Amazonian soil fungi are efficient degraders of glyphosate herbicide; novel isolates of Penicillium, Aspergillus, and Trichoderma / Os fungos do solo amazônico são degradadores eficientes do herbicida glifosato; novos isolados de Penicillium, Aspergillus e Trichoderma

Abstract Pesticide residues that contaminate the environment circulate within the hydrological cycle can accumulate within the food chain and cause problems to both environmental and human health. Microbes, however, are well known for their metabolic versatility and the ability to degrade chemically stable substances, including recalcitrant xenobiotics. The current study focused on bio-prospecting within Amazonian rainforest soils to find novel strains fungi capable of efficiently degrading the agriculturally and environmentally ubiquitous herbicide, glyphosate. Of 50 fungal strains isolated (using culture media supplemented with glyphosate as the sole carbon-substrate), the majority were Penicillium strains (60%) and the others were Aspergillus and Trichoderma strains (26 and 8%, respectively). All 50 fungal isolates could use glyphosate as a phosphorous source. Eight of these isolates grew better on glyphosate-supplemented media than on regular Czapek Dox medium. LC-MS revealed that glyphosate degradation by Penicillium 4A21 resulted in sarcosine and aminomethylphosphonic acid.

Resumo Resíduos de agrotóxicos que contaminam o meio ambiente circulam no ciclo hidrológico, podendo se acumular na cadeia alimentar e causar problemas tanto à saúde ambiental quanto humana. Por sua vez, microrganismos são bem conhecidos por sua versatilidade metabólica e capacidade de degradar substâncias quimicamente estáveis, incluindo xenobióticos recalcitrantes. O estudo atual se concentrou na bioprospecção nos solos da floresta amazônica para encontrar novas linhagens de fungos capazes de degradar com eficiência o herbicida onipresente na agricultura e no meio ambiente, o glifosato. Entre os 50 fungos isolados (usando meio de cultura suplementado com glifosato como única fonte de carbono), a maioria eram isolados do gênero Penicillium (60%) e os outros eram isolados de Aspergillus e Trichoderma (26 e 8%, respectivamente). Todos os 50 isolados de fungos foram capazes de usar glifosato como fonte de fósforo. Oito desses isolados cresceram melhor em meio suplementado com glifosato do que em meio Czapek Dox regular. LC-MS revelou que a degradação do glifosato por Penicillium 4A21 resultou nos metabólitos sarcosina e ácido aminometilfosfônico.

Exposure of Artemia salinato glyphosate and bioremediation by isotherapy

Artemia salina, an ecotoxicity bioindicator, isa microcrustacean belonging to the order Anostraca. Glyphosate is an herbicide widely used to control weeds. However, its intoxication can cause serious damage to human health and the balance of the environment, given its effects as an endocrine disruptor.Objective:verify the possible protection of the artemia exposed to glyphosate by the addition of its isotherapic into water, by means of the evaluation of the behavioral and morphological features of nauplii and of the physical properties of remedies andtreated water, to elucidate the involved mechanisms. Methods:Artemia salinacysts were kept in culture bottles containing artificial seawatercontaining glyphosate at LC 10 (lethal concentration 10%), to promote hatching within 48 hours. The isotherapic preparations were inserted in each bottle in a 10% of the total water volume. Part of the nauplii was distributed in transparent tubes, being 10 nauplii per tube and 6 tubes per group, for behavioral analysis, and part were collected for a detailed morphological analysis, under an optical microscope. About 80 to 270 nauplii were analyzed per group. The reserved water was divided into aliquots for physicochemical analysis, that is, evaluation of the water dipole behavior by Cartwright ́s method.Results:Gly 6cH presented selective effects on nauplii hatching (p=0.02) and on defected/healthy ratio (p=0.001), representing some protective action. This result was dependent of the salinity of water and presented correspondence with the effects on solvatochromic dyes, indicating that charges and ions can be critical factors involved in the mechanism of action. We concluded that the use of isotherapics could be a plausible tool to reduce the environmental impact of the indiscriminate use of glyphosate, since these results can be reproduced in further studies.

Clinical characteristics of 83 patients with acute glyphosate herbicide poisoning / 中华急诊医学杂志

Objective:To analyze the clinical characteristics of patients with acute glyphosate herbicide poisoning and the differences in the severity of poisoning.Methods:A retrospective analysis was performed on patients with acute glyphosate herbicide poisoning admitted to the First Affiliated Hospital of Zhengzhou University from January 2014 to December 2020. The general information, exposure time, poisoning dose, poisoning cause, poisoning route, clinical manifestations, laboratory examination results during hospitalization, treatment measures, hospital stays and prognosis of the patients were collected. The patients were graded according to the poisoning severity scoring standard of Chinese Expert Consensus on Diagnosis and Treatment of Acute Poisoning in 2016. The highest severity score during hospitalization was used as the final grade. According to the final grade, asymptomatic and mild patients were included in the mild group, and moderate, severe and death patients were included in the severe group. The independent sample T test or Mann-Whitney U test was used for measurement data, and χ2 test or Fisher's exact test was used for counting data. The differences of general data and clinical data between the two groups were compared. Results:According to the inclusion and exclusion criteria, 83 patients with acute glyphosate herbicide poisoning were selected as the study subjects. All patients survived, mainly mild poisoning (56.6%), with a male to female ratio of 33∶50, and an average age of 39 years. The number of poisoning cases increased yearly (the highest in 2019), and most cases occurred in spring and summer. The main cause of poisoning was suicide (71.1%), direct oral administration (83.1%) was the primary route of poisoning, and the dominating clinical manifestations were digestive symptoms (71.1%). Laboratory tests showed increased white blood cell count (WBC), neutrophil percentage (NEUT %) and D-dimer, and decreased hemoglobin and potassium. Compared with the mild group, patients in the severe group were older [(51±17) years vs. (35±19) years], had a higher proportion of suicide and direct oral administration, a longer hospital stay [8.0 (4.8, 12.0) d vs. 3.0 (2.0, 5.5) d], a higher dose of poisoning [200.0 (50.0, 200.0) mL vs. 30.0 (11.3, 57.5) mL], and higher NEUT % within 24 h of admission [(83.4±10.4) vs. (73.2±12.8)]. The increase of WBC, NEUT %, aspartate aminotransferase, prothrombin time, D-dimer and the decrease of serum potassium were more common in the severe group than the mild group, with statistical significance (all P<0.05). Conclusions:The number of patients with acute glyphosate herbicide poisoning is increasing yearly. Generally, the condition is mild and the prognosis is satisfying. The severity is more serious in the middle-aged and elderly patients andthose with direct oral administration, high toxic dose, and high NEUT % within 24 h of admission. Severe poisoning is more likely to cause changes in laboratory indicators.

Determination of glyphosate in breast milk of lactating women in a rural area from Paraná state, Brazil

The aim of this study was to verify the presence of glyphosate in breast milk and to characterize maternal environmental exposure. Sixty-seven milk samples were collected from lactating women in the city of Francisco Beltrão, Paraná, living in urban (n=26) and rural (n=41) areas, at the peak of glyphosate application in corn and soy crops in the region (April and May 2018). To characterize the study population, socio-epidemiological data of the women were collected. To determine glyphosate levels, a commercial enzyme immunosorbent assay kit was used. Glyphosate was detected in all breast milk samples analyzed with a mean value of 1.45 µg/L. Despite some descriptive differences, there were no statistically significant differences (P<0.05) between the categories of the variables tested. Also, glyphosate was detected in drinking water samples from the urban area and in artesian well water from the rural area of the region where the studied population lived. The estimation of the total amount of glyphosate ingested by breastfeeding babies in a period of 6 months was significant. These results suggest that the studied lactating population was contaminated with glyphosate, possibly through continued environmental exposure.

Exposure to glyphosate-based herbicide during early stages of development increases insulin sensitivity and causes liver inflammation in adult mice offspring

ABSTRACT Objective: To investigate the effect of pre and postnatal exposure to a glyphosate-based herbicide on glucose metabolism and liver histology in adult F1 mice offspring. Methods: Female mice (C57Bl/6) received 0.5% of glyphosate (Roundup Original DI®) in drinking water or purified water (Glyphosate Group and Control Group respectively) during pregnancy and lactation. Offspring (F1) were submitted to glucose and insulin tolerance tests and euthanized on postnatal day 150. Body and plasma parameters, and liver histology were analyzed. Results: Exposure to glyphosate reduced maternal body weight gain during pregnancy and lactation, with no impacts on litter size. Pre and postnatal exposure to glyphosate did not affect body parameters but increased glucose tolerance on postnatal day 60. In spite of glucose tolerance normalization by postnatal day 143, this effect was associated with higher insulin sensitivity relative to mice in the Control-F1 Group. Mice in the Glyphosate-F1 Group had mild and moderate lobular inflammation in the liver. Conclusion: Maternal exposure to glyphosate affected insulin sensitivity and caused hepatic inflammation in adult F1 mice offspring.

Ultrafiltração na remoção de glifosato e ácido aminometilfosfônico em águas de abastecimento / Ultrafiltration for removal of glyphosate and aminomethylphosphonic acid in drinking water

RESUMO O glifosato é um herbicida não seletivo utilizado no controle de ervas daninhas em ambientes agrícolas e urbanos. É encontrado em mananciais associado ao seu principal metabólito, o ácido aminometilfosfônico, sendo necessária a remoção destes em estações de tratamento de água. O objetivo do trabalho foi avaliar a ultrafiltração direta na rejeição do glifosato e do ácido aminometilfosfônico em água de manancial superficial fortificada nas concentrações de 630, 800 e 1.250 μg L-1 e variação de pH entre 4 e 10. Para o glifosato, os aumentos do pH e da concentração e a presença de matéria orgânica natural contribuíram para a taxa de rejeição. A massa de glifosato adsorvida foi de 1,2 μg cm-2, sendo a adsorção na superfície e nos poros da membrana o principal mecanismo de rejeição. A ultrafiltração direta não foi efetiva para rejeição do ácido aminometilfosfônico. Para concentração do glifosato de 630 μg L-1 e pH entre 6,1 e 7,2, a ultrafiltração direta produziu permeado com concentração inferior a 500 μg L-1, atendendo ao padrão de potabilidade brasileiro, Portaria Gabinete do Ministro/Ministério da Saúde nº 888, publicada em 7 de maio de 2021.

ABSTRACT Glyphosate is a non-selective herbicide used to control weeds in agricultural and urban environments. It is found in water sources associated with its main metabolite, the aminomethylphosphonic acid, and it must be removed in the public water treatment. The aim of this work was to evaluate the direct ultrafiltration in the removal of glyphosate and aminomethylphosphonic acid in fortified surface water at concentrations of 630, 800, and 1,250 μg L-1 and pH variation between 4 and 10. For glyphosate, the increase in pH, concentration, and the presence of natural organic matter contributed to the rejection rate. The herbicide mass adsorbed was 1.2 μg cm-2, being adsorption onto the membrane surface/pores the main mechanism of herbicides retention. The UF process was not effective for removal of aminomethylphosphonic acid. For glyphosate concentration of 630 μg L-1, pH of 6.1-7.2, the UF was effective to produce the permeated in concentration of less than 500 μg L-1, as recommended by the Brazilian Drinking Water Legislation of the Ministry of Health, Portaria Gabinete do Ministro/Ministério da Saúde nº 888, publicada em 7 de maio de 2021.

The Effect of Glyphosate on Human Sperm: In Vitro Approximation / Efecto del glifosato sobre los espermatozoides humanos: una aproximación in vitro

Introduction Glyphosate is an herbicide used to eradicate illicit crops; however, its use is controversial due to different health problems associated with it. The present study aims to evaluate the effects of glyphosate on human sperm in vitro. Methods Twenty-two semen samples from healthy normozoospermic men were included; 11 semen samples were incubated with Panzer (INVESA S.A., Antiquia, Colombia) and 11 with Roundup (Monsanto Company, MO, USA). The changes in motility and viability were observed. Functional seminal parameters were evaluated as well. Results The samples exposed to glyphosate showed less motility and viability; a decrease in the potential of the mitochondrial membrane was observed, and an increase in the lipoperoxidation of the membrane was evidenced. Conclusion Based on the present results, we concluded that glyphosate has cytotoxic potential for exposed people and may affect their fertility.

Introducción El glifosato es un herbicida utilizado ampliamente para la erradicación de cultivos ilícitos; sin embargo, su uso es polémico debido a diferentes problemas de salud asociados con él. El objetivo del presente estudio fue evaluar los efectos del glifosato sobre los espermatozoides humanos in vitro. Métodos Se incluyeron 22 muestras de semen de hombres sanos normozoospérmicos, de las cuales 11 se incubaron con Panzer y 11 con Roundup, y se evaluaron los cambios en la movilidad y la viabilidad espermática, además de valorar los parámetros seminales funcionales. Resultados Las muestras expuestas al glifosato presentaron una menor movilidad y viabilidad, una disminución en el potencial de la membrana mitocondrial, y un aumento en la lipoperoxidación de la membrana. Conclusiones El glifosato es potencialmente citotóxico para las personas que estén expuestas, y puede afectar su fertilidad.

Destino ambiental y efectos ecológicos de los tres herbicidas más utilizados en Colombia / Environmental fate and ecological effects of the three most commonly used herbicides in Colombia / Destino ambiental e efeitos ecológicos dos três herbicidas mais comumente usados na Colômbia

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Abstract Knowledge of the environmental and transport fate of pesticides is integral to characterizing environmental exposures. Agencies such as the Environmental Protection Agency (EPA) 27 integrate the characterization of an exposure with ecological effects to determine the risks of pesticide use. The three most widely used herbicides in Colombia are glyphosate, propanil, and glufosinate ammonium. Although glyphosate had, since the early 1970's, been considered an environmentally friendly herbicide, the weight of the evidence of studies beginning in the 1990s show that inadvertent applications in wetlands can be fatal to the larvae of amphibians. Although glyphosate formulations are typically considered moderately toxic (LC50 = 1-10 ppm) for most aquatic organisms, for many amphibian species they are highly toxic (LC50 ˂ 1 ppm). This concern is compounded by the fact that Colombia has an incredibly high diversity of amphibians (746 species). In addition, chronic exposures of some amphibian species to sublethal concentrations, which are more likely found in the environment, have been shown to cause decreased survival, intersex gonads, tail damage, prolongation of time to metamorphosis, and liver lipidosis. In Colombia, aerial applications of Glyphos-Cosmo-Flux have been used since 1997 to fumigate illicit coca crops. As of 2019, the International Union for Conservation of Nature (IUCN) Red List had 285 species of amphibians in Colombia considered threatened. We found that 132 of the endangered (EN) and critically endangered (CR) amphibian species were present in 11 of the 32 Colombian departments with illegal coca plantations. Therefore, studies are needed to evaluate the impact of the coca eradication program on the amphibian population of the country. Propanil is the main herbicide used in rice fields. Despite its rapid environmental degradation ((1 day in aerobic conditions), studies on the impact of propanil have found that at projected application rates (4.5 kg/ha), within 48 hours after application, it reduces dissolved oxygen to levels critically important for survival of fish. At very low concentrations of 50 ppb or less of propanil, this effect correlates with lower phytoplankton production rather than higher community respiration. Despite being mild to moderately toxic at acute exposures, chronic exposure data also show that concentrations greater than 9 ppb affect the growth, reproduction, and development of fish and invertebrates. With respect to glufosinate ammonium, although it is highly likely to contaminate surface (runoff) and groundwater due to its physicochemical properties, data suggest that it is practically non-toxic to aquatic and terrestrial species. The acute hazard ratios were well below the level of concern, and therefore the EPA determined that the acute risk to non-target terrestrial and aquatic organisms is very low.

Resumo O conhecimento do destino ambiental e de transporte dos pesticidas é parte integrante da caracterização das exposições ambientais. Agências como a Agência de Proteção Ambiental (EPA) 27 integram a caracterização de uma exposição com efeitos ecológicos para determinar os riscos do uso de pesticidas. Os três herbicidas mais usados na Colômbia são glifosato, propanil e glufosinato de amônio. Embora o glifosato tenha sido considerado um herbicida ambientalmente correto desde o início da década de 1970, foi evidenciado por estudos iniciados na década de 1990 que aplicações inadvertidas em pântanos podem ser fatais para larvas de anfíbios. Embora as formulações de glifosato sejam tipicamente consideradas moderadamente tóxicas (CL50 = 1-10 ppm) para a maioria dos organismos aquáticos, para muitas espécies de anfíbios elas são altamente tóxicas (CL50 ˂ 1 ppm). Essa preocupação é agravada pelo fato de que a Colômbia tem uma diversidade incrivelmente alta de anfíbios (791 espécies). Além disso, foi demonstrado que exposições crônicas de algumas espécies de anfíbios a concentrações subletais, que são mais prováveis no ambiente, causam diminuição da sobrevivência, gônadas intersex, dano na cauda, prolongamento do tempo até a metamorfose e lipidose hepática. Na Colômbia, aplicações aéreas de Glyphos-Cosmo-Flux têm sido usadas desde 1997 para fumigar plantações ilícitas de coca. Em 2019, a Lista Vermelha da União Internacional para a Conservação da Natureza (IUCN) tinha 285 espécies de anfíbios na Colômbia consideradas ameaçadas. Descobrimos que 132 das espécies de anfíbios em perigo (EN) e criticamente em perigo (CR) estavam presentes em 11 dos 32 departamentos colombianos onde há mais plantações ilegais de coca. Portanto, são necessários estudos para avaliar o impacto do programa de erradicação da coca na população de anfíbios do país. O propanil é o principal herbicida usado nos campos de arroz. Apesar de sua rápida degradação ambiental ((1 dia em condições aeróbicas), estudos sobre o impacto do propanil descobriram que nas taxas de aplicação projetadas (4,5 kg / ha), ele reduz o oxigênio dissolvido a níveis criticamente importantes para a sobrevivência dos peixes em 48 horas. após a aplicação. Este efeito se correlaciona com a produção de fitoplâncton mais baixa, em vez de maior respiração da comunidade em concentrações muito baixas de 50 ppb ou menos de propanil. Apesar de ser leve a moderadamente tóxico em exposições agudas, os dados de exposição crônica também mostram que concentrações maiores que 9 ppb afetam o crescimento, a reprodução e o desenvolvimento de peixes e invertebrados. Com relação ao glufosinato de amônio, embora tenha grande probabilidade de contaminar a superfície (escoamento) e as águas subterrâneas devido às suas propriedades físico-químicas, os dados sugerem que é praticamente atóxico para as espécies aquáticas e terrestres. As razões de risco agudo estavam bem abaixo do nível de preocupação e, portanto, a EPA determinou que o risco agudo para organismos terrestres e aquáticos não visados é muito baixo.

Importance of sequential herbicide application in the control of Elephantopus mollis / A importância da aplicação sequencial no controle de Elephantopus mollis

RESUMO: Popularmente conhecida como pata de elefante ou sussuaiá, Elephantopus mollis, é uma espécie que recentemente surgiu como uma planta daninha em lavouras de cultivos de grãos. Objetivou-se com este estudo avaliar a sensibilidade diferencial de biótipos de E. mollis oriundos de áreas agrícolas e não agrícolas e determinar as melhores combinações de herbicidas aplicados em diferentes estádios de desenvolvimento para o controle desta espécie. Foram realizados três experimentos. No experimento de sensibilidade diferencial foram utilizados dez herbicidas em dois biótipos em duas fases de desenvolvimento. O experimento de dose resposta foi conduzido com oito doses de sete herbicidas. O experimento a campo foi realizado em uma área que apresenta problemas com a planta daninha para validar os resultados dos experimentos anteriores. O biótipo oriundo de área agrícola apresentou menor sensibilidade aos herbicidas quando comparado ao biótipo oriundo de área não agrícola. Os herbicidas de contato inicialmente apresentam controle superior, mas logo a planta apresenta rebrote em função de gemas presentes no colo da planta. A aplicação isolada de 2,4-D com doses entre 1005 e 1675 g e.a ha-1 possui controle de 25%. Quando o 2,4-D é seguido da aplicação de paraquat (400 g i.a ha-1) o controle passa a ser entre 51 e 68%. O melhor tratamento para o controle de plantas desenvolvidas é a mistura de 2,4-D + glyphosate (1340 + 1080 g e.a ha-1) com aplicação sequencial de paraquat.

ABSTRACT: Elephantopus mollis, popularly known as elephant paw or sussuaiá, is a species that has recently emerged as a weed affecting grain crops. This study aimed to evaluate the differential sensitivity of biotypes of E. mollis obtained from agricultural and nonagricultural areas and to determine the best combinations of herbicides applied at different stages of development for controlling this species. Three experiments were conducted. In the differential sensitivity experiment, 10 herbicides were used in 2 biotypes in 2 stages of development. The dose-response experiment was conducted using 8 doses of 7 herbicides. The field experiment was conducted in an area with weed issues to validate the results of previous experiments. The biotype from the agricultural area showed lower sensitivity to herbicides than the biotype from the nonagricultural area. The contact herbicides initially presented better control; however, the weed shortly showed re-growth due to the presence of buds in the plant crown. Isolated application of 2.4-D amine at doses between 1005 and 1675 g a.e. ha−1 demonstrated 25% control. When 2,4-D amine was followed by paraquat application (400 g a.i. ha−1), the control observed was between 51% and 68%. The best treatment for the control of completely developed weeds is 2,4-D amine + glyphosate mixture (1340 + 1080 g a.e. ha−1), followed by sequential paraquat application.

Study of muscle fibers of the extensor digitorium longus and soleus muscles of C57BL/6 females exposed to glyphosate during pregnancy and lactation / Estudo das fibras musculares dos músculos extensor longo dos dedos e sóleo de fêmeas C57BL/6 expostas ao glifosato durante a prenhez e a lactação

ABSTRACT Objective To evaluate the morphology and morphometry of the muscles extensor digitorium longus and soleus of C57BL/6 females, who were exposed to glyphosate during pregnancy and lactation. Methods Twelve female mice from the C57BL/6 lineage were used. After detection of pregnancy, they were divided into a Control Group, which received only water, and a Glyphosate Group, which received water with 0.5% glyphosate during pregnancy and lactation. Both groups received ad libitum standard diet. After weaning, the females were euthanized and weighed; naso-anal length was measured, and fats were collected and weighed. The muscles extensor digitorium longus and soleus were collected, and their length and weight were measured. Then, the muscles were fixed in Methacarn to perform the histological study of muscle fibers. Results Glyphosate Group presented lower weight gain during pregnancy and also lower final body weight and naso-anal length; however, the other body parameters evaluated did not present a significant difference in relation to the Control Group. Significant differences were also not observed in the analysis of muscle fibers and connective tissue. Conclusion Exposure to 0.5% glyphosate during pregnancy and lactation resulted in lower weight gain during pregnancy, final weight, and naso-anal length. Despite not directly altering the morphology of muscle tissue, these results may indicate enough exposure to interfere with animal metabolism.

RESUMO Objetivo Avaliar a morfologia e a morfometria dos músculos extensor longo dos dedos e sóleo de fêmeas C57BL/6 expostas ao glifosato durante a prenhez e lactação. Métodos Foram utilizados 12 camundongos fêmeas da linhagem C57BL/6. Após detecção da prenhez, foram separadas em Grupo Controle, que recebeu somente água, e Grupo Glifosato, que recebeu água com 0,5% de glifosato durante a prenhez e lactação. Ambos os grupos receberam dieta padrão ad libitum. Após o desmame, as fêmeas foram eutanasiadas e pesadas; o comprimento nasoanal foi mensurado, e as gorduras foram coletadas e pesadas. Os músculos extensor longo dos dedos e sóleo foram coletados, e seu comprimento e peso foram mensurados. Em seguida, os músculos foram fixados em Methacarn para a realização do estudo histológico das fibras musculares. Resultados O Grupo Glifosato apresentou menor ganho de peso durante a prenhez e também menor peso corporal final e comprimento nasoanal, entretanto os demais parâmetros corporais avaliados não apresentaram diferença significativa em relação ao Grupo Controle. Na análise das fibras musculares e do tecido conjuntivo, também não foram observadas diferenças significativas. Conclusão A exposição a 0,5% de glifosato durante a prenhez e lactação resultou em menor ganho de peso na gestação, peso final e comprimento nasoanal, o que pode indicar que, apesar de não alterar a morfologia do tecido muscular diretamente, a exposição foi suficiente para interferir no metabolismo dos animais.

Cloning and Characterization of 5-enopy ruvylshikimate-3-phosphate Synthase from Fragaria vesca

Abstract To discover and isolate a glyphosate-resistant gene from Fragaria vesca through gene mining. An open reading frame (ORF) of 1563 bp encoding EPSPSwas amplified from Fragaria vesca (FvEPSPS). FvEPSPS (Genebank: XP004306932.1) encodes a polypeptide of 520 amino acids and it has hightly homologous with EPSPS from other plants. qRT-PCR analysis showed that the FvEPSPS was expressed extensively in all tissues including leaves, roots and stems, with higher expression in leaves. Furthermore, transgenic Arabidopsis Thaliana exhibited 10 mM glyphosate to resistance. Therefore, this research offers a new glyphosate-resistant gene for development of transgenic crops.

Integrated management of tough lovegrass (Eragrostis plana Nees): associating chemical control tools and plant physiology / Manejo integrado de capim-annoni (Eragrostis plana Nees): associando ferramentas do controle químico e fisiologia da planta

ABSTRACT: The objective of the study was to evaluate the efficiency of herbicides use in the control of tough lovegrass according to the availability of solar radiation and the presence or absence of flooded. Two experiments were conducted in a completely randomized design, in a 2x2x4 and 2x2x5 factorial scheme, where factor A was equivalent to the environment (natural or reduced radiation); factor B to water condition (with a 2 cm flooded or without); and, factor C the herbicides: cyhalofop butyl (315 g ha-1), glyphosate (1080 g ha-1), cyhalofop butyl + glyphosate (315 g ha-1 + 1080 g ha-1) and control (without application) in the first experiment. In the second experiment, the herbicides used were cyhalofop butyl (315 g ha-1), glyphosate (1080 g ha-1), sethoxydim (184g ha-1), imazethapyr (106 g ha-1) and control (without application). Glyphosate efficiently controls plants of tough lovegrass with four tillers (>90%), being superior to cyhalofop, imazethapyr and sethoxydim; independently, of resource conditions. Reduction in the availability of solar radiation generates less shoot dry mass production from the weed, and improves the control only by imazethapyr and cyhalofop. In general, a flooded condition does not affect tough lovegrass control by herbicides.

RESUMO: O objetivo do trabalho foi avaliar a eficiência de uso de herbicidas no controle de capim-annoni em função da disponibilidade de radiação solar e da presença ou ausência de lâmina de água. Para isso, foram conduzidos dois experimentos em delineamento inteiramente casualizado, em esquema fatorial 2x2x4 e 2x2x5, em que o fator A equivaleu ao ambiente (radiação natural ou reduzida); fator B à condição hídrica (com lâmina d'água de 2 cm ou sem); e, o fator C aos herbicidas: cyhalofop butyl (315 g ha-1), glyphosate (1080 g ha-1), cyhalofop butyl + glyphosate (315 g ha-1+ 1080 g ha-1) e testemunha (sem aplicação) no primeiro experimento. No segundo experimento, os herbicidas utilizados foram: cyhalofop butyl (315 g ha-1), glyphosate (1080 g ha-1), sethoxydim (184 g ha-1), imazethapyr (106 g ha-1) e testemunha (sem aplicação). Glyphosate controla eficientemente plantas de capim-annoni com quatro afilhos (>90%), sendo superior a cyhalofop butyl, imazethapyr e sethoxydim, independente das condições. A redução da disponibilidade de radiação solar gera menor produção de matéria seca da parte aérea pelo capim-annoni, podendo inclusive, contribuir para maior eficiência de controle somente por imazethapyr e cyhalofop. De maneira geral, a condição de lâmina d'água não afeta o controle de capim-annoni pelos herbicidas.

Isolation of culturable mycota from Argentinean soils exposed or not-exposed to pesticides and determination of glyphosate tolerance of fungal species in media supplied with the herbicide / Aislamiento de la microbiota cultivable de suelos argentinos expuestos y no expuestos a plaguicidas y determinación de la tolerancia a glifosato de especies fúngicas en medios suplementados con el herbicida

Abstract The current agricultural system has led to the development of glyphosate (GP)-resistant weeds, causing an increase in GP doses and applications. Native mycota of pesticide-contaminated sites are the major source of pesticide-degrading microorganisms. The aims of the present study were to isolate the GP-tolerant culturable mycota in two soils with different pesticide exposure from Córdoba, Argentina, and to evaluate the growth parameters in native fungal isolates in the presence of GP and the effective dose that caused 50% growth reduction. The results showed that the genera Fusarium, Aspergillus, Mucor, Penicil-lium and Sterilia were the prevalent fungi isolated from soils both exposed and not-exposed to pesticides. The highest value (>100 mM) of effective concentration of herbicide that caused 50% growth inhibition (EC50), was found for Trichoderma isolates. Sterilia spp. had EC50 val-ues of 100mM, while Aspergillus spp. and Mucor had EC50 values between 50 and 100mM. The growth rate evaluation varied according to the isolates and GP concentrations. The data showed that all Aspergillus spp., Trichoderma spp., Mucor and three Sterilia spp. had the best growth performance in media supplied with GP after a variable acclimation period. This study provides valuable data for further studies that would allow to know the metabolic capacity of these fungal species that can be potential candidates for GP removal from contaminated environments.

Resumen El sistema agrícola actual condujo ai desarrollo de malezas resistentes a glifos-ato (GP), lo que deriva en el aumento de las dosis y el número de aplicaciones requeridas. La microbiota nativa de sitios contaminados con plaguicidas constituye la fuente principal de microorganismos capaces de degradar este herbicida. Los objetivos del presente estudio fueron aislar la microbiota cultivable tolerante a GP a partir de dos suelos agrícolas con diferente grado de exposición a plaguicidas, ambos de la provincia de Córdoba, Argentina, y evaluar en los hongos nativos aislados los parámetros de crecimiento en presencia de GP y la dosis efectiva que produce un 50% de reducción del crecimiento (EC50). Los géneros Fusarium, Aspergillus, Mucor, Penicillium y Sterilia fueron los de mayor prevalencia, tanto en suelos expuestos como no expuestos a plaguicidas. La mayor EC50 hallada (> 100 mM) correspondió a dos aislamientos de Trichoderma. En dos aislamientos de Sterilia, los valores de EC50 fueron de 100 mM, mientras que en los dos de Aspergillus y en uno de Mucor fueron de entre 50 y 100 mM. La velocidad de crecimiento varió según la concentración de GP empleada y el aislamiento considerado. Todos los aislamientos de Aspergillus, Trichoderma y Mucor, y tres aislamientos de Sterilia mostraron el mejor rendimiento en cuanto al crecimiento en los medios con GP después de un período de aclimatación variable. Este trabajo proporciona datos preliminares valiosos referidos a la capacidad metabólica de estos aislados y su potencial para remover GP en ambientes contaminados.

Effect of Roundup 41% (glyphosate) on blood serum biochemical parameters of freshwater fish, Rasbora daniconius

Aim: The present study was carried out to investigate the impact of Roundup 41% (glyphosate) on serum enzymes, like aminotransferases activity (ALT and AST), alkaline phosphates (ALP), total protein and glucose of fresh water fish, Rasbora daniconius.Methodology: LC50 of glyphosate for R. daniconius was calculated by static bioassays and recorded as 5.6 ppm at 96 hr. The fish were exposed to sub-lethal concentration (1/10th 96 hr LC50) of glyphosate for a period of 7, 14, 21 and 28 days. The serum ALT, AST, ALP, glucose and total protein were determined by standard methods. Results: Statistically important differences were observed in alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total protein and glucose levles. ALT, AST and ALP values were significantly higher (p<0.05) in treated group, compared with the control group. There was also a significant increase (p<0.05) in the level of serum glucose while the reduced amount of protein content as compared to control fish. Interpretation: Increased metabolic enzymes, protein metabolism and hyperglycemia in the serum may be a possible indicator of liver damage caused by Roundup 41% (glyphosate) exposure in fish, Rasbora daniconius.

Acute obstructive fi brinous laryngotracheobronchitis induced by severe glyphosate surfactant intoxication: A case report / 世界急诊医学杂志（英文）

@#Glyphosate is a nonselective herbicide commonly used in agriculture worldwide. Glyphosate targets the shikimate metabolic pathway, which is found in plants but not in animals. Therefore, it is considered to be of low toxicity to humans, supported by the finding of high concentrations of oral and dermal LD50 (>5,000 mg/kg body weight) of glyphosate in experimental animals.[1,2] Ingesting commercial formulations of glyphosate surfactant herbicide (GlySH), however, has been reported to be associated with various toxic effects, including gastrointestinal injury, laryngeal injury, impaired renal function, metabolic acidosis, arrhythmias, pulmonary toxicity, shock, and even death.[3] Recently, glyphosate surfactant has been used with increasing frequency in suicide attempts, and clinical toxicologists have encountered fatal cases of glyphosate surfactant poisoning.[4] Mortality from ingesting glyphosate surfactant varies from 0% to 29.3%, depending on patients’ characteristics such as age and intent of exposure. [5–7] Acute fi brinous laryngotracheobronchitis is often seen in infants and children, but it is rare in adults. Here we present a case with acute obstructive fibrinous laryngotracheobronchitis induced by severe glyphosatesurfactant intoxication.

Optimization and validation of a highly sensitive method for determining glyphosate in human urine by solid-phase extraction and liquid chromatography with tandem mass spectrometry: a methodological study

BACKGROUND@#Glyphosate and its salt formulations are nonselective herbicides that have been extensively used worldwide, both for residential and agricultural purposes. The possible carcinogenicity and teratogenicity of glyphosate remain to be elucidated. We developed a sensitive and high-throughput analytical method for urinary glyphosate using liquid chromatography-tandem mass spectrometry with the aim of contributing to glyphosate exposure assessment in epidemiological studies.@*METHODS@#After urine dilution (creatinine matching dilution to 0.05 g creatinine/L), glyphosate was extracted using two types of solid phase extraction columns (SCX and NH2) with automated sample preparation instruments. The eluate was dried and dissolved in the mobile phase, followed by liquid chromatography-tandem mass spectrometry analysis. The optimized method was applied to urine samples obtained from 54 Japanese adults and children.@*RESULTS@#The results from the validation study demonstrated good recoveries (91.0-99.6%), within- and between-run precisions (< 15%), low detection limits (0.1 μg/L), and lower limit of quantification (0.3 μg/L). The detection frequency and median concentration of the urinary glyphosate in Japanese subjects were 59% and 0.25 μg/L (0.34 μg/g creatinine).@*CONCLUSIONS@#Our reliable determination method was successful in measuring urinary glyphosate concentration. Moreover, this is the first biomonitoring report of urinary glyphosate levels in the Japanese general population.

Control of Conyza spp. with sequential application of glufosinate in soybean pre-sowing / Controle de Conyza spp. com aplicação sequencial de glufosinate em pré-semeadura da soja

ABSTRACT: Conyza spp. is among the main weeds reported worldwide. Due to its aggressiveness, such as high seed production and dispersion, and the growing reports of biotypes resistant to glyphosate, paraquat, and other herbicides, different control practices are required. Thus, this study aimed to evaluate the efficacy of herbicides with sequential application of glufosinate in soybean pre-sowing for control of Conyza spp. with indicative of resistant to paraquat. The study was carried out in the field, at Assis Chateaubriand and Palotina, state of Paraná, Brazil, in the 2018/19 season. The experiments were conducted in a randomized block design with four replications. The treatments consisted in application of glyphosate, 2,4-D, saflufenacil, glufosinate, saflufenacil/imazethapyr, diclosulam, paraquat/diuron, paraquat and imazethapyr/flumioxazin, at different combinations, in soybean pre-sowing. Control of Conyza spp., crop injury to soybean plants and variables related to agronomic performance (plant height and yield) were evaluated. All treatments were selective for soybean, which showed stronger crop injury in the presence of diclosulam herbicide, but this did not compromise soybean agronomic performance. In general, control levels were high for the treatments used. Except for paraquat treatments, in the area with the highest frequency of Conyza spp. with indicative of resistant to paraquat, and imazethapyr/flumioxazin treatment in both areas. These control results emphasized the importance of glufosinate in this management system and showed promising results for saflufenacil/imazethapyr.

RESUMO: A planta daninha Conyza spp. está entre as principais encontradas em todo o mundo. Devido a sua agressividade, como elevada produção e dispersão de sementes e os crescentes relatos de biótipos resistentes ao glyphosate, paraquat e outros herbicidas, faz-se necessário o uso de diferentes ferramentas de manejo. Assim, o objetivo deste estudo foi avaliar a eficácia de herbicidas com aplicação sequencial de glufosinate em pré-semeadura da soja, no controle de Conyza spp. com indicativo de resistência ao paraquat. O estudo foi realizado em campo nos municípios de Assis Chateaubriand e Palotina, estado do Paraná (PR), Brasil, na safra 2018/19. O delineamento experimental utilizado foi o de blocos casualizados, com quatro repetições. Os tratamentos foram compostos pela aplicação de glyphosate, 2,4-D, saflufenacil, glufosinate, saflufenacil/imazethapyr, diclosulam, paraquat/diuron, paraquat e imazethapyr/flumioxazin, em diferentes combinações em pré-semeadura da soja. Foram avaliados controle de Conyza spp., sintomas de injúria nas plantas de soja e variáveis relacionadas ao desempenho agronômico (altura de plantas e produtividade). Todos os tratamentos mostraram-se seletivos a soja. Visualizou-se maiores sintomas de injúria quando se tinha a presença do herbicida diclosulam, mas não resultou em danos ao desempenho agronômico da soja. De forma geral os níveis de controle foram altos para os tratamentos utilizados, em exceção para os tratamentos com paraquat na área que apresentava maior frequência de Conyza spp. com indicativo de resistência a este herbicida, e para o tratamento com imazethapyr/flumioxazin nas duas áreas. Estes resultados de controle enfatizam a importância do glufosinate neste sistema de manejo e demostram resultados promissores para saflufenacil/imazethapyr.

Toxicidade aguda (LC50) e efeitos comportamentais e morfológicos de formulado comercial com princípio ativo glifosato em girinos de Physalaemus cuvieri (Anura, Leptodactylidae) e Rhinella icterica (Anura, Bufonidae) / Acute Toxicity (CL50) and behavioral and morphological effects of a commercial formulation with glyphosate active ingredient in tadpoles of Physalaemus cuvieri (Anura, Leptodactylidae) and Rhinella icterica (Anura, Bufonidae)

RESUMO O declínio de anfíbios no mundo está cada vez maior e a perda de riqueza, muitas vezes, está associada a áreas agrícolas. O uso de agrotóxicos, em especial daqueles à base de glifosato, é cada vez maior e novas formulações estão sendo desenvolvidas. O presente trabalho avaliou a toxicidade aguda de um composto comercial com o princípio ativo glifosato e os efeitos comportamentais e morfológicos em girinos de Physalaemus cuvieri e Rhinella icterica. O primeiro mostrou-se mais sensível em relação ao formulado comercial do que o segundo e não houve diferenças significativas em relação às mudanças comportamentais e o aumento das concentrações do formulado entre as espécies, porém houve alterações morfológicas de peso e comprimento dos girinos de ambas espécies.

ABSTRACT The decline of amphibians in the world is increasing and the loss of wealth is often associated with agricultural areas. The use of pesticides, especially those based on glyphosate is increasing and new formulations are being developed. This study evaluated the acute toxicity of a commercial compound with glyphosate and behavioral and morphological effects on Physalaemus cuvieri and Rhinella icterica tadpoles; the former was more sensitive to the commercially formulated compound than R. icterica and there were no significant differences in relation to behavioral changes and increase compound concentrations among species, but there were morphological changes of weight and length of tadpoles of both species.

Preliminary Environmental Risk Assessment of Genetically Modified Oilseed Rape MON 88302 for Food and Feed Uses, Import and Processing under Regulation (EC) No 1829/2003 (Application EFSA/GMO/BE/2011/101)

The environmental risk assessment of the herbicide tolerant genetically modified oilseed rape MON 88302 (Reference EFSA/GMO/BE/2011/101) has been performed by the Panel on Genetically Modified Organisms (GMO) of the Norwegian Scientific Committee for Food Safety (VKM). VKM has been requested by the Norwegian Directorate for Nature Management and the Norwegian Food Safety Authority to issue a preliminary scientific opinion on the safety of the genetically modified oilseed rape MON 88302 (Unique identifier MON-88Ø2-9) for food and feed uses, import and processing, and submit relevant scientific comments or questions to EFSA on the application EFSA/GMOBE/2011/101. The environmental risk assessment of the MON 88302 is based on information provided by the applicant in the application EFSA/GMO/BE/2011/101, and scientific comments from EFSA and other member states made available on the EFSA website GMO Extranet. The risk assessment also considered peer-reviewed scientific literature as relevant. The VKM GMO Panel has evaluated MON 88302 with reference to its intended uses in the European Economic Area (EEA), and according to the principles described in the Norwegian Food Act, the Norwegian Gene Technology Act and regulations relating to impact assessment pursuant to the Gene Technology Act, Directive 2001/18/EC on the deliberate release into the environment of genetically modified organisms, and Regulation (EC) No 1829/2003 on genetically modified food and feed. The Norwegian Scientific Committee for Food Safety has also decided to take account of the appropriate principles described in the EFSA guidelines for the risk assessment of GM plants and derived food and feed (EFSA 2006, 2011a), the environmental risk assessment of GM plants (EFSA 2010), the selection of comparators for the risk assessment of GM plants (EFSA 2011b), and for the post-market environmental monitoring of GM plants (EFSA 2006, 2011c). The scientific risk assessment of oilseed rape MON 88302 include molecular characterisation of the inserted DNA and expression of target proteins, comparative assessment of agronomic and phenotypic characteristics, unintended effects on plant fitness, potential for horizontal and vertical gene transfer, and evaluations of the post-market environmental plan. In line with its mandate, VKM emphasized that assessments of sustainable development, societal utility and ethical considerations, according to the Norwegian Gene Technology Act and Regulations relating to impact assessment pursuant to the Gene Technology Act, shall not be carried out by the Panel on Genetically Modified Organisms. The GMO Panel has therefore not considered possible health and environmental effects of cultivation and processing of oilseed rape MON 88302 outside the EU/EEA area. The genetically modified oilseed rape MON 88302 was developed to provide tolerance to the herbical active substance glyphosate by the introduction of a gene coding for the enzyme 5enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium tumefaciens, strain CP4 (CP4 EPSPS). Glyphosate is a non-selective herbicide and is normally phytotoxic to a broad range of plants. Its mode of action occurs by binding to and inactivating the EPSPS protein, which is a key enzyme in the shikimate pathway that leads to the biosynthesis of the aromatic amino acids tyrosine, tryptophan and phenylalanine. The disruption of this pathway and the resulting inability to produce key amino acids prevents growth and ultimately leads to plant death. Molecular characterization: The VKM Panel on Genetically Modified Organisms find the conclusion that no major section of the T-DNA plasmid backbone is inserted in MON88302 oilseed rape justified. We also find it justified that there is only one major T-DNA insert in MON88302. Comparative assessment: Based on results from comparative analyses of data from field trials located at representative sites and environments in the USA, Canada and Chile, it is concluded that oilseed rape MON 88302 is agronomically and phenotypically equivalent to the conventional counterpart and commercial available reference varieties, with the exception of the herbicide tolerance conferred by the CP4 EPSPS protein. The field evaluations support a conclusion of no phenotypic changes indicative of increased plant weed/pest potential of MON 88302 compared to conventional oilseed rape. Furthermore, the results demonstrate that in-crop applications of glyphosate herbicide do not alter the phenotypic and agronomic characteristics of MON 88302 compared to conventional oilseed rape. Evaluations of environmental interactions between genetically modified oilseed rape MON 88302 and the biotic and abiotic environment, and studies of seed dormancy, seed germination, pollen morphology and viability indicates no unintended effects of the introduced trait on these characteristics in MON 88302 oilseed rape. Environmental risk: Considering the scope of the application EFSA/GMO/BE/2011/101, excluding cultivation purposes, the environmental risk assessment is limited to exposure through accidental spillage of viable seeds of MON 88302 into the environment during transportation, storage, handling, processing and use of derived products. Oilseed rape is mainly a self-pollinating species, but has entomophilous flowers capable of both self- and cross-pollinating. Normally the level of outcrossing is about 30 %, but outcrossing frequencies up to 55 % are reported. Several plant species related to oilseed rape that are either cultivated, occurs as weeds of cultivated and disturbed lands, or grow outside cultivation areas to which gene introgression from oilseed rape could be of concern. These are found both in the Brassica species complex and in related genera. A series of controlled crosses between oilseed rape and related taxa have been reported in the scientific literature. Because of a mismatch in the chromosome numbers most hybrids have a severely reduced fertility. Exceptions are hybrids obtained from crosses between oilseed rape and wild turnip (B. rapa ssp. campestris) and to a lesser extent, mustard greens (B.juncea), where spontaneously hybridising and transgene introgression under field conditions have been confirmed. Wild turnip is native to Norway and a common weed in arable lowlands. There is no evidence that the herbicide tolerant trait results in enhanced fitness, persistence or invasiveness of oilseed rape MON 88302, or hybridizing wild relatives, compared to conventional oilseed rape varieties, unless the plants are exposed to glyphosate-containing herbicides. However, accidental spillage and loss of viable seeds of MON 88302 during transport, storage, handling in the environment and processing into derived products is likely to take place over time, and the establishment of small populations of oilseed rape MON 88302 on locations where glyphosate is frequently applied to control weeds e.g. on railway tracks, cannot be excluded. Feral oilseed rape MON 88302 arising from spilled seed could theoretically pollinate conventional crop plants if the escaped populations are immediately adjacent to field crops, and shed seeds from cross-pollinated crop plants could emerge as GM volunteers in subsequent crops. However, both the occurrence of feral oilseed rape resulting from seed import spills and the introgression of genetic material from feral oilseed rape populations to wild populations are likely to be low in an import scenario. Apart from the glyphosate tolerance trait, the resulting progeny will not possess a higher fitness and will not be different from progeny arising from cross-fertilisation with conventional oilseed rape varieties. The VKM GMO Panel concludes that this route of gene flow would not introduce significant numbers of transgenic plants into agricultural areas or result in any environmental consequences in Norway. The environmental risk assessment will be completed and finalized by the VKM Panel on Genetically Modified Organisms when requested additional information from the applicant is available.

Influência do etanol como cosubstrato na biorremediação de água contaminada com formulação comercial à base de glifosato / Influence of ethanol as a co-substrate on bioremediation of water contaminated with glyphosate-based comercial formulation

RESUMO O presente trabalho objetivou avaliar a eficiência e a cinética de degradação de formulação comercial à base de glifosato em amostras de água, previamente contaminadas, via processo aeróbio, utilizando etanol como cosubstrato no processo. Utilizou-se um reator aeróbio com capacidade volumétrica de 1 L, operado em sistema de batelada sequencial com ciclos de 24 horas e biomassa em suspensão. Foram testadas quatro fases sob diferentes condições nutricionais: 1 sem a presença do etanol e outras 3 com relação etanol:glifosato de 7,8:1, 27,5:1 e 18,3:1. Quando o sistema operou sem a presença de etanol, a eficiência de remoção do glifosato foi de 18%, a qual aumentou para 78% ao se adicionar etanol na relação 7,8:1. No entanto, ao aumentar a concentração de etanol nas demais fases, não foram observadas melhoras na eficiência de remoção devido, provavelmente, à preferência dos organismos pelo uso do etanol como fonte de carbono em relação ao glifosato. Com relação aos ensaios cinéticos, os resultados foram ajustados ao modelo de degradação de pseudoprimeira ordem, considerando o residual de glifosato. A fase 3, com relação etanol:glifosato 27,5:1, foi a que apresentou maior valor da constante cinética aparente, k = 0,01746.min-1. Os resultados obtidos neste estudo indicam que a utilização de etanol como cosubstrato pode ser uma alternativa para a descontaminação de águas contaminadas com glifosato.

ABSTRACT This study aimed to evaluate the efficiency and degradation kinetics of commercial glyphosate formulation in previously contaminated water samples via aerobic process using ethanol as a co-substrate. An aerobic reactor with 1L volumetric capacity operated in sequential batch system with 24-hour cycles and suspended biomass was used. Four different nutritional conditions were tested: one with no ethanol addition; and three others with ethanol: glyphosate ratios of 7.8:1, 27.5:1 and 18.3:1. Operating with no ethanol addition, the efficiency of glyphosate removal was 18% and increased to 78% when ethanol was added to the system in a 7.8:1 ratio. However, when glyphosate concentration was further increased in the next phases, it did not result in higher removal efficiency, probably due to the preference of organisms for ethanol use as a carbon source over glyphosate. The results of the kinetic tests adjusted to first-order degradation models considering residual glyphosate. Phase 3 with 27.5:1 ethanol/glyphosate ratio showed the highest apparent kinetic constant value, k = 0.01746. The results of this study suggest that using ethanol as a co-substrate can be an alternative for decontamination of glyphosate contaminated waters.