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.

Chemically induced herbicide tolerance in rice by the safener metcamifen is associated with a phased stress response.

The closely related sulphonamide safeners, metcamifen and cyprosulfamide, were tested for their ability to protect rice from clodinafop-propargyl, a herbicide normally used in wheat. While demonstrating that both compounds were equally bioavailable in planta, only metcamifen prevented clodinafop from damaging seedlings, and this was associated with the enhanced detoxification of the herbicide. Transcriptome studies in rice cultures demonstrated that whereas cyprosulfamide had a negligible effect on gene expression over a 4 h exposure, metcamifen perturbed the abundance of 590 transcripts. Changes in gene expression with metcamifen could be divided into three phases, corresponding to inductions occurring over 30 min, 1.5 h and 4 h. The first phase of gene induction was dominated by transcription factors and proteins of unknown function, the second by genes involved in herbicide detoxification, while the third was linked to cellular homeostasis. Analysis of the inducible genes suggested that safening elicited similar gene families to those associated with specific biotic and abiotic stresses, notably those elicited by abscisic acid, salicylic acid, and methyl jasmonate. Subsequent experiments with safener biomarker genes induced in phase 1 and 2 in rice cell cultures provided further evidence of similarities in signalling processes elicited by metcamifen and salicylic acid.

Evolution of Target-Site Resistance to Glyphosate in an Amaranthus palmeri Population from Argentina and Its Expression at Different Plant Growth Temperatures.

The mechanism and expression of resistance to glyphosate at different plant growing temperatures was investigated in an Amaranthus palmeri population (VM1) from a soybean field in Vicuña Mackenna, Cordoba, Argentina. Resistance was not due to reduced glyphosate translocation to the meristem or to EPSPS duplication, as reported for most US samples. In contrast, a proline 106 to serine target-site mutation acting additively with EPSPS over-expression (1.8-fold increase) was respectively a major and minor contributor to glyphosate resistance in VM1. Resistance indices based on LD50 values generated using progenies from a cross between 52 PS106 VM1 individuals were estimated at 7.1 for homozygous SS106 and 4.3 for heterozygous PS106 compared with homozygous wild PP106 plants grown at a medium temperature of 24 °C day/18 °C night. A larger proportion of wild and mutant progenies survived a single commonly employed glyphosate rate when maintained at 30 °C day/26 °C night compared with 20 °C day/16 night in a subsequent experiment. Interestingly, the P106S mutation was not identified in any of the 920 plants analysed from 115 US populations, thereby potentially reflecting the difference in A. palmeri control practices in Argentina and USA.

Mechanism of resistance to mesotrione in an Amaranthus tuberculatus population from Nebraska, USA.

Amaranthus tuberculatus is a troublesome weed in corn and soybean production systems in Midwestern USA, due in part to its ability to evolve multiple resistance to key herbicides including 4-hydroxyphenylpyruvate dioxygenase (HPPD). Here we have investigated the mechanism of resistance to mesotrione, an important chemical for managing broadleaf weeds in corn, in a multiple herbicide resistant population (NEB) from Nebraska. NEB showed a 2.4-fold and 45-fold resistance increase to mesotrione compared to a standard sensitive population (SEN) in pre-emergence and post-emergence dose-response pot tests, respectively. Sequencing of the whole HPPD gene from 12 each of sensitive and resistant plants did not detect any target-site mutations that could be associated with post-emergence resistance to mesotrione in NEB. Resistance was not due to HPPD gene duplication or over-expression before or after herbicide treatment, as revealed by qPCR. Additionally, no difference in mesotrione uptake was detected between NEB and SEN. In contrast, higher levels of mesotrione metabolism via 4-hydroxylation of the dione ring were observed in NEB compared to the sensitive population. Overall, the NEB population was characterised by lower levels of parent mesotrione exported to other parts of the plant, either as a consequence of metabolism in the treated leaves and/or impaired translocation of the herbicide. This study demonstrates another case of non-target-site based resistance to an important class of herbicides in an A. tuberculatus population. The knowledge generated here will help design strategies for managing multiple herbicide resistance in this problematic weed species.

Synthesis and evaluation of hydroxyazolopyrimidines as herbicides; the generation of amitrole in planta.

BACKGROUND: Exploiting novel herbicidal modes of action is an important method to overcome the challenges faced by increasing resistance and regulatory pressure on existing commercial herbicides. Recent reports of inhibitors of enzymes in the non-mevalonate pathway of isoprenoid biosynthesis led to the design of a novel class of azolopyrimidines which were assessed for their herbicidal activity. Studies were also undertaken to determine the mode of action responsible for the observed herbicidal activity. RESULTS: In total, 30 novel azolopyrimidines were synthesised and their structures were unambiguously determined by 1 H NMR, mass spectroscopy and X-ray crystallographic analysis. The herbicidal activity of this new chemical class was assessed against six common weed species, with compounds from this series displaying bleaching symptomology in post-emergence tests. A structure-activity relationship for the novel compounds was determined, which showed that only those belonging to the hydroxytriazolopyrimidine subclass displayed significant herbicidal activity. Observed similarities between the bleaching symptomology displayed by these herbicides and amitrole suggested that hydroxytriazolopyrimidines could be acting as elaborate propesticides of amitrole, and this was subsequently demonstrated in plant metabolism studies using Amaranthus retroflexus. It was shown that selected hydroxytriazolopyrimidines that displayed promising herbicidal activity generated amitrole, with peak concentrations of amitrole generally being observed 1 day after application. Additionally, the herbicidal activity of selected compounds was profiled against tobacco plants engineered to overexpress 4-diphosphocytidyl-2C-methyl-d-erythritol synthase (IspD) or lycopene ß-cyclase, and the results suggested that, where significant herbicidal activity was observed, inhibition of IspD was not responsible for the activity. Tobacco plants overexpressing lycopene ß-cyclase showed tolerance to amitrole and the two most herbicidally active triazolopyrimidines. CONCLUSIONS: Inhibition of IspD leading to herbicidal activity has been ruled out as the mode of action for the hydroxytriazolopyrimidine class of herbicides. Additionally, tobacco plants overexpressing lycopene ß-cyclase showed tolerance to amitrole, which indicates that this is the main herbicidal mode of action for amitrole. Results from the metabolic fate study of selected hydroxytriazolopyrimidines suggested that the herbicidal activity displayed by these compounds is due to amitrole production, which was confirmed when tobacco plants overexpressing lycopene ß-cyclase also showed tolerance towards two triazolopyrimidines from this study. © 2016 Society of Chemical Industry.