Effets des pesticides de la vigne sur le cycle biologique de l'escargot dans divers contextes d'exposition

An increasing number of chemicals are produced worldwide and a large part of them are plant protection products (PPPs). France is the first European consumer of PPPs and the third on the global level. The aim of our research was to investigate the fate and the effects of pesticides in the environment, particularly in the agrosystem, with the use of a bioindicator model, the snail Helix aspersa. We chose to characterize the exposure, the transfer and the effects on snails of 8 commercial herbicide and fungicide formulations (Roundup® and Bypass® = glyphosate; Basta® = glufosinate; Valiant® Flash = folpet + cymoxanil + fosetyl-Al; Corail® = tebuconazole; Cabrio® Top = pyraclostrobin + metiram-Zn; Thiovit® = sulphur; Bordeaux mixture = copper salts), commonly employed in vineyards. A field study in real conditions of pesticide application allowed the quantification of the spray deposits on soil in the parcel and in its surroundings showing the PPP drift deposits. At the same time, few effects were demonstrated on the survival and growth of juvenile snails exposed in microcosms to the different treatments, except for growth inhibition after Roundup® exposure, probably in relation to heavy spray deposits on the soil, accumulation and metabolisation of the active ingredient in the tissues. A soil extraction and analysis method involving two herbicides (glyphosate, glufosinate) was developed allowing the determination of exposure concentrations along the experiments and their persistence in a vineyard soil. The effects of Bypass® and Basta® were studied on the whole life cycle of snails in laboratory conditions. No significant effects were shown on survival and growth although an accumulation of glyphosate was revealed in snails (6 mg kg-1) exposed continuously to contaminated food (30 mg kg-1). However, we found that exposure of snails to glyphosate during their embryonic development tended to disrupt reproduction in the resulting adult snail (by decreasing fertility) due to a delayed effect of the herbicide. The potential targets affected at the level of neuro-endocrine control are discussed. All pesticides inhibited egg hatching success at lower concentrations than those applied in the field and the influence of soil characteristics on the toxicity of the pesticides was demonstrated. These effects were showcased by laboratory experiments that led to the development of embryotoxicity bioassays. This work has contributed to current understanding of the in- and on-soil fate and effects on snails (transfer and toxicity) of various pesticides applied to the vineyard. It allowed the development of tools (laboratory bioassays and field exposure device) that could help improving pesticide risk assessment proceedings in a terrestrial environment. The analysis methods designed for use in complex matrixes (soil and animal tissues) make an original contribution to environmental and human health risk assessment. Extensive analytic difficulties were encountered and it was not possible to finalize the analytical methods for all of the substances studied. Thus, the determination of pesticide residues in animal tissues still represents an essential challenge for current environmental concerns.De plus en plus de substances chimiques sont produites dans le monde et une grande partie d'entre elles entrent dans la composition des produits phytosanitaires. La France en est le premier consommateur européen et le troisième au niveau mondial. L'objectif de nos recherches est de comprendre le devenir et les effets de ces molécules dans l'environnement, notamment agricole, grâce à l'utilisation d'un invertébré bioindicateur, l'escargot Helix aspersa. Nous avons choisi de caractériser l'exposition, le transfert et les effets sur l'escargot de 8 formulations commerciales de fongicides et d'herbicides (Roundup® et Bypass® = glyphosate; Basta® = glufosinate; Valiant® Flash = folpel + cymoxanil + fosétyl-Al; Corail® = tébuconazole; Cabrio® Top = pyraclostrobine + métirame-Zn; Thiovit® = soufre; Bouillie bordelaise = cuivre), communément utilisées en viticulture. Une étude réalisée en conditions in situ a permis de quantifier les retombées au sol des pesticides appliqués sur la vigne, dans la parcelle et en périphérie mettant en évidence les dépôts suite à la dérive des substances. En parallèle, peu d'effets ont été démontrés sur la survie et la croissance d'escargots juvéniles placés en microcosmes et soumis aux différents traitements de pesticides, mis à part un effet inhibiteur du Roundup® sur la masse des organismes, probablement en relation avec d'importants dépôts au sol, l'accumulation et la métabolisation de la substance active dans les tissus. Une méthode d'extraction et d'analyse de deux herbicides (glyphosate et glufosinate) dans le sol a été développée afin de pouvoir mesurer les concentrations d'exposition pour diverses expériences et de caractériser leur persistance dans ce sol de vigne. Les effets du Bypass® et du Basta® ont été étudiés sur l'ensemble du cycle de vie de l'escargot en conditions contrôlées. Aucun effet significatif sur la survie et la croissance n'a été démontré bien qu'une accumulation de glyphosate ait été quantifiée chez des escargots (6 mg kg-1) nourris continuellement par de l'aliment contaminé (à 30 mg kg-1). Cependant, une exposition au glyphosate des escargots durant le développement embryonnaire semble perturber leur reproduction (diminution de la fertilité) du fait d'un effet différé de cet herbicide. Les cibles potentiellement affectées au niveau du contrôle neuroendocrine sont discutées. Tous les pesticides inhibent l'éclosion des œufs de l'escargot à des concentrations inférieures à celles appliquées au champ et l'influence des caractéristiques du sol sur leur toxicité a été démontrée. Ces effets ont été mis en évidence lors d'expériences réalisées en laboratoire dans le cadre d'un développement de bioessais d'embryotoxicité. Ces travaux de thèse ont apporté des connaissances sur le comportement, en surface et dans le sol, ainsi que dans l'escargot (transfert, toxicité), de plusieurs pesticides appliqués sur la vigne. Ils ont également permis de développer plusieurs outils (bioessais de laboratoire, dispositif d'exposition sur le terrain) visant à améliorer les procédures d'évaluation du risque des pesticides dans l'environnement terrestre. Les techniques d'analyse de ces composés mises au point pour des matrices complexes (sols, tissus animaux), constituent un apport original pour l'évaluation du risque environnemental et sanitaire. D'importantes difficultés analytiques ont été rencontrées et il n'a pas été possible de finaliser les méthodes de dosage pour la totalité des substances. Ainsi, la mesure des résidus de pesticides dans les tissus d'origine animale reste un challenge fort et qui plus est, au cœur des préoccupations environnementales actuelles

Towards the development of an embryotoxicity bioassay with terrestrial snails: Screening approach for cadmium and pesticides

International audienceCurrently no bioassays are available to assess the embryotoxicity of chemicals with terrestrial soil invertebrates. We therefore presented a new method for embryotoxicity testing with snail eggs: a relevant biological material that incubates in soil and that can be exposed to contaminants from leachates and soil solution. The effects of aqueous solutions of two herbicide formulations, Reglone® (active ingredient (a.i.), diquat) and Roundup® or its a.i., glyphosate, of a surfactant (Agral® 90, a.i., nonylphenol polyethoxylates) and of cadmium (Cd) were studied. Endpoints were the hatching success and observations of embryo abnormalities after exposure. Roundup® was found to be more toxic than its a.i. alone (EC50a.i. = 18 mg/l and about 1300 mg/l, respectively). Reglone® (EC50a.i. = 0.72 mg/l) and Agral® (EC50a.i. ? 50 mg/l) were also tested together, revealing that Reglone® accounted for more than 99% of the mixture's toxicity. An antagonistic interaction between the two substances was found. For Cd (EC50 = 3.9 mg/l), a significant transfer from exposure medium to eggs was emphasized, particularly affecting the albumen. Abnormalities of embryogenesis in non-hatched embryos depended on the substance and the concentration considered

Ecotoxicity evaluation of industrial discharge waters and metallic solutions using two organisms (Lactuca sativa and Daphnia magna

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Monitoring and Origin of Polycyclic Aromatic Hydrocarbons (PAHs) in Effluents from a Surface Treatment Industry

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Landsnail eggs bioassays: A new tool to assess embryotoxicity of contaminants in the solid, liquid or gaseous phase of soil

International audienceBioassays for ecotoxicity testing for the same non-target soil organism are not currently available for contaminants that can be present in the liquid, solid or gaseous phase of soil. Here, three bioassays with three modes (liquid, solid or gaseous phase, LPB, SPB and GPB, respectively) allowing the assessment of the embryotoxicity of chemicals or pluri-contaminated matrices on land snail eggs are presented. Eight pesticides commonly used in vineyards (based on 10 active ingredients: copper, sulfur, metiram-zinc+pyraclostrobin, cymoxanil+folpet+fosetyl-Aluminium, tebuconazole, glyphosate, glufosinate) were tested by LPB: all of them reduced egg hatching success at concentrations lower than those applied in the field. The SPB was developed with one artificial ISO substrate and seven natural soils: three non-contaminated agricultural soils (Agr1-3) and four metal-contaminated soils (Me1-4). The moisture content (from 40 to 60% of the water holding capacity) in the natural soils did not influence the hatching success. Hatching success did not vary in the three agricultural soils suggesting the relative insensitivity of eggs to some soil properties. Among the two pesticides tested in SPB, Corail® (tebuconazole based-fungicide) was more toxic than Bypass® (glyphosate based-herbicide) to snail embryos with EC50 values of 1 and 219 mg kg-1 respectively in a natural soil (Agr2). Both pesticides were less toxic when tested in the ISO substrate (EC50 of 7.8 and higher than 400 mg kg-1, respectively for Corail® and Bypass®), highlighting the influence of the organic matter content (lower in soil Agr2) on the bioavailability and thus the toxicity of the chemicals. LPB showed that soluble compounds of the most toxic soil in SPB (Me4) did not affect embryos. Similarly, GPB did not reveal toxic volatile compounds from this soil. These bioassays are complementary and efficient tools for soil risk assessment

A full life-cycle bioassay with Cantareus aspersus shows reproductive effects of a glyphosate-based herbicide suggesting potential endocrine disruption

International audienceA full life-cycle (240 days) bioassay using the terrestrial snail, Cantareus aspersus, allowing exposureduring embryogenesis and/or the growth and reproduction phases, was used to assess the effects ofBypass®, a glyphosate-based herbicide (GlyBH), on a range of endpoints, including parameters underendocrine control. As a positive control, a mixture (R-A) made of diquat (Reglone®) and nonylphenols (NP, Agral®), known for its endocrine disrupting effects in other organisms, was tested. At environmental concentrations, both pesticides (R-A mixture and GlyBH) enhanced growth but reduced reproduction. The R-A mixture acted mainly on the fecundity through a delay in egg-laying of approximately 20 days and a strongly reduced number of clutches. This latter dysfunction may be caused by a permanent eversion of the penis, suggesting a disrupting effect at the neuro-endocrine level, which prevented normal mating. GlyBH acted on fertility, possibly due to a decrease in the fertilization of eggs laid by adults exposed during their embryonic development. These results, associated with the absence of observed effects on gonad histology of GlyBH exposed snails, suggested that the underlying mechanisms are neuro-endocrine

Snails in grapevine: active bioindication and drift effects of pesticides

More and more chemicals are produced in the world and a big part entry in the composition of pesticides. The intensive use of pesticides has ended up in chronic contamination of the ecosystem compartments: air, water and soil and their effects on the environment aren't much known. The snail is an appropriate model to bring data about effects of these contaminants because its particular place at interface soil-air-plant. Moreover it can be itself a source of contamination for their predators included humans. The aim of this study is to expose snails in real conditions of applications of pesticides and to measure the effects of drift on mortality. Snails were encaged in microcosms placed in a parcel of grapevine and in kerb (from 1m to the parcel). An area without contamination, ranged in 1.2 km, served as control. Five treatments (2 herbicides and 3 fungicides) occurred from May to July. The day of each treatment, soil deposition of pesticides was measured from 0 to 20 m, under wind, of the parcel. The results indicate no significant mortality due to pesticides in the parcel and in kerb. For the soil deposition of fungicides, average ranged from 0.308 to 106.6 g/ha in the parcel following the poised active ingredient and the poised treatment. These differences can be explained by meteorological conditions, height of vegetation and physic-chemical properties of the active ingredient among other things. For the herbicides, average concentrations ranged from 83.98 to 856.92 g/ha. The soil depositions (in percentage of applied doses) progressively decreased from 31.15 % to 0 % from 0 to 20 m of the parcel for the fungicides whereas for the herbicides, there is only deposition at 0 m reaching an average of 61.2 % for one of two formulations

Monitoring metal ions present in the effluent discharged from a surface treatment plant: Analytical results

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Embryotoxicity testing of pesticides on snail eggs

To assess the hazards of chemicals for the soil, juveniles and adults of several soil organisms such as earthworms or snails have been used as biological models. To our knowledge, no terrestrial standardised bioassay concerns the effects of pollutants on the embryonic development of soil invertebrates even though this stage is a crucial part of an organism's life cycle. Moreover, embryos are generally more sensitive to pollutants than juveniles and adults. In this context, we aim to develop a bioassay allowing the assessment of the effects of chemicals on snail eggs, first focussing on some widely used pesticides. The chosen pesticides were commercial formulations of herbicides (Roundup®, Bypass®, and Basta®) and fungicides (Corail®, Valiant Flash®, Cabrio Top®, Bordeaux mixture and Thiovit®). Glyphosate, the active ingredient (a.i.) of two formulations (Roundup® and Bypass®), was also tested alone. Aqueous phase bioassay was performed to mimic exposure of eggs to the soil solution. For each product, hatching success was determined at the end of the embryonic development. For all contaminants, EC50 were lower than recommended concentrations for agricultural usage. The toxicity of glyphosate alone (EC50 = 1.3 g/l) was lower than those of commercial formulations (EC50 Bypass® = 0.09 g a.i./l and Roundup® = 0.018 g a.i./l), possibly in relation to the presence of surfactants in the commercial products. This work shows that snail embryos are a sensitive model to assess the hazard of pesticides for a non-target soil organism and constitutes a first step in the development of a new terrestrial embryotoxicity bioassay

Ecotoxicity testing of pesticides at the soil surface using a non-target terrestrial gastropod.

International audienceFate and effects of pesticides in agrosystem were studied using a non target species living at the interface soil-plant-air, the landsnail Helix aspersa. Fate in and effects on snails of 8 commercial herbicide and fungicide formulations (Roundup® and Bypass® = glyphosate; Basta® = glufosinate; Valiant® Flash = folpet + cymoxanil + fosetyl-Al; Corail® = tebuconazole; Cabrio® Top = pyraclostrobin + metiram-Zn; Thiovit® = sulphur; Bordeaux mixture = copper salts) commonly employed in vineyards were studied in the field and for some of them, in laboratory experiments. Here we present a synthesis of a four-years program that aims to develop various biological methods for in situ biomonitoring and life cycle exposure under controlled conditions. A field study in real conditions of pesticide application revealed few effects on the survival and growth of juvenile snails exposed in microcosms to each different treatment. Glyphosate (4 mg kg-1) and AMPA (8 mg kg-1) were detected in snail tissues. The transfer of pyraclostrobin and tebuconazole was also demonstrated. Internal concentrations were low but however higher than LMR drawn for animal products. The effects of two herbicides, Bypass® and Basta® were studied on the whole life cycle of snails in laboratory conditions. No toxicity was found on survival and growth although an accumulation of glyphosate was revealed in snails (6 mg kg-1) exposed continuously to contaminated food (30 mg kg-1). Bioassays for embryotoxicity testing of chemicals in solution or in the soil showed that all pesticides inhibited egg hatching success at lower concentrations than those applied in the field and the influence of soil characteristics on the toxicity of the pesticides was demonstrated. As bioassays using eggs are considered as in vitro methods, efforts should be given to improve and promote the use of such method. This work led to the proposal of complementary tools available for pesticide risk assessment in terrestrial environment. Analytical difficulties were encountered and it was not possible to finalize the analytical methods for all substances studied. Thus, the determination of pesticide residues in animal tissues still represents an essential challenge for current environmental concerns