High Concentrations Of Glyphosate Research Articles

Glyphosate Herbicide Induces Changes in the Growth Pattern and Somatic Indices of Crossbred Red Tilapia (O. niloticus × O. mossambicus)

Simple SummaryIn this study, a chronic, seven-week study of the effect of technical grade glyphosate on the toxicity parameters of crossbred red tilapia (O. niloticus × O. mossambicus) was carried out. The results show that the bodyweight index was the most sensitive toxicity parameter wherein a reduction in body weight was observed at 25 mg/L of glyphosate. Negative correlations between the glyphosate concentration and toxicity parameters such as specific growth rate (SGR), hepato-somatic index (HIS), and gonado-somatic index (GSI) were observed. The fish condition factor and feed conversion ratio were found to be unaffected at the highest glyphosate concentration tested (150 mg/L). The development of glyphosate-resistant genetically modified organisms (GMO) has increased the use of herbicide glyphosate by several magnitudes in recent years. It is now the most commonly used pesticide globally that affects aquatic habitats, especially fish. This study aims to add new knowledge on the effect of technical grade glyphosate on several toxicity parameters and to identify the most effective parameter in predicting technical grade glyphosate chronic toxicity (seven weeks) to fish, especially Malaysia’s heavily farmed red tilapia. The results show that a relatively high concentration of technical grade glyphosate is needed to induce significant changes in all tested parameters. However, the results also indicate that the bodyweight index is the most sensitive toxicity parameter in that a reduction in body weight was observed at 25 mg/L of glyphosate. Negative correlations between the glyphosate concentration and toxicity parameters such as specific growth rate (SGR), hepato-somatic index (HIS), and gonado-somatic index (GSI) were observed. The fish condition factor and feed conversion ratio were found not to be affected at the highest glyphosate concentration tested (150 mg/L). To conclude, crossbred red tilapia (O. niloticus × O. mossambicus) is one potential species for evaluating the toxic effects of technical grade glyphosate on fish.

Characterization of a Glyphosate-Tolerant Enzyme from Streptomyces svecius: A Distinct Class of 5-Enolpyruvylshikimate-3-phosphate Synthases.

Natural and modified versions of the 5-enolpyruvylshikimate-3-phosphate synthase (epsps) gene have been used to confer tolerance to the broad-spectrum herbicide glyphosate in a variety of commercial crops. The most widely utilized trait was obtained from the Agrobacterium tumefaciens strain CP4 and has been commercialized in several glyphosate-tolerant crops. The EPSPS gene products are enzymes that have been divided into three classes based on sequence similarity, sensitivity to glyphosate, and steady-state catalytic parameters. Herein, we describe the informatics-guided identification and biochemical and structural characterization of a novel EPSPS from Streptomyces sviceus (DGT-28 EPSPS). The data suggest DGT-28 EPSPS and other closely related homologues exemplify a distinct new class (Class IV) of EPSPS enzymes that display intrinsic tolerance to high concentrations of glyphosate (Ki ≥ 5000 μM). We further demonstrate that dgt-28 epsps, when transformed into stable plants, provides robust (≥4× field rates) vegetative/reproductive herbicide tolerance and has utility in weed-control systems comparable to that of commercialized events.

Effects of microplastics and glyphosate on growth rate, morphological plasticity, photosynthesis, and oxidative stress in the aquatic species Salvinia cucullata.

Glyphosate and microplastics are widely found in marine, terrestrial, and freshwater environments due to their globally widespread application. Further, they have proved to have specific ecotoxicity effects on aquatic plants. However, few studies have focused on the effects of small plastic particles and glyphosate, or especially, their combined effect on vascular plants in freshwater ecosystems. This study aimed to conduct a simulated greenhouse experiment to investigate the ecotoxicity of polystyrene microplastics and glyphosate on the floating plant Salvinia cucullata by exposure to fluorescent polystyrene microplastics (1μm; concentration, 3, 15, and 75mg/L), glyphosate (5, 25, and 50mg/L), and a mixture of the two (3+5, 15+25, and 75+50mg/L) for seven days. Glyphosate significantly reduced the relative growth rate, photosynthetic capacity, and root activity of S.cucullata. Polystyrene microplastics did not significantly influence photosynthesis or leaf morphological characteristics but they significantly reduced relative growth rate and root activity in S.cucullata, indicating that the effects of microplastics on aquatic plants are potentially associated with different organs exposed to pollution. Polystyrene microplastics and glyphosate activated the plant antioxidant defense systems by increasing antioxidative enzyme activities including, superoxide dismutase, ascorbate peroxidase, and catalase to cope with oxidative stress. Synergistic effects (only observed in percent leaf yellowing) were observed when S.cucullata was exposed to a high concentrations (≥15+25mg/L) of glyphosate and microplastics. Our results indicate that pervasive microplastics and herbicide contamination in freshwater may potentially affect the growth of aquatic plants.

Ecotoxicological Studies on the Effect of Roundup® (Glyphosate Formulation) on Marine Benthic Microalgae.

Glyphosate is a very effective herbicide and the main active ingredient in Roundup®—the most extensively used herbicide in the world. Since glyphosate is highly water soluble it reaches water bodies easily in surface water runoff. This prompted us to undertake an experiment to evaluate the effects of glyphosate in Roundup® on natural communities of marine microphytobenthos. Microphytobenthos communities were obtained from the environment, and after transporting them to the laboratory and acclimatizing them, they were tested under controlled conditions. Changes in microphytobenthos composition and structure and the deteriorating condition of the cells of community-forming organisms (assessed by analyzing changes in chloroplast shape) were used to assess the impact of Roundup® on endpoints. The tests indicated that microphytobenthic communities were relatively resistant to herbicide. The species richness of the communities probably enabled them to rebuild effectively. Sensitive species were replaced by those more tolerant of glyphosate. Only at the highest glyphosate concentration (8.5 g·dm−3) tested was a strong negative effect noted that limited community abundance and eliminated some of the organisms. The dominant diatoms in the communities were replaced by intensively developing cyanobacteria, which ultimately comprised nearly 60% of all the cells observed in the communities.

Changes in microtubule stability in zebrafish (Danio rerio) embryos after glyphosate exposure

Glyphosate, the most commonly used pesticide worldwide, blocks aromatic amino acid biosynthetic pathways and inhibits growth in plants. Although the specific mode of action of glyphosate in animals remains unclear, adverse effects during embryonic development have been reported, including epiboly delays, morphological alterations, and changes in central nervous system development and cardiogenesis. In this study, we suggest a possible toxicity mechanism for this herbicide related to changes in microtubule stability, which could alter the distribution and dynamics of cytoskeleton components. Using zebrafish embryos to evaluate in vivo effects of glyphosate exposure (5, 10, and 50 μg/ml), we found significant reductions in the levels of acetylated α-tubulin (50 μg/ml) and in the polymeric tubulin percentage in zebrafish embryos that had been exposed to 10 and 50 μg/ml glyphosate, without any changes in either the expression patterns of α-tubulin or the stability of actin filaments. These results indicate that high concentrations of glyphosate were associated with reduced levels of acetylated α-tubulin and altered microtubule stability, which may explain some of the neurotoxic and cardiotoxic effects that have been attributed to this herbicide.

Fatty Acids as Glyphosate Toxicity Indicators in Pseudomonas Fluorescens

Herbicides have harmful effects on bacteria, despite not being their primary targets. Herbicide active molecules cause unspecific oxidative stress, affecting soil microbiome function. Microorganisms possess enzymatic and non-enzymatic oxidative stress control systems that enable them to survive in contaminated environments. It is uncertain whether these systems require specific selective pressures to remain effective against contaminants and how the toxicity is signaled. The objective of this work was to analyze the antioxidant enzymes and lipid profiles of a Pseudomonas fluorescens strain, isolated from water used in washing of pesticide containers, as response systems to the herbicide glyphosate, not present at the isolation site. The bacteria showed low growth rates, cell viability and high hydrogen peroxide quantities at 40x and 50x glyphosate concentrations used in agriculture. These findings demonstrate the toxicity of this herbicide to Pseudomonas strain. However, growth rates in the presence of up to 10x were equivalent to the control group, probably because of superoxide dismutase and the catalase isoenzyme activities that managed to maintain peroxide levels under control in the early- and mid-log phases. Identification and quantification of lipids, by gas chromatography and mass spectrometry, showed that specific fatty acids were correlated with high concentrations of glyphosate and specific growth phases in P. fluorescens , through Principal Component Analysis, suggesting a role in modulating responses to glyphosate toxicity. Nevertheless, glyphosate was not present in the isolation site and, therefore, was not previously selective for P. fluorescens . This suggest that the response system is generalist, based on physiological plasticity, indicating substantial adaptative values in environments subjected to intense pesticide contamination. A matter that emerges is what would be the consequences of a system that is able to respond to toxic substances to which soil microbiomes have not had contact on for bacterial diversity and functionality in agricultural soils.

Fate of glyphosate in wheat during milling and bread production

AbstractBackground and objectivesGlyphosate is an active ingredient in widely used herbicides; its residues have been observed in grain and grain‐based foods. This work investigated the fate of glyphosate and its degradation product aminomethylphosphonic acid (AMPA) in milling fractions of wheat and during the bread making process. Samples of wheat were pearled to obtain successive fractions representing outer kernel layers. Straight grade flour and bread were prepared from the two samples with the highest glyphosate concentration (2.1 and 2.8 mg/kg).FindingsAll pearling and milling product fractions, dough, fermented dough, bread crust, and bread crumb were analyzed for glyphosate and AMPA using solvent extraction with derivatization and liquid chromatography–tandem mass spectrometry. Glyphosate was the only residue detected in samples. On average, the pearling experiment demonstrated that 50% of the total glyphosate mass resided in the outer 17% of the kernels. Similarly, 81% of the total glyphosate mass in the wheat was associated with the bran, shorts, and feeds milling fractions. No changes in glyphosate concentration were observed during the preparation of dough, fermented dough, and bread.ConclusionsMilling provides the best opportunity for reducing exposure to glyphosate, as the baking process did not affect glyphosate residues. Concentrations in bread made from straight grade flour will be approximately 4× lower than that made from whole grain flour.Significance and noveltyThe use of Canada Western Red Spring from commercial wheat shipments provide realistic glyphosate residue concentrations and also reflect the predominance of this type of wheat used in Canada and elsewhere for bread. Therefore, the outcomes from this study are particularly relevant for estimating consumers’ dietary exposure.

Proper Glyphosate Application at Post-anthesis Lowers Grain Moisture Content at Harvest and Reallocates Non-structural Carbohydrates in Maize.

Glyphosate (GP)-based herbicides have been widely applied to crops for weed control and pre-harvest desiccation. The objective of this research was to evaluate the effects of pre-harvest GP application on maize or how it physiologically alters this crop. Here, we applied four GP treatment (Control, GP150, GP200, and GP250) on maize lines of Z58 and PH6WC belonging to different maturity groups at grain-filling stages form DAP30 to DAP45. GP application significantly decreased the grain moisture content at harvest by 22–35% for Z58 and by 15–41% for PH6WC. However, the responses of grain weight to glyphosate vary with inbred lines and application time. A high concentration of glyphosate (GP250) reduced the grain weight of Z58 and low concentrations (GP150 and GP200) did not affect, while the grain weight of PH6WC significantly decreased under glyphosate treatment. In summary, our results revealed that timely and appropriate GP application lowers grain moisture content without causing seed yield and quality loss. GP application adversely affected photosynthesis by promoting maturation and leaf senescence. Meanwhile, it also enhanced non-structural carbohydrate (soluble sugars and starch) remobilization from the vegetative organs to the grains. Hence, GP treatment coordinates plant senescence and assimilate remobilization. RNA sequencing revealed that glyphosate regulated the transcript levels of sugar signaling-related genes and induced assimilate repartitioning in grains. This work indicates the practical significance of GP application for maize seed production and harvest, which highlights the contributions of source-sink communication to maize yield in response to external stress or pre-harvest desiccant application.

Differential impact of dose-range glyphosate on locomotor behavior, neuronal activity, glio-cerebrovascular structures, and transcript regulations in zebrafish larvae

The presence of glyphosate represents a debated ecotoxicological and health risk factor. Here, zebrafish larvae were exposed, from 1.5 to 120 h post-fertilization, to a broad concentration range (0.05–10.000 μg/L) of glyphosate to explore its impact on the brain. We evaluated morphology, tracked locomotor behavior and neurophysiological parameters, examined neuro-glio-vascular cell structures, and outlined transcriptomic outcomes by RNA sequencing.At the concentration range tested, glyphosate did not elicit gross morphological changes. Behavioral analysis revealed a significant decrease in locomotor activity following the exposure to 1000 μg/L glyphosate or higher. In parallel, midbrain electrophysiological recordings indicated abnormal, and variable, spike activity in zebrafish larvae exposed to 1000 μg/L glyphosate. Next, we asked whether the observed neurophysiological outcome could be secondary to brain structural modifications. We used transgenic zebrafish and in vivo 2-photon microscopy to examine, at the cellular level, the effects of the behavior-modifying concentration of 1000 μg/L, comparing to low 0.1 μg/L, and control. We ruled out the presence of cerebrovascular and neuronal malformations. However, microglia morphological modifications were visible at the two glyphosate concentrations, specifically the presence of amoeboid cells suggestive of activation. Lastly, RNAseq analysis showed the deregulation of transcript families implicated in neuronal physiology, synaptic transmission, and inflammation, as evaluated at the two selected glyphosate concentrations.In zebrafish larvae, behavioral and neurophysiological defects occur after the exposure to high glyphosate concentrations while cellular and transcript signatures can be detected in response to low dose. The prospective applicability to ecotoxicology and the possible extension to brain-health vulnerability are critically discussed.

Novel aroA of Glyphosate-Tolerant Bacterium Pseudomonas sp. Strain HA-09 Isolated from Roundup-Contaminated Garden Soils in Iran.

Background: Glyphosate is a non-selective systemic herbicide with a broad spectrum of weed control that inhibits a key enzyme, 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase, in the shikimate pathway.Objectives: Isolation and analysis of the epsps (aroA) gene responsible for glyphosate-tolerance in bacteria from Roundup- contaminated soils was the aim of this study.Materials and Methods: Sampling was done from the soil of the gardens which were heavily contaminated by Roundup herbicide and then bacterial screening was performed in the presence of high concentrations of glyphosate. The genus of bacterium was identified via molecular methods such as 16S rRNA sequencing. The aroA gene of this bacterium (aroAHA-09) was isolated using the primers designed-upon specific regions of aroA genes available in NCBI GenBank database. The PCR product was cloned, sequenced and subcloned into pET28a as an expression vector and transferred into E. coli strain BL21(DE3). The cells were inoculated in liquid M9 minimal medium containing IPTG and different concentrations of glyphosate.Results: The genus of bacterium was identified as Pseudomonas sp. strain HA-09. The isolated aroAHA-09 gene from this bacterium was approximately 2.2 kb in size. Bioassay of E. coli expressing this gene showed high tolerance to glyphosate (up to 300 mM).Conclusion: The aroAHA-09 gene could be considered as a novel and efficient candidate for development of glyphosate-tolerant crop plants.

DNA of crayfish spermatozoa as a target of waterborne pesticides – An ex vivo approach as a tool to short-term spermiotoxicity screening

The spermiotoxic properties of aquatic contaminants might be the cause of low fertilization rate and decreased prolificacy, affecting the success of the impacted populations. The genotoxic potential of pesticides in spermatozoa as an undesirable effect on non-target organisms, namely aquatic invertebrates with external fertilization, emerges as a key question in ecogenotoxicological research. Thus, this study aimed to clarify if DNA integrity of red swamp crayfish (Procambarus clarkii) spermatozoa is affected by waterborne pesticides at environmentally relevant concentrations. By adopting an ex vivo approach, six pesticides were addressed in a short-term assay: herbicides glyphosate (9 and 90 μg L−1) and penoxsulam (2.3 and 23 μg L−1); insecticides dimethoate (2.4 and 24 μg L−1) and imidacloprid (13.1 and 131 μg L−1); fungicides pyrimethanil (2.2 and 22 μg L−1) and imazalil (16 and 160 μg L−1). Genotoxicity was observed in higher concentrations of glyphosate, penoxsulam, dimethoate, pyrimethanil, and imazalil. Imidacloprid was the only pesticide that did not cause non-specific DNA damage, although displaying pro-oxidant properties. Overall, the present study demonstrated the suitability of the ex vivo approach on spermiotoxicity screening, highlighting the potential ecological impact of pesticides on non-target species, such as P. clarkii, compromising sperm DNA integrity and, subsequently, the population success.

Dermal, histological anomalies with variations in enzyme activities of the earthworms Lampito mauritii and Drawida willsi after short term exposure to organophosphate pesticides

Monocrotophos and glyphosate are two potent organophosphate pesticides used on agricultural farms in India to control insect pests and weeds, respectively. Consistent application of these chemicals poses a risk of residual soil contamination with possible adverse implications on non-target organisms, like earthworms. The present study evaluates the impacts of these pesticides on the skin, muscles and certain biochemical parameters such as protein content, lipid peroxidation (LPX) level, activities of lactate dehydrogenase (LDH), acetylcholinesterase (AChE) and catalase (CAT) of two tropical earthworms Drawida willsi and Lampito mauritii. Monocrotophos at 1.0, 2.0, 3.0 g/kg soil and glyphosate at 0.1, 0.15, 0.2 g/kg soil were used for the experiment. At high concentrations, both pesticides induced lesions and skin undulation in the earthworms. In L. mauritii, the postclitellar region indicated muscle disorganization with high concentrations of monocrotophos. The lowest protein level was recorded in D.willsi and L. mauritii with high concentrations of monocrotophos. L. mauritii exhibited maximum LPX at high concentrations of glyphosate. Both the earthworms indicated the least LDH activity with high pesticide concentration. Minimal AChE activity in L. mauritii was observed with a high concentration of glyphosate. A high concentration of monocrotophos inhibited CAT activity in L. mauritii. The variable response of the selected morpho-histological and biochemical parameters in the earthworms to different pesticide concentrations could be useful early warning biomarkers to evaluate soil residual toxicity.

The Effects of Glyphosate and Its Commercial Formulations to Marine Invertebrates: A Review

Glyphosate is the active ingredient of numerous commercial formulations of herbicides applied in different sectors, from agriculture to aquaculture. Due to its widespread use around the world, relatively high concentrations of glyphosate have been detected in soil and aquatic environments. The presence of glyphosate in aquatic ecosystems has aroused the attention of researchers because of its potential negative effects on living organisms, both animals and plants. In this context, this review intends to summarize results of studies aimed at evaluating the effects of glyphosate (both as active ingredient and component of commercial formulations) on marine invertebrates. Generally, data obtained in acute toxicity tests indicate that glyphosate and its commercial formulations are lethal at high concentrations (not environmentally realistic), whereas results of long-lasting experiments indicate that glyphosate can markedly affect biological responses of marine invertebrates. Consequently, more efforts should be addressed at evaluating chronic or sub-chronic effects of such substances to marine invertebrate species.

Effects of Glyphosate on Locomotion and Anxiety‐like Behavior in Rats

Glyphosate is the active ingredient in several commercial herbicides. The development of glyphosate‐resistant seeds has led to an increase in the use of glyphosate in agriculture. This has resulted in the exposure of non‐agricultural lands and bodies of water to glyphosate. Though initially considered safe for mammals, recent studies in rats have shown that administration of glyphosate leads to alterations in locomotion and anxiety‐like behavior. These studies generally used unrealistically high concentrations of glyphosate. Therefore, the effects of a dose of glyphosate considered safe by the Environmental Protection Agency through drinking water remain unclear. To determine if these levels affect locomotion and anxiety‐like behavior, we provided rats with access ad libitum to glyphosate‐contaminated drinking water or filtered water for four weeks. Glyphosate‐contaminated water was prepared for a target dose of 1.75mg/kg of glyphosate daily. We hypothesize that intake of glyphosate‐contaminated drinking water will decrease locomotion and increase anxiety‐like behavior. Locomotion was assessed in an open field. We observed that glyphosate did not decrease the overall distance travelled (control: 30.67m, glyphosate: 23.70m; p=0.058) nor did it affect average speed (control: 0.102m/s, glyphosate: 0.079; p=0.058). Anxiety‐like behaviors were also measured in the open field. Here, glyphosate did not increase the amount of entries to the center (control: 25.5, glyphosate: 27.5; p=0.726) nor time spent in the center (control: 74.22s, glyphosate: 87.9; p=0.572). In conclusion, this dose and exposure time of glyphosate did not affect locomotion or anxiety‐like behavior. Future directions include performing immunohistochemistry to assess neuronal activity in brain regions involved in locomotion and anxiety‐like behaviors.Support or Funding InformationYoung Investigator grant (NARSAD) of the Brain & Behavior Research Foundation, PRCTRC Pilot, NIGMS COBRE and RCMI8G12MD00760 to DS‐M; NSF PRCEN fellowships to CJR‐S, MC‐C, HH‐M, RR‐S and MR‐L, UPR Med Sci Campus Chancellor’s Office and School of Medicine Deanship.

The role of glyphosate in infertility: the mechanistic link

In light of the recent Roundup lawsuit, glyphosate has been widely accepted as a significant environmental toxin that may affect humans in various ways including cancer and infertility. Exposure to even low doses of glyphosate-based herbicides during pregnancy has been found to impair fertility, cause intrauterine growth restriciton and induce fetal malformations. In this study we sought to determine the underlying mechanism by which glyphosate negatively impacts oocyte quality, fertilization rates as well as embryo development. Experimental case-control study of mouse oocytes and pronuclear embryos, exposed in vitro to increasing glyphosate concentrations and followed through day 5 of development. We utilized multiple assays including reactive oxygen species (ROS) generation and zinc depletion to examine the possible underlying detrimental mechanisms. Metaphase II mouse oocytes (n=200) were retrieved from 8-10 week female mice and a subset (n=100) were fertilized using IVF. The oocytes as well as the fertilized mouse embryos were then exposed to increasing concentrations of glyphosate (0-200 μM) for 2h - 4h as per protocol. The oocytes were divided into four groups that were treated as follows: Group A: ROS detection assay, Group B: Zinquin ethyl ester assay, Group C: fixed, stained and scored based on the spindle structure (microtubule morphology -MT and chromosomal alignment - CH) as indicators of the oocyte's capacity to sustain exposure. All groups were compared to Group 4: untreated controls. Exposed embryos were incubated for up to 120 hours post fertilization and evaluated for full and hatching blast rate conversion. They were photographed and graded daily based on their appearance and development using published embryo grading protocols. A subset of the treated embryos (n = 10 for each concentration) were treated, in a similar fashion as the oocytes, in order to evaluate for ROS overproduction and zinc depletion. Confocal microscopy was used to assess the embryos. Statistical analysis was performed using t-test, ANOVA and chi-square. A p-value < 0.05 was considered statistically significant. Oocytes treated with increasing glyphosate concentrations > 50 μΜ were found to have poor scores for MT and CH and that difference was statistically significant as compared to controls (p< 0.05). Embryos followed to 96 hours post fertilization (early blastocyst) and 120 hours (full and hatching blastocyst) after glyphosate exposure (0-200 μM) were assessed and those exposed to glyphosate concentrations > 100 μM showed significantly increased arrest rates and poor morphology scores compared to controls. ROS overproduction as well as zinc depletion was evident in embryos treated with high glyphosate concentrations. These observations were statistically significant compared to untreated controls (p<0.05). This work suggests the possible underlying mechanisms by which glyphosate negatively affects reproductive health in the mouse model. Possible fertility implications in humans will require further research.

The glyphosate influence on cytogenetic and biochemical aspects of wheat (Triticum aestivum L) seedlings development

The influence of glyphosate on the germination and development of bread wheat (Triticum aestivum) was researched. The chronical influence during all the time of growth was estimated, together with the consequences of the contact with glyphosate of the already formed seedlings. A dose-dependent decrease in the growth rate of seedlings in the presence of glyphosate was found, and even in normally developing seedlings damage to the root tips was observed. In particular, in seedlings, the integrity of the root cap was disrupted, degenerative changes of the rhizodermis and root hairs were detected. It has been shown that a decrease in the root growth rate during chronic exposure is largely due to the suppression of mitoses. In addition, high glyphosate concentrations had a general toxic effect, stopping the development of seedlings yet at the stage of primary roots growth through enlarging the cells. It was found that the antioxidant system of seedlings prevented the development of oxidative stress at the concentration of herbicide 50 μM, in particular, due to the activation of peroxidase. Placing seedlings grown on distilled water in 100 μM glyphosate solution induced lipid peroxidation. Chronic effects of herbicide in this concentration inhibited lipid metabolism in general.

How glyphosate and its associated acidity affect early development in zebrafish (Danio rerio).

BackgroundGlyphosate is among the most extensively used pesticides worldwide. Following the ongoing highly controversial debate on this compound, its potential impact on non-target organisms is a fundamental scientific issue. In its pure compound form, glyphosate is known for its acidic properties.MethodsWe exposed zebrafish (Danio rerio) embryos to concentrations between 10 μM and 10 mM glyphosate in an unbuffered aqueous medium, as well as at pH 7, for 96 hours post fertilization (hpf). Furthermore, we investigated the effects of aqueous media in the range of pH 3 to 8, in comparison with 1 mM glyphosate treatment at the respective pH levels. Additionally, we exposed zebrafish to 7-deoxy-sedoheptulose (7dSh), another substance that interferes with the shikimate pathway by a mechanism analogous to that of glyphosate, at a concentration of one mM. The observed endpoints included mortality, the hatching rate, developmental delays at 24 hpf, the heart rate at 48 hpf and the malformation rate at 96 hpf. LC10/50, EC10 and, if reasonable, EC50 values were determined for unbuffered glyphosate.ResultsThe results revealed high mortalities in all treatments associated with low pH, including high concentrations of unbuffered glyphosate (>500 μM), low pH controls and glyphosate treatments with pH < 3.4. Sublethal endpoints like developmental delays and malformations occurred mainly at higher concentrations of unbuffered glyphosate. In contrast, effects on the hatching rate became particularly prominent in treatments at pH 7, showing that glyphosate significantly accelerates hatching compared with the control and 7dSh, even at the lowest tested concentration. Glyphosate also affected the heart rate, resulting in alterations both at pH 7 and, even more pronounced, in the unbuffered system. In higher concentrations, glyphosate tended to accelerate the heart rate in zebrafish embryos, again, when not masked by the decelerating influence of its low pH. At pH > 4, no mortality occurred, neither in the control nor in glyphosate treatments. At 1 mM, 7dSh did not induce any mortality, developmental delays or malformations; only slightly accelerated hatching and a decelerated heart rate were observed. Our results demonstrate that lethal impacts in zebrafish embryos can be attributed mainly to low pH, but we could also show a pH-independent effect of glyphosate on the development of zebrafish embryos on a sublethal level.

Physiological and molecular analysis of glyphosate resistance in non-rapid response Ambrosia trifida from Wisconsin.

We previously identified a glyphosate-resistant A. trifida phenotype from Wisconsin USA that showed a non-rapid response to glyphosate. The mechanism of glyphosate resistance in this phenotype has yet to be elucidated. We conducted experiments to investigate non-target-site resistance and target-site resistance mechanisms. The roles of glyphosate absorption, translocation, and metabolism in resistance of this phenotype have not been reported previously, nor have EPSPS protein abundance or mutations to the full-length sequence of EPSPS. Whole-plant dose-response results confirmed a 6.5-level of glyphosate resistance for the resistant (R) phenotype compared to a susceptible (S) phenotype. Absorption and translocation of 14 C-glyphosate were similar between R and S phenotypes over 72 h. Glyphosate and AMPA concentrations in leaf tissue did not differ between R and S phenotypes over 96 h. In vivo shikimate leaf disc assays confirmed that glyphosate EC50 values were 4.6- to 5.4-fold greater for the R than S phenotype. Shikimate accumulation was similar between phenotypes at high glyphosate concentrations (>1000 μM), suggesting that glyphosate entered chloroplasts and inhibited EPSPS. This finding was supported by results showing that EPSPS copy number and EPSPS protein abundance did not differ between R and S phenotypes, nor did EPSPS sequence at Gly101, Thr102, and Pro106 positions. Comparison of full-length EPSPS sequences found five nonsynonymous polymorphisms that differed between R and S phenotypes. However, their locations were distant from the glyphosate target site and, therefore, not likely to affect enzyme-glyphosate interaction. The results suggest that a novel mechanism confers glyphosate resistance in this A. trifida phenotype. © 2019 Society of Chemical Industry.

Potential of terracing to reduce glyphosate and AMPA surface runoff on Latosol

Glyphosate is the world’s most used herbicide and monitoring glyphosate in the environment is a relevant topic. The present study aims to develop a methodology to extract glyphosate from the soil and sediments, and assess the potential of the terracing system to mitigate contamination by glyphosate and AMPA in soil and water. Collections were performed on a weekly basis in two different periods of the agricultural calendar, totaling 24 Latosol soil samples, 12 sediment samples, and 10 water samples. The sampling was performed in two distinct areas: in the cultivation area where the lots with and without terrace were installed (soil and water of the reservoirs) and in the creek (sediment) present in the middle of the property. The analytes were extracted from the soil and sediment samples using alkaline extraction with KH2PO4 and NH4OH. The supernatant resulting from the extraction and the water samples were submitted to the derivatization (FMOC-Cl) and solid-phase extraction steps. The samples were then analyzed by high-performance liquid chromatography equipped with fluorescence detector (HPLC-FD). The soil samples showed AMPA content in all samples ranging from 0.50 to 1.11 μg g−1 of soil. Glyphosate could be quantified in 37.5% of the samples, and the concentrations ranged from 0.21 to 0.49 μg g−1. High concentrations of glyphosate were detected in the water samples (20.74 and 31.24 μg L−1) in the first rain events after application, decreasing significantly in the following rainfall events. The concentrations found were similar for both lots, but the volume of runoff water was higher in the lot without terrace, thus a greater mass of analyte was transported. None of the analytes under investigation could be quantified in the analysis of riverbed sediments. Glyphosate and AMPA, however, were detected in 50 and 75% of the sediment samples respectively. The results indicate the presence of analytes in the cultivation areas and prove the effectiveness of the terracing system in agricultural areas, limiting the dumping of the material originated from the surface runoff into water bodies and reducing the risk of contamination.

Antioxidative status, immunological responses, and heat shock protein expression in hepatopancreas of Chinese mitten crab, Eriocheir sinensis under the exposure of glyphosate

As a broad-spectrum herbicide, glyphosate was extensively utilised in China for several decades. The contradiction between glyphosate spraying and crab breeding in the rice-crab co-culture system has become more obvious. In this study, the antioxidative status and immunological responses of Chinese mitten crab, Eriocheir sinensis, under sublethal exposure of glyphosate were investigated by detecting the antioxidative and immune-related enzyme activity, acetylcholinesterase (AChE) activity and relative mRNA expression of heat shock proteins (HSPs) in hepatopancreas. The results showed that high concentrations of glyphosate (44 and 98 mg/L) could induce significant alteration of superoxide dismutase (SOD), peroxidase (POD), acid phosphatase (ACP), alkaline phosphatase (AKP), and phenoloxidase (PO) activities by first rising then falling during the exposure. However, AChE activity in all treatments including 4.4 mg/L was inhibited markedly after 6 h of exposure. In addition, the relative mRNA expression of HSP 60, HSP 70, and HSP 90 was significantly upregulated at both 48 h and 96 h. These results revealed that glyphosate has a prominent toxic effect on E. sinensis based on antioxidative and immunological response inhibition and AChE activity reduction even at the lowest concentration of 4.4 mg/L, and a protective response by upregulation of HSPs was carried out by the species to ease the environmental stress.