compared to glyphosate. Applications with the flat-fan nozzle resulted in 32.3 and 11.5% drift of the applied rate at 1.0 and 3.0 m downwind, respectively, whereas the air-inclusion nozzle decreased the dose exposure in the same distances (11.4 and 2.7%, respectively). Herbicide drift towards field boundaries was influenced by nozzle design and exposed weeds to herbicide rates previously reported to select for herbicide-resistant biotypes

Reversing resistance to tembotrione in an Amaranthus tuberculatus (var. rudis) population from Nebraska, USA with cytochrome P450 inhibitors

Background: A population of Amaranthus tuberculatus (var. rudis) was confirmed resistant to 4-hydroxyphenylpyruvate dioxygenase (HPPD)-inhibitor herbicides (mesotrione, tembotrione, and topramezone) in a seed corn/soybean rotation in Nebraska. Further investigation confirmed a non-target-site resistance mechanism in this population. The main objective of this study was to explore the role of cytochrome P450 inhibitors in restoring the efficacy of HPPD-inhibitor herbicides on the HPPD-inhibitor resistant A. tuberculatus population from Nebraska, USA (HPPD-R). Background: A population of Amaranthus tuberculatus (var. rudis) was confirmed resistant to 4-hydroxyphenylpyruvate dioxygenase (HPPD)-inhibitor herbicides (mesotrione, tembotrione, and topramezone) in a seed corn/soybean rotation in Nebraska. Further investigation confirmed a non-target-site resistance mechanism in this population. The main objective of this study was to explore the role of cytochrome P450 inhibitors in restoring the efficacy of HPPD-inhibitor herbicides on the HPPD-inhibitor resistant A. tuberculatus population from Nebraska, USA (HPPD-R). Results: Enhanced metabolism via cytochrome P450 enzymes is the mechanism of resistance in HPPD-R. Amitrole partially restored the activity of mesotrione, whereas malathion, amitrole, and piperonyl butoxide restored the activity of tembotrione and topramezone in HPPD-R. Although corn was injured through malathion followed by mesotrione application a week after treatment, the injury was transient, and the crop recovered. Includes supplementary file

Herbicide-resistant weeds : from research and knowledge to future needs

Synthetic herbicides have been used globally to control weeds in major field crops. This has imposed a strong selection for any trait that enables plant populations to survive and reproduce in the presence of the herbicide. Herbicide resistance in weeds must be minimized because it is a major limiting factor to food security in global agriculture. This represents a huge challenge that will require great research efforts to develop control strategies as alternatives to the dominant and almost exclusive practice of weed control by herbicides. Weed scientists, plant ecologists and evolutionary biologists should join forces and work towards an improved and more integrated understanding of resistance across all scales. This approach will likely facilitate the design of innovative solutions to the global herbicide resistance challenge

On the discovery of genes involved in metabolism-based resistance to herbicides using RNA-Seq transcriptome analysis in Lolium rigidum

Weed control failures due to herbicide resistance are an increasing and worldwide problem significantly impacting crop yields. Herbicide resistance due to increased herbicide metabolism in weeds is not well characterized at the genetic level. An RNA-Seq transcriptome analysis was used to identify genes conferring metabolism-based herbicide resistance (MBHR) in a population (R) of a major global weed (Lolium rigidum), in which resistance to the herbicide diclofop-methyl was experimentally evolved through recurrent selection from a susceptible (S) progenitor population. A reference transcriptome of 19,623 contigs was assembled using 454 sequencing technology on a cDNA library and annotated using UniProt and Pfam databases. Transcriptomic-level gene expression was measured using Illumina 100 bp reads from untreated control, mock, and diclofop-methyl treatments of R and S. Due to the established importance of cytochrome P450 (CytP450), glutathione-S-transferase (GST), and glucosyltransferase (GT) genes in MBHR, 11 contigs with these annotations and higher constitutive expression in untreated R than in untreated S were selected as candidate genes for hypothesis testing, along with 17 additional differentially expressed contigs with annotations related to metabolism or signal transduction. In a forward genetics validation experiment, higher constitutive expression of nine contigs co-segregated with the resistance phenotype in an F2 population, including 3 CytP450, 3 GST, and 1 GT. At least nine genes with heritable increased constitutive expression are associated with MBHR trait. In a physiological validation experiment where 2, 4-D pre-treatment induced diclofop-methyl protection in S individuals due to increased metabolism, seven of the nine genetically-validated contigs were significantly induced. These data help explain accumulation of resistance-endowing genes and rapid evolution of MBHR, and provide the opportunity to improve diagnostics of MBHR using molecular tools such as transcriptional markers. Keywords: 2,4-Dichlorophenoxyacetic acid (2,4-D), diclofop-methyl, evolution, herbicide metabolism, herbicide resistance, next-generation sequencing, transcriptional markers Zur Entdeckung der beteiligten Gene an der metabolischen Herbizidresistenz in Lolium rigidum durch RNA-Seq Transkriptom Analyse. Zusammenfassung Herbizidresistenz ist weltweit ein zunehmendes Problem in der Landwirtschaft, vor allem der enzymatische Abbau von Wirkstoffen bzw. die metabolische Herbizidresistenz (MBHR) ist bislang noch weitgehend unbekannt, besonders auf genetischer Ebene. Um die an einer MBHR beteiligten Gene zu identifizieren wurde das Transkriptom von Herbizid-resistenten und sensitiven Weidelgräsern (Lolium rigidum) in einem RNA-Seq-Ansatz verglichen. Die verwendete Diclofop-Methyl herbizidresistente Population wurde experimentell aus einer sensitiven Population rekurrent selektiert. Ein 19623 Contig umfassendes Referenztranskriptom wurde aus einer cDNA Bibliothek der resistenten Pflanzen durch 454 Sequenzierung erstellt und mit Hilfe von UniProt und Pfam annotiert. Mit 100 bp Illumina-Reads wurde die Genexpression in unbehandelten, mit Blindformulierung und mit formuliertem Herbizid Diclofop-methyl behandelten resistenten und sensitiven Pflanzen untersucht. Es ist bekannt, dass Cytochrom P450 (CytP450), Glutathion-S-transferasen (GST) oder Glycosyltransferasen (GT) eine wichtige Rolle in der MBHR spielen. Elf Contigs, die als solche annotiert und in unbehandelten resistenten Pflanzen konstitutiv höher exprimiert sind als in Sensitiven, wurden als Kandidatengene ausgewählt und getestet. Zusätzlich wurden 17 weitere Contigs untersucht, die zwischen resistenten und sensitiven Pflanzen unterschiedlich exprimiert und durch ihre Annotation in Metabolismus oder Signaltransduktion eingebunden sind. Eine höhere konstitutive Expression konnte in neun selektierten Contigs (u.a. 3 CytP450, 3 GST, 1 GT) auch in der F2 Generation bestätigt werden, cosegregierend mit den resistenten Phänotypen. Daher ist die Expression von mindestens neun Genen mit metabolischer Diclofop-methyl Herbizidresistenz gekoppelt und vererbbar. Eine Vorbehandlung sensitiver Weidelgras-Pflanzen mit 2,4-D induziert metabolische Resistenz gegenüber dem ACCase Inhibitor Diclofop-methyl. Dies wurde verwendet, um die Expression der 28 selektierten Contigs zusätzlich physiologisch zu validieren. Sieben der neun zuvor beschriebenen Contigs sind in der physiologischen Validierung auch in sensitiven Pflanzen durch 2, 4-D induziert. Diese Ergebnisse bieten neue Diagnosemöglichkeiten und verdeutlichen den Zusammenhang zwischen einer vererbbaren Expressionsregulierung von Resistenz vermittelnden Genen und der schnellen Entwicklung von MBHR. Stichwörter: 2,4-Dichlorophenoxyessig Säure (2, 4-D), Diclofop-Methyl, Evolution, Herbizidmetabolisierung, Herbizidresistenz, Next-Generation Sequenzierung, transkriptionelle Marker, Transkriptomi

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.Fil: Gaines, Todd A. State University of Colorado - Fort Collins; Estados UnidosFil: Slavov, Gancho. No especifíca;Fil: Hughes, David. No especifíca;Fil: Kupper, Anita. State University of Colorado - Fort Collins; Estados UnidosFil: Sparks, Crystal. State University of Colorado - Fort Collins; Estados UnidosFil: Oliva, Julian. Universidad Católica de Córdoba; ArgentinaFil: Vila Aiub, Martin Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; ArgentinaFil: García, Alejandro Marcelo. Instituto Nacional de Tecnología Agropecuaria; ArgentinaFil: Merotto, Aldo. Universidade Federal do Rio Grande do Sul; BrasilFil: Neve, Paul. No especifíca

Metabolism of 2,4-dichlorophenoxyacetic acid contributes to resistance in a common waterhemp (\u3ci\u3eAmaranthus tuberculatus\u3c/i\u3e) population

BACKGROUND: Synthetic auxins such as 2,4-D have been widely used for selective control of broadleaf weeds since the mid-1940s. In 2009, an Amaranthus tuberculatus (common waterhemp) population with 10-fold resistance to 2,4-D was found in Nebraska, USA. The 2,4-D resistance mechanism was examined by conducting [14C] 2,4-D absorption, translocation and metabolism experiments. RESULTS: No differences were found in 2,4-D absorption or translocation between the resistant and susceptible A. tuberculatus. Resistant plants metabolized [14C] 2,4-D more rapidly than did susceptible plants. The half-life of [14C] 2,4-D in susceptible plants was 105 h, compared to 22 h in resistant plants. Pre-treatment with the cytochrome P450 inhibitor malathion inhibited [14C] 2,4-D metabolism in resistant plants and reduced the 2,4-D dose required for 50% growth inhibition (GR50) of resistant plants by 7-fold to 27 g ha-1, similar to the GR50 for susceptible plants in the absence of malathion. CONCLUSIONS: Our results demonstrate that rapid 2,4-D metabolism is a contributing factor to resistance in A. tuberculatus, potentially mediated by cytochrome P450. Metabolism-based resistance to 2,4-D could pose a serious challenge for A. tuberculatus control due to the potential for cross-resistance to other herbicides