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.

Soil organic carbon, aggregation and fungi community after 44 years of no-till and cropping systems in the Central Great Plains, USA.

Implementing sustainable agricultural land management practices such as no-till (NT) and diversified crops are important for maintaining soil health properties. This study focuses on the soil health of three long-term (44 years) tillage systems, NT, reduced tillage (RT), and conventional tillage (CT), in monoculture winter wheat-fallow (W-F) (Triticum aestivum L.) and wheat-soybean (W-S) (Glycine max (L.) Merrill) rotation. Soil organic carbon (C) was higher in NT than CT in the surface 0-5 cm, but not different in the 5-15 cm, demonstrating SOC stratification on the soil profile. The soil water content was higher in NT followed by RT and CT in the top 0-5 cm. We found an association between increased carbon, aggregation, and AMF biomass. Greater soil aggregation, carbon and AMF were observed in NT at 0-5 cm soil depth. The W-S cropping system had greater soil microbial community composition based on fungi biomass, AMF and fungal to bacteria ratio from phospholipid fatty acid analysis (PLFA). Large macroaggregates were positively correlated with total C and N, microbial biomass, Gram + , and AMF. Soil water content was positively correlated with macroaggregates, total C and N, and AC. No-till increased soil carbon content even after 44 years of cultivation. By implementing conservation tillage systems and diversified crop rotation, soil quality can be improved through greater soil organic C, water content, greater soil structure, and higher AMF biomass than CT practice in the Central Great Plains.

Inheritance of 2,4-dichlorophenoxyacetic acid (2,4-D) resistance in Amaranthus palmeri.

In this study, the inheritance of 2,4-D resistance in a multiple herbicide-resistant Palmer amaranth (KCTR) was investigated. Direct and reciprocal crosses were performed using 2,4-D-resistant KCTR and susceptible KSS plants to generate F1 progenies. 2,4-D dose-response assays were conducted to evaluate the response of progenies from each F1 family along with KCTR and KSS plants in controlled environmental growth chambers. Additionally, 2,4-D-resistant male and female plants from each of the F1 families were used in pairwise crosses to generate pseudo-F2 families. Segregation (resistance or susceptibility) of progenies from the F2 families in response to a discriminatory rate of 2,4-D (i.e., 560 g ae ha-1) was evaluated. Dose-response analysis of F1 progenies derived from direct and reciprocal crosses suggested that the 2,4-D resistance in KCTR is a nuclear trait. Chi-square analyses of F2 segregation data implied that 2,4-D resistance in KCTR is controlled by multiple gene(s). Overall, our data suggest that the 2,4-D resistance in KCTR Palmer amaranth is a nuclear inherited trait controlled by multiple genes. Such resistance can spread both via pollen or seed-mediated gene flow. In future, efforts will be directed towards identifying genes mediating 2,4-D resistance in KCTR population.

Corrigendum: Predominance of Metabolic Resistance in a Six-Way-Resistant Palmer Amaranth (Amaranthus palmeri) Population.

[This corrects the article DOI: 10.3389/fpls.2020.614618.].

Predominance of Metabolic Resistance in a Six-Way-Resistant Palmer Amaranth (Amaranthus palmeri) Population.

Evolution of multiple herbicide resistance in Palmer amaranth across the United States is a serious challenge for its management. Recently, a Palmer amaranth population (KCTR; Kansas Conservation Tillage Resistant) from a long-term conservation tillage research project in Kansas, United States, was found uncontrolled by several commonly used herbicides. Importantly, this field did not have a history of repeated use of some of the herbicides for which the KCTR Palmer amaranth population showed lack of control. The objectives of this study were to confirm the evolution of multiple resistances and determine possible mechanism(s) of resistance in KCTR Palmer amaranth plants. In response to post-emergence application, 28-100% of KCTR Palmer amaranth survived field recommended rates of 2,4-D, ALS-, PS II-, EPSPS-, PPO-, HPPD-inhibitor herbicides, or tank- or pre-mixture of PS II- and HPPD-inhibitor herbicides, confirming evolution of six-way resistance in this Palmer amaranth population. However, this population was found susceptible to the PS I- and glutamine synthetase inhibitor herbicides. Chlorsulfuron-, imazethapyr-, and atrazine-resistant plants did not show any previously reported mutation in ALS and psbA genes, the target sites of these herbicides, respectively. However, the survivors of glyphosate treatment showed amplification of EPSPS gene (up to 88 copies). The KCTR plants pretreated with cytochrome P450 or GST inhibitors along with atrazine, 2,4-D, lactofen, or mesotrione had significantly less biomass accumulation than those treated with herbicides alone. Plants treated with P450 inhibitor followed by imazethapyr showed moderate reduction of biomass in KCTR which was statistically similar to a susceptible Palmer amaranth population treated with imazethapyr. These results suggest predominance of metabolic resistance possibly mediated by cytochrome P450 and GST enzyme activity that may have predisposed the KCTR Palmer amaranth population to evolve resistance to multiple herbicides. This is the first report of evolution of six-way resistance in a single Palmer amaranth population. Appropriate management strategies, including integration of cultural, and mechanical, and herbicide mixtures, are warranted to control such Palmer amaranth populations.

Gene Duplication and Aneuploidy Trigger Rapid Evolution of Herbicide Resistance in Common Waterhemp.

An increase in gene copy number is often associated with changes in the number and structure of chromosomes, as has been widely observed in yeast and eukaryotic tumors, yet little is known about stress-induced chromosomal changes in plants. Previously, we reported that the EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) gene, the molecular target of glyphosate, was amplified at the native locus and on an extra chromosome in glyphosate-resistant Amaranthus tuberculatus Here, we report that the extra chromosome is a ring chromosome termed extra circular chromosome carrying amplified EPSPS (ECCAE). The ECCAE is heterochromatic, harbors four major EPSPS amplified foci, and is sexually transmitted to 35% of the progeny. Two highly glyphosate resistant (HGR) A. tuberculatus plants with a chromosome constitution of 2n = 32+1 ECCAE displayed soma cell heterogeneity. Some cells had secondary ECCAEs, which displayed size polymorphisms and produced novel chromosomal variants with multiple gene amplification foci. We hypothesize that the ECCAE in the soma cells of HGR A. tuberculatus plants underwent breakage-fusion-bridge cycles to generate the observed soma cell heterogeneity, including de novo EPSPS gene integration into chromosomes. Resistant soma cells with stable EPSPS amplification events as de novo insertions into chromosomes may survive glyphosate selection pressure during the sporophytic phase and are plausibly transmitted to germ cells leading to durable glyphosate resistance in A. tuberculatus This is the first report of early events in aneuploidy-triggered de novo chromosome integration by an as yet unknown mechanism, which may drive rapid adaptive evolution of herbicide resistance in common waterhemp.

Rapid detoxification via glutathione S-transferase (GST) conjugation confers a high level of atrazine resistance in Palmer amaranth (Amaranthus palmeri).

BACKGROUND: Palmer amaranth (Amaranthus palmeri) is an economically troublesome, aggressive and damaging weed that has evolved resistance to six herbicide modes of action including photosystem II (PS II) inhibitors such as atrazine. The objective of this study was to investigate the mechanism and inheritance of atrazine resistance in Palmer amaranth. RESULTS: A population of Palmer amaranth from Kansas (KSR) had a high level (160 - 198-fold more; SE ±21 - 26) of resistance to atrazine compared to the two known susceptible populations MSS and KSS, from Mississippi and Kansas, respectively. Sequence analysis of the chloroplastic psbA gene did not reveal any known mutations conferring resistance to PS II inhibitors, including the most common Ser264Gly substitution for triazine resistance. However, the KSR plants rapidly conjugated atrazine at least 24 times faster than MSS via glutathione S-transferase (GST) activity. Furthermore, genetic analyses of progeny generated from reciprocal crosses of KSR and MSS demonstrate that atrazine resistance in Palmer amaranth is a nuclear trait. CONCLUSION: Although triazine resistance in Palmer amaranth was reported more than 20 years ago in the USA, this is the first report elucidating the underlying mechanism of resistance to atrazine. The non-target-site based metabolic resistance to atrazine mediated by GST activity may predispose the Palmer amaranth populations to have resistance to other herbicide families, and the nuclear inheritance of the trait in this dioecious species further exacerbates the propensity for its rapid spread. © 2017 Society of Chemical Industry.

Physiological and Molecular Characterization of Hydroxyphenylpyruvate Dioxygenase (HPPD)-inhibitor Resistance in Palmer Amaranth (Amaranthus palmeri S.Wats.).

Herbicides that inhibit hydroxyphenylpyruvate dioxygenase (HPPD) such as mesotrione are widely used to control a broad spectrum of weeds in agriculture. Amaranthus palmeri is an economically troublesome weed throughout the United States. The first case of evolution of resistance to HPPD-inhibiting herbicides in A. palmeri was documented in Kansas (KS) and later in Nebraska (NE). The objective of this study was to investigate the mechansim of HPPD-inhibitor (mesotrione) resistance in A. palmeri. Dose response analysis revealed that this population (KSR) was 10-18 times more resistant than their sensitive counterparts (MSS or KSS). Absorbtion and translocation analysis of [14C] mesotrione suggested that these mechanisms were not involved in the resistance in A. palmeri. Importantly, mesotrione (>90%) was detoxified markedly faster in the resistant populations (KSR and NER), within 24 hours after treatment (HAT) compared to sensitive plants (MSS, KSS, or NER). However, at 48 HAT all populations metabolized the mesotrione, suggesting additional factors may contribute to this resistance. Further evaluation of mesotrione-resistant A. palmeri did not reveal any specific resistance-conferring mutations nor amplification of HPPD gene, the molecular target of mesotrione. However, the resistant populations showed 4- to 12-fold increase in HPPD gene expression. This increase in HPPD transcript levels was accompanied by increased HPPD protein expression. The significant aspects of this research include: the mesotrione resistance in A. palmeri is conferred primarily by rapid detoxification (non-target-site based) of mesotrione; additionally, increased HPPD gene expression (target-site based) also contributes to the resistance mechanism in the evolution of herbicide resistance in this naturally occurring weed species.

Physical Mapping of Amplified Copies of the 5-Enolpyruvylshikimate-3-Phosphate Synthase Gene in Glyphosate-Resistant Amaranthus tuberculatus.

Recent and rapid evolution of resistance to glyphosate, the most widely used herbicides, in several weed species, including common waterhemp (Amaranthus tuberculatus), poses a serious threat to sustained crop production. We report that glyphosate resistance in A tuberculatus was due to amplification of the 5-enolpyruvylshikimate-3-P synthase (EPSPS) gene, which encodes the molecular target of glyphosate. There was a positive correlation between EPSPS gene copies and its transcript expression. We analyzed the distribution of EPSPS copies in the genome of A tuberculatus using fluorescence in situ hybridization on mitotic metaphase chromosomes and interphase nuclei. Fluorescence in situ hybridization analysis mapped the EPSPS gene to pericentromeric regions of two homologous chromosomes in glyphosate sensitive A tuberculatus In glyphosate-resistant plants, a cluster of EPSPS genes on the pericentromeric region on one pair of homologous chromosomes was detected. Intriguingly, two highly glyphosate-resistant plants harbored an additional chromosome with several EPSPS copies besides the native chromosome pair with EPSPS copies. These results suggest that the initial event of EPSPS gene duplication may have occurred because of unequal recombination mediated by repetitive DNA. Subsequently, gene amplification may have resulted via several other mechanisms, such as chromosomal rearrangements, deletion/insertion, transposon-mediated dispersion, or possibly by interspecific hybridization. This report illustrates the physical mapping of amplified EPSPS copies in A tuberculatus.