Philosophiae Doctor - PhD / Pesticides are designed to control or eliminate pests such as insects, rodents, weeds, bacteria, and fungi. They are used at a global scale for agricultural produce. Although pesticides play a significant role in increasing food production and eliminating diseases, exposure to pesticides may be harmful to non-target organisms. As a result concern over safety and esistance to pesticides has increased and there is pressure to reduce use and search for more environmentally and toxicologically safe and efficacious pesticides. Most pesticides currently in use are synthetic; therefore an alternative to synthetic pesticides is the use of naturally occurring products/ botanicals with pesticidal properties. Two plants indigenous to South African with pesticidal properties were chosen for this study. Dicerothamnus rhinocerotis (D. rhinocerotis) and Galenia africana (G. africana) have potential antifungal properties thus, may have potential use on agricultural produce as fungicides. Galenia africana and D. rhinocerotis extracts inhibit growth of B. cinerea (a fungal pathogen) at concentrations greater than 31.25 mg/ml and 125mg/ml respectively. A major consideration in approving pesticides for use is whether they pose an unreasonable risk to humans and to the environment. Toxicity studies are required to determine the safety of the plant extracts. The purpose of this study was to evaluate potential toxicity of ethanol extracts of D. rhinocerotis and G. africana, which is important when designing practices to reduce or eliminate excess exposure to them. Natural plant products with pesticidal properties could provide an alternative to synthetic pesticides and may thus effectively reduce resistance levels. This first objective of this study was to assess the cytotoxicity of D. rhinocerotis and G. africana on human cell cultures. Human whole blood and the human breast adenocarcinoma cell line (MCF-7) were treated with varying concentrations of the plant extracts and cytotoxicity determined. Cytotoxicity was measured using several biomarkers. Inhibiotory concentration for a 50% effect (IC50) and no observable effect level (NOEL) values were obtained for D. rhinocerotis and G. africana. The results showed that extracts of D. rhinocerotis and G. africana had cytotoxic effects on the cell cultures. The second objective of this study was to determine the ecotoxicity of D. rhinocerotis and G. africana. A series of acute toxicity tests, with effective concentration for a 50% effect (EC50) and lethal concentration for a 50% effect (LC50) as biomarkers, were conducted to estimate the potential environmental effect of the two plants. The tests were carried out using Vibrio fischeri, Selenastrum capricornutum, Daphnia pulex, and Poecilia reticulata as bioindicators. Results obtained showed that G. africana had higher toxicity units than D. rhinocerotis, thus showing that G. africana is more toxic to the aquatic species used as compared to D. rhinocerotis. The third objective of this study was to investigate the immunomodulatory effects of the two plant extracts. This was achieved by using mouse spleen cell cultures. Exposure of pesticides to the immune system may result in alteration of the normal immune functions. The cytokines IFN-γ and IL-4 were used as biomarkers to determine the T-cell activity of the immune system when exposed to the two botanical products. The results obtained showed that both D. rhinocerotis and G. africana decrease of the cytokines interferongamma (IFN-γ) and interleukin-4 (IL-4), thus may have immunotoxic effects. The fourth objective was to investigate the hepatotoxicity of the two plant extracts. Injury to the liver was investigated using a range of clinical biochemical tests that monitor liver enzyme activity and metabolic activity. Primary liver cell cultures were exposed to the plant products in question, after which the biochemical tests were carried out. The enzymes that were monitored were alanine aminotrasferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and lactate dehydrogenase (LDH). The results obtained showed that both of D. rhinocerotis and G. africana may have effects on the liver, as shown by the increased levels of enzymes released from cells upon exposure to plant extracts. The final objective of this study was to investigate the effect of the two plants on the male reproductive system. Injury to the male reproductive system was investigated using testicular cell cultures. Primary cell cultures were stimulated with luteinizing-hormone (LH) and exposed to the plant extracts. LH results in the production of testosterone, thus testosterone was used as a biomarker for assessing reproductive toxicity. The results obtained showed that both of D. rhinocerotis and G. africana have effects on the male reproductive system, as shown by the decreased testosterone secretion. Botanicals provide a simple, inexpensive and environmentally friendly (non-pollution and lesser toxicological concerns) alternative for pest control. However, motivation for the commercial use of botanicals as pesticides requires validating the efficacy of the plant as a pesticide, and also assessing its effects on human health and the environment. An important component of this evaluation involves toxicity studies, which enables cautions of dangerous practices and toxic effects of the plants to be issued. / South Africa

En Lorraine, le drainage fréquent des sols argileux favorise les transferts des intrants agricoles (pesticides et engrais) vers les eaux de surface. Dans ce cas, les bandes enherbées réglementaires (BCAE de la PAC) installées en bordure de cours d’eau sont court-circuitées et ne jouent plus leur rôle épurateur vis-à-vis des eaux de surface. Ainsi, des zones tampons humides artificielles (ZTHA) sont préconisées pour prendre le relais des bandes enherbées. Généralement, ces ZTHA sont installées à l’échelle du bassin versant ou du groupe de parcelles et sont donc de taille conséquente, les rendant concurrentielles de la surface agricole utile (SAU). L’objectif de ce projet de recherche est donc d’évaluer l’efficacité de deux ZTHA de taille réduite, installées dans la bande enherbée entre la parcelle drainée et le ruisseau. Pour cela, une approche multi-échelles intégrant des expériences de laboratoire en batch, des suivis dynamiques en bacs pilotes et un monitoring à la parcelle a été mise en place. Afin d’évaluer l’efficacité des ZTHA au cours des saisons de drainage 2012/13 et 2013/14, le suivi des teneurs en nitrate et de 79 pesticides a été réalisé dans les eaux en entrée et en sortie des ZTHA, ainsi que dans diverses matrices environnementales (sols, sédiment, paille, végétaux). Pour les deux dispositifs, la réduction du flux de nitrate s’est montrée stable, comprise entre 5,4 et 10,9 %, alors que la réduction du flux de pesticides s’est montrée très variable, comprise entre -618,5 et 100 %. Cette variabilité a été expliquée par quatre comportements distincts des molécules. Les fortes valeurs négatives d’efficacité ont été expliquées par une contamination des dispositifs à la suite d’épisodes de ruissellement connexes pour des molécules solubles telles que le 2,4-MCPA ou l’isoproturon. Pour ces deux molécules, l’étude de sorption a montré leur faible affinité avec les matrices environnementales, facilitant leur transport par les eaux de ruissellement et de drainage. D’autres molécules, telles que le boscalide ou l’OH-atrazine, ont présenté des abattements proches de 0. Pour le boscalide, ce comportement a été montré au laboratoire, en accord avec ses fortes capacités d’adsorption et de désorption observées lors des études de sorption et en bacs pilotes. Enfin, certaines molécules ont montré des abattements, allant de 9,9 à 100 %, en lien avec leur faible persistance (2,4-MCPA, clopyralid, mésosulfuron-méthyl,…) ou leur forte affinité pour les matrices environnementales (diflufenicanil, propiconazole, propyzamide,…). Ces ZTHA de taille réduite, installées dans les bandes enherbées, sont donc efficaces pour limiter la contamination des eaux de surface par les eaux de drainage agricole. Ainsi, la multiplication de ces dispositifs à l’échelle de la parcelle dans le paysage agricole permettrait de réduire la contamination des eaux de surface par les pesticides et les nitrates tout en préservant la SAU. Toutefois, l’efficacité de ces ZTHA restant limitée vis-à-vis de certains pesticides présentant un Koc faible et une DT50 élevée, leur installation ne doit pas remettre en cause la réduction en amont des apports phytosanitaires sur les parcelles / In Lorraine, the drainage of clay soils contributes to transfer of agricultural inputs (pesticides and fertilizers) towards surface waters. In this case, the regulatory grass strips (CAP regulation) installed along rivers are bypassed and no longer effective in purifying surface water. Thus, constructed wetlands (CWs) are recommended to take over grass cover bands. Generally, these CWs are installed at watershed scale or at that of several plots and they are therefore quite huge, making them competitive in the cultivated land. The objective of this project is to assess the effectiveness of two small CW, installed in the grass cover band between the drained plot and the river. For this, a multi-scale approach was set up integrating laboratory experiments in batch, pilot study taking into account the water dynamics and monitoring at field scale. The effectiveness of CW was assessed during the 2012/13 and 2013/14 drainage seasons. The nitrate and 79 pesticides concentrations were monitored at inlet and outlet of CWs and in various substrates (soils, sediment, straw and plants). For both devices, the removal efficiencies measured for nitrate were stable and ranged from 5.4 to 10.9 % of inlet amounts while the reduction of pesticide flows was very variable, ranging from -618.5 to 100 %. This variability was explained by four distinct behaviors. The high negative efficiency values were attributed to neighboured runoff events after heavy rains, for soluble molecules such as 2,4-MCPA or isoproturon. For both molecules, the sorption study showed low affinity for environmental substrates, facilitating their transport by runoff and drainage water. Other molecules such as boscalid or OH-atrazine showed effectiveness close to 0 %. For boscalid, this behavior has been observed in laboratory sorption and pilots studies, in agreement with its strong adsorption and desorption capacities. Finally, some molecules have shown reductions ranging from 9.9 to 100 %, in agreement with their low persistence (2,4-MCPA, clopyralid, mesosulfuron-methyl, ...) or their high affinity for environmental substrates (diflufenican, propiconazole or propyzamide). These small CWs, located in the grass cover bands, are effective in minimizing the contamination of surface waters by agricultural drainage water. Thus, multiplying the number of these devices at the plot scale in the agricultural landscape would reduce the contamination of surface waters by pesticides and nitrates while preserving the cultivated land. However, the effectiveness of these CWs are limited for pesticides with a low Koc and a long DT50; thus their installation in plot must not call into question the reduction in the use of pesticides

Dans cette étude, des aspects importants du rapport entre pesticides et abeilles domestiques (Apis mellifera L.) ont été traités. Dans la première partie, les effets de l’exposition des abeilles aux poussières contaminées avec trois neonicotinoides et le fipronil ont été étudiés. En fait, des quantités considérables de ces pesticides, utilisés pour l’enrobage des semences, sont dispersées pendant le semis du mais, et peuvent donc représenter une voie d’exposition des abeilles.En particulier, une voie spécifique d’exposition, le contact indirect, a été pris en compte. Les effets létaux et sub-létaux (mortalité aigue, butinage, développement des colonies, capacité d’orientation) des poussières ont été évalués en laboratoire, en tunnel et en plein champ. La dispersion réelle des poussières pendant le semis avait été évalué précédemment.Les résultats ont montré un effet significatif, sur la mortalité, de l’exposition aux poussières contaminées avec neonicotinoides et fipronil, en laboratoire et en tunnel. Par contre, la capacité d’orientation des abeilles n’a été pas influencée par l’exposition aux concentrations testées.Dans la deuxième partie de la recherche, on a étudié l’impact de différentes pesticides (chimiques et biologiques) sur les changements biochimiques et physiologiques des abeilles exposées. Ces paramètres on été évalué pour différentes façons et durées d’exposition. En particulier, trois expérimentations ont été effectuées en combinant les spores de Bacillus thuringiensis avec la deltamethine, les spores de Bt avec le fipronil et le traitement avec deltamethrine et le fongicide difenoconazole. Certains enzymes impliqués dans la détoxification, le stress oxydant et le métabolisme énergétique (GST, ALP, SOD, CAT, G6PDH, GAPDH) ont été sélectionnés pour évaluer les variations de leur activité suite à l’exposition aux pesticides. L’analyse des différents indicateurs biochimiques, comme le GST et le ALP, a mis en évidence des variations physiologiques qui peuvent être liés à l’exposition aux pesticides. Cette méthodologie pourrait donc représenter un nouvel aspect de l’évaluation des effets sub-létaux des pesticides chez l’abeille. / In this study, some important aspects of the relationship between honey bees (Apis mellifera L.) and pesticides have been investigated. In the first part of the research, the effects of the exposure of honey bees to neonicotinoids and fipronil contaminated dusts were analyzed. In fact, considerable amounts of these pesticides, employed for maize seed dressing treatments, may be dispersed during the sowing operations, thus representing a way of intoxication for honey bees. In particular, a specific way of exposure to this pesticides formulation, the indirect contact, was taken into account. To this aim, we conducted different experimentations, in laboratory, in semi-field and in open field conditions in order to assess the effects on mortality, foraging behaviour, colony development and capacity of orientation. The real dispersal of contaminated dusts was previously assessed in specific filed trials. The results showed a significant effect on mortality of neonicotinoids and fipronil contaminated dusts, both in laboratory and in semi-field trials. However, no effects were evidenced in honey bees orientation capacity.In the second part, the impact of various pesticides (chemical and biological) on honey bee biochemical-physiological changes, was evaluated. Different ways and durations of exposure to the tested products were also employed. Three experimentations were performed, combining Bt spores and deltamethrin, Bt spores and fipronil, difenoconazole and deltamethrin. Several important enzymes (GST, ALP, SOD, CAT, G6PDH, GAPDH) were selected in order to test the pesticides induced variations in their activity. In particular, these enzymes are involved in different pathways of detoxification, oxidative stress defence and energetic metabolism. The analysis of different biochemical indicators highlighted some interesting physiological variations that can be linked to the pesticide exposure. We therefore stress the attention on the possibility of using such a methodology as a novel toxicity endpoint in environmental risk assessment.

La zone non saturée joue un rôle clé sur le transfert des pesticides et la qualité des eaux souterraines. Les connaissances sur les processus d’écoulement et de rétention dans les matériaux géologiques de la zone non saturée au-delà des sols sont toutefois parcellaires. Le transfert du glyphosate et du S-métolachlore (SMOC), et de leurs métabolites AMPA, ESA-métolachlore (MESA) et OXA-métolachlore (MOXA) est étudié en colonne pour deux matériaux fluvio-glaciaires issus d’un aquifère de l’Est lyonnais : un sable, S-x, et un mélange bimodal de graviers et de sables, Gcm,b. Pour des conditions de non-saturation en eau, l’écoulement dans les colonnes est fractionné en deux zones, eau mobile et eau immobile, d’importance variable suivant le solide. La sortie du SMOC est retardée par rapport au traceur de l’eau ; son bilan de masse déficitaire traduit une rétention de la molécule lors de son transfert. A l’inverse, le MESA et le MOXA se comportent comme le traceur de l’eau. Le glyphosate et l’AMPA sont très peu mobiles dans la colonne de Gcm,b (seul matériau étudié) avec des quantités éluées inférieures à 1% de la quantité appliquée. La modélisation montre que le transfert des molécules est affecté de manière variable suivant le matériau par la cinétique physique d’échange entre les zones d’eau mobile et immobile et par la cinétique chimique des molécules. Cette cinétique chimique est décrite par des expérimentations complémentaires de sorption en batch. La caractérisation des matériaux révèle la présence d’oxydes et de minéraux argileux qui pourrait expliquer leur forte réactivité, qui s’avère parfois supérieure à celle des sols de la zone d’étude. / Vadose zone play a key role in pesticides transfer and groundwater quality. Knowledge’s about leaching and retention processes in the vadose zone below the shallow soil zone are still poorly understood. Transfer of glyphosate, S-metolachlor (SMOC), and their metabolites AMPA, ESA-metolachlor (MESA) and OXA-metolachlor (MOXA) is studied in unsaturated columns filled with two glaciofluvial materials collected in the East of Lyon: a sand, S-x, and a bimodal gravel, Gcm,b. Experiments show water fractionation into mobile and immobile compartments with variable importance according to material column. SMOC outflow is delayed compared to the conservative tracer. SMOC mass balance is in deficit revealing retention in columns. At the opposite, complete mass elution associated with retardation factors close to unity shows that there is no adsorption of MESA and MOXA in either lithofacies. Glyphosate and AMPA mobility is very low in the one Gcm,b column studied with amounts in leachates inferior to 1% of applied. Modelling show pesticides and metabolites transfer is affected by both flow regionalisation and non-equilibrium sorption. Chemical kinetic of sorption mechanisms is studied with complementary batch experiments. The high glaciofluvial materials reactivity, in some cases upper than soil reactivity from the study site, could be attributed to oxides and clay minerals.

Amitraz is a formamide acaricide used predominantly in the control of ectoparasites in livestock and honeybees. Amitraz hydrolysis is rapid and occurs under acidic conditions, exposure to sunlight and biodegradation by microorganisms. The main hydrolysis product of amitraz, 2,4-dimethylaniline, is recalcitrant in the environment and toxic to humans. An electrochemical method for the determination of total amitraz residues and its final breakdown product, 2,4-dimethylaniline, in spent cattle dip, is presented. Cyclic voltammetry at a glassy carbon electrode showed the irreversible oxidation of amitraz and 2,4-dimethylaniline. A limit of detection in the range of 8.5 x 10⁻⁸ M for amitraz and 2 x 10⁻⁸ M for 2,4-dimethylaniline was determined using differential pulse voltammetry. Feasibility studies in which the effect of supporting electrolyte type and pH had on electroanalysis of amitraz and its degradants, showed that pH affects current response as well as the potential at which amitraz and its degradants are oxidised. Britton-Robinson buffer was found to be the most suitable supporting electrolyte for detection of amitraz and its degradants in terms of sensitivity and reproducibility. Studies performed using environmental samples showed that the sensitivity and reproducibility of amitraz and 2,4-dimethylaniline analyses in spent cattle dip were comparable to analyses of amitraz and 2,4-dimethylaniline performed in Britton-Robinson buffer. In addition, the feasibility qf measuring amitraz and 2,4-dimethylaniline in environmental samples was assessed and compared to amitraz and 2,4-dimethylaniline analyses in Britton-Robinson buffer. Amitraz and 2,4-dimethylaniline were readily detectable in milk and honey. Furthermore, it was elucidated that 2,4-dimethylaniline can be metabolised to 3-methylcatechol by Pseudomonas species and the proposed breakdown pathway is presented. The biological degradation of amitraz and subsequent formation of 2,4-dimethylaniline was readily monitored in spent cattle dip. The breakdown of amitraz to 2,4-dimethylaniline and then to 3-MC was monitored using cyclic voltammetry.

Chaque année en France, les semenciers sont contraints de déclasser plus de 10 000 tonnes de semences. Ces semences sont actuellement incinérées dans des cimenteries au frais des semenciers et non valorisables dans des procédés de valorisation des agro-ressources, compte tenu de leur teneur non négligeable en pesticides. Par conséquent, il apparaît nécessaire de réduire, dans un premier temps, la quantité de produits phytosanitaires présents dans les enrobages avant de pouvoir valoriser ces semences déclassées.L’objet de cette thèse consiste donc à développer un outil permettant de décomposer une grande partie des produits phytosanitaires présents sur les enrobages par ozonation. Dans un premier temps, une méthode d’extraction par liquide pressurisé a été développée pour la quantification des produits phytosanitaires présents sur les enrobages avant et après ozonation. Cette méthode d’extraction a été validée et a permis de déterminer, par la suite,l’efficacité du traitement d’ozonation sur les semences enrobées.Ensuite, afin d’évaluer l’efficacité d’un traitement à l’ozone, deux pesticides présents dans la solution d’enrobage Gaucho Blé® -l’imidaclopride et le bitertanol- ont été ozonés, en solution aqueuse dans un réacteur agité. Les deux produits ont été dégradés complètement en moins de 30 minutes et la loi cinétique de leurs réactions d’ozonation a montré que l’ordre partiel de la réaction a été de 1 pour l’ozone et pour le pesticide. Les produits de dégradation,formés au cours de l’ozonation, ont aussi été analysés par spectrométrie de masse ESI(+)-MSet la structure de la plupart des produits de dégradation a été déterminée. Un schéma des différentes voies de dégradation de l’imidaclopride et du bitertanol a pu être proposé.Enfin, l’ozonation des pesticides a été étudiée sur les semences de blé tendre enrobées avec la solution de Gaucho Blé® dans un réacteur à lit fixe. L’humidification des grains avant ozonation, la concentration en ozone dans le gaz et la durée de traitement ont semblé être les facteurs importants pour accélérer la cinétique de dégradation des pesticides dans les enrobages. Les pesticides ont été dégradés (respectivement 99 et 90 % de dégradation pour l’imidaclopride et le bitertanol). La loi cinétique de la réaction d’ozonation de l’imidaclopride et du bitertanol a présenté des ordres partiels égaux à 1 pour l’ozone et pour le pesticide. Ce procédé a ensuite été appliqué à d’autres types de semences enrobées et s’est avéré efficace sur tous les principes actifs étudiés. Enfin, les semences enrobées ont été ozonées dans un réacteur prototype (réacteur à plateau vibrant) dans lequel tous les pesticides étudiés ont été dégradés mais à des taux de dégradation plus faibles qu’en réacteur à lit fixe. / Every year in France, seed producers must downgrade more than 10,000 tons of seeds. These seeds are currently incinerated in cement factories at seed producers’ expenses and non available for the processes of valorization of renewable resources, due to their significant pesticide content. Consequently, in the first step, the quantity of pesticides present in the seed loading must be reduced before a subsequent valorization of the downgraded seeds. The purpose of this PhD work is thus to develop a process for the decomposition of a large amount of the pesticides present in the seed loading. First, an extraction procedure by pressurized liquid was developed for the quantitation of phytopharmaceuticals present on the loading before and after the ozonation. This extraction method was validated and enabled to determine, subsequently, the efficiency of the ozonation process on the loaded seeds. In the second time, in order to evaluate the efficiency of an ozonation treatment, both pesticides present in the Gaucho Blé® seed loading solution – imidacloprid and bitertanol – were ozonated, in aqueous solutions in a stirred reactor. Both compounds were completely decomposed in 30 minutes and the kinetic law for the ozonation of both compounds displayed a partial order of 1 for ozone and 1 for the pesticide. Degradation products, formed during the ozonation, were analyzed by mass spectrometry ESI(+)-MS and the structure of most of the compounds was characterized. Degradation pathways for imidacloprid and bitertanol were thus proposed. Finally, the ozonation of pesticides were applied to soft wheat seeds loaded with Gaucho Blé® loading solution, in a fixed bed reactor. The humidification of seeds before ozonation, the concentration of ozone in the inlet gas and the ozonation time were the predominant factors to accelerate the kinetic of degradation of pesticides in the seed loadings. The pesticides were successfully decomposed (respectively 99 and 90 % for the degradation of imidacloprid and bitertanol). The kinetic laws for the ozonation reactions of imidacloprid and bitertanol were determined and the partial orders of the reaction were 1 for both ozone and the pesticide. This process was then applied to the treatment of other loaded seeds and showed good degradation rates for all the studied pesticides. Subsequently, the loaded seeds were ozonated in a prototype reactor equipped with a dynamic plate in which all the pesticides were decomposed but with lower degradation rates than in a fixed bed reactor.

Les marchés réclament des fruits parfaits sans symptômes de maladies ou piqures. En conséquence les arboriculteurs appliquent des pesticides ayant des effets négatifs sur l’environnement et la santé. Des méthodes alternatives existent mais peuvent être risquées et réduire les performances agronomiques. Face à ces enjeux et sur l’exemple de la protection en culture de pommes, la plus traitée en pesticides, l’étude vise à répondre à la problématique suivante: quelle démarche mettre en place pour aider les arboriculteurs à évaluer leurs marges de manœuvre et les effets d’une évolution de leur stratégie de production sur le fonctionnement et les performances de leur exploitation?La première phase de la démarche vise à comprendre les stratégies de protection des arboriculteurs et évaluer leurs effets sur les performances des exploitations. Les pratiques d’utilisation des pesticides d’une coopérative ont été analysées à partir de 557 calendriers de traitements, à l’aide d’indicateurs de fréquence, de toxicité et d’efficience. Puis 35 enquêtes qualitatives ont été conduites dans deux régions pour comprendre les processus qui aboutissent à ces pratiques et identifier les leviers et freins à leurs évolutions.Malgré un contexte commun, quatre grands types de pratiques ont été mis en évidence. Trois types se distinguent par une utilisation relativement importante d’un type de pesticide: anti-tavelure, anti-insecte, pesticides naturels, et la quatrième est équilibrée. Pour chaque type, les indicateurs couvrent une diversité de valeurs. Les liens entre eux sont complexes, et montrent que l’indicateur de fréquence cache une diversité de pratiques, n’est pas synonyme d’efficience, et n’explique qu’en partie la toxicité environnementale des parcelles. Des actions pour diminuer l’utilisation des pesticides ont été discutées, comme la prise en compte de la diversité et l’amélioration de la surveillance des ravageurs et maladies correspondants aux pesticides les plus utilisés.L’étude des processus aboutissant aux choix des pratiques de protection à l’échelle de l’exploitation a mis en évidence trois stratégies, répondant à des objectifs environnementaux et économiques différents. Pour chaque stratégie, ces objectifs sont cohérents avec la stratégie commerciale, les ressources en main d’œuvre, la diversification/spécialisation et la surface cultivée. L’influence de l’organisation du travail et de la stratégie commerciale sont à prendre en compte pour aider les stratégies à évoluer et anticiper les freins aux transitions.La seconde phase vise à développer un processus d’accompagnement des producteurs dans leurs réflexions d’évolution de leur stratégie. Un outil de simulation (CoHort) a été coconstruit avec deux techniciens de coopérative. Il représente la structure et le fonctionnement d’une exploitation arboricole, dans sa dimension technique et son organisation du travail. Chaque scénario est évalué après simulation en termes économique, environnemental et de bilan travail. Ces résultats alimentent la discussion entre producteurs et techniciens sur les évolutions des exploitations envisageables. CoHort a été utilisé sur des exploitations ayant des projets d’évolution, puis sur des scénarios de rupture élargissant les types de changement explorés : cas de l’introduction d’ovins dans les vergers.En combinant les échelles parcelle et exploitation, la démarche saisit la complexité et la diversité des processus analysés. Ces connaissances ont permis de développer un processus d’accompagnement des producteurs basé sur un outil générique, qui prend en compte leurs contraintes et la diversité des situations.A moyen terme, l’outil de simulation CoHort sera transmis aux techniciens. La démarche globale pourrait être déployée sur un territoire plus large ou utilisée pour d’autres types de productions arboricoles. Mais pour déclencher des transitions et produire des fruits sans pesticides, la filière entière doit être intégrée aux réflexions. / Markets ask for perfect looking fruits without any symptoms of diseases or insect bites. To fulfill these criteria, fruit tree farmers spray pesticides that have negative impacts on human health and on the environment. Alternative methods can be risky and affect the economic performances of the farm. Based on the example of apple crop protection, the most sprayed fruit tree crop, this study addresses the following issue: which approach can be implemented to support fruit tree farmers in evaluating ways of changing their production strategies and their impacts on farm performances?The first phase of the study aims at describing and understanding the protection strategies implemented by farmers, and at evaluating their impacts on farm performances. Practices of pesticide use were analyzed using 557 spray records of a cooperative. Frequency, toxicity and efficiency indicators were used to evaluate practices’ environmental and health impacts. Then, 35 interviews were conducted in two regions of France, to understand farmers’ decision making processes that influence these practices, and to identify the levers and barriers for evolving towards less pesticide use.Despite a common climatic and marketing context, four types of pesticide use sets were identified. Three types showed a relatively important use of a given pesticide category: anti-scab, anti-insect, natural pesticides. The fourth one was balanced. Within each type, the different indicators covered a large range of scores. The frequency indicator covered a diversity of practices. It was not linked with the efficiency indicator, and it explained only a part of the plots’ environmental toxicity. Possible actions for improvements towards less pesticide use were discussed, such as taking into account the diversity of practices and improving the monitoring of the pests and diseases corresponding to the most sprayed pesticides.Studying the decision making processes at the farm scale highlighted three different strategies, depending on the farmers’ environmental and economic objectives. Each strategy was consistent regarding the farmers’ objectives, their marketing strategy, their labor resources, their position between diversification and specialization, and the apple area. Work organization and marketing strategy were considered as instrumental components for defining alternative strategies and for anticipating implementation difficulties.A methodology aiming at supporting fruit tree farmers in their strategical thinking towards innovative production strategies was developed during the second phase of the study. A simulation tool called CoHort was developed with two technicians. CoHort depicts the structure of a tree fruit farm, the crop management per homogeneous block and its work organization. Each scenario corresponds to a specific configuration of the farm. It is evaluated in terms of economic and environmental performances and of impacts on work organization. The simulation results feed the discussions between farmers and their technicians on the potential evolutions of the farm. CoHort was used at first by addressing some farmers’ projects regarding their farm evolution projects. Then it was used on de novo scenarios allowing exploring new types of evolutions such as combining apple and sheep production within a farm.The analysis at both farm and plot scales permitted to capture the complexity and the diversity of the management processes observed in the farm sample surveyed. It provided the knowledge required to develop a support process based on a generic simulation tool, which can cover a large diversity of farm contexts and projects. A mid-term objective is to transfer CoHort to the technicians for extending their advice activities to strategic issues. The whole approach could also be used at the supply chain scale, or on other fruit tree crops. Indeed, enhancing transitions towards less pesticide used will require to involve the whole fruit supply chains.

L’utilisation des pesticides peut être à l’origine de graves nuisances pour la santé humaine,comme l’ont montré les études en milieu professionnel agricole.L’objectif de cette thèse était d’étudier les manifestations, principalement respiratoires etsurvenant à court terme, dans la population générale exposée aux pesticides utilisés enagriculture.Les résultats de deux études épidémiologiques, Phytoriv, menée en Gironde et Phytoniger, plusexploratoire, sont présentés, toutes deux menées au sein de population d’adultes et d’enfantsvivant en milieu rural.L’étude Phytoriv a mis en évidence la présence de concentrations faibles mais plus élevées depesticides dans l’air ambiant à proximité de zones viticoles traitées et a permis de recueillir surune période de une à trois semaines les symptômes ressentis par les riverains de manièrejournalière. L’absence d’effet majeur de l’exposition sur la santé respiratoire chez les riverainsobservée dans cette étude demande à être confirmée, en améliorant notamment l’estimationtemporelle et spatiale de l’exposition.L’étude Phytoniger a permis de démontrer la faisabilité d’une étude de santé environnementaledans un pays en développement. L’utilisation de pesticides non autorisés et potentiellementdangereux, pour des usages agricoles mais également domestiques, est importante au Niger etpose la question de leur impact sur la santé des populations. Certains symptômes respiratoiresétaient plus fréquemment observés en zone agricole humide, où les usages de pesticides, maiségalement les feux de démoustication, sont plus importants qu’en zone sahélienne agropastorale.Enfin, dans les deux études Phytoriv et Phytoniger, des risques plus élevés de symptômesrespiratoires étaient observés, notamment chez les enfants, en lien avec l’utilisation domestiquede pesticides.A l’avenir, une caractérisation plus précise des sources, niveaux et déterminants d’exposition dela population générale permettrait d’améliorer les connaissances sur le lien avec la survenue desymptômes et pathologies respiratoires et d’identifier les axes d’intervention les plus pertinents. / Pesticides are widely used in agriculture worldwide. However, it is now recognised thatoccupational pesticide exposure, especially in agriculture, can pose serious health concerns. Weaimed to study short term respiratory symptoms in general populations exposed to pesticidesused in agriculture.Here we present results of two epidemiological studies, Phytoriv, performed in Bordeaux area,France, and Phytoniger, performed in a Sahelian African country, both among rural adult andchildren populations.Higher pesticide levels in ambient air were observed in the surrounding of vineyards in Phytoriv.However, no major effect was observed on resident’s respiratory health. This need to beconfirmed by studies with improved temporal and spatial exposure assessment.Phytoniger allowed us to show the feasibility of environmental health studies in a developingcountry. Unauthorized and hazardous pesticides are widely used in Niger, for agricultural as wellas for residential purposes, and concerns are growing on their potential health effects. Some ofthe short term respiratory symptoms were more frequent in the agricultural wetland area,compared to the Sahelian agro-pastoral area. Pesticides are more susceptible to be used in thefirst area, but it is also more subject to mosquito control fires.Finally in both surveys, higher risks of respiratory symptoms have been observed for peopleliving in houses treated with indoor pesticides, and especially in children.All these results warrants further studies to improve assessment of sources, exposure levels anddeterminants of pesticide exposure in the general population, in order to improve knowledge onits respiratory and short term health effects. Thus, relevant public health interventions could beperformed.

MSc (Chemistry) / Department of Chemistry / In this work, modern pre-concentration techniques, namely dispersive liquid-liquid micro-extraction (DLLME) and QuEChERS, were used to analyse pesticides and heavy metals in complex matrices. The work is divided into six papers. In Papers 1, the recent developments and applications of DLLME during analysis of pesticides in food matrices were reviewed. The DLLME technique has captured the interests of many researchers in recent years. The major advantage, among others, of DLLME is miniaturisation in which the acceptor-to-donor ratio is reduced tremendously leading to high enrichment compared to other sample preparation techniques. In the present work, the different complex matrices where the DLLME technique has been employed for the analysis of pesticides are reviewed as well as the challenges associated with this technique. Papers II reviewed the recent applications and developments of the QuEChERS technique during the analysis of pesticides in food matrices. QuEChERS is a versatile pre-concentration method whose application spans the whole breath of organic compounds. There are three common standard methods used during QuEChERS and these are the original QuEChERS, AOAC and the EN methods. In this paper, recent developments and applications of QuEChERS techniques in the analysis of pesticides in food samples were reviewed. In Paper III, green pre-concentration techniques employed during analysis of pesticides were reviewed. Recently, the parameter of “greenness” during sample pre-concentration of pesticides in food matrices is as important as selectivity in order to avoid using large amounts of harmful organic solvents during sample preparation. Developing new green pre-concentration techniques is one of the key subjects in green chemistry in order to minimize the release of large volume of toxic organic solvents into the environment. Thus, to reduce the impact on the environment during trace analysis of pesticides in food matrices, new developments in pesticide pre-concentration have gone in three separate directions (which are reviewed in this paper): one is the search for more environmentally friendly solvents, the second one is miniaturization and the third one is the development of solvent-free pre-concentration techniques. Eco-friendly solvents such as supercritical fluids, ionic liquids and natural deep eutectic solvents have been developed for use as extraction solvents during pre-concentration of pesticides in food matrices. Also miniaturized pre-concentration techniques such as QuEChERS, dispersive liquid-liquid micro-extraction and hollow-fibre liquid phase micro-extraction have been used during trace analysis of pesticides in food samples as well as solvent-free techniques such as solid phase micro-extraction and stir bar sorptive extraction. All these developments are geared to ensure that pesticide pre-concentration in food matrices is green and were reviewed in this paper. The effect of vehicular emissions on the concentrations of selected heavy metals was investigated in Paper IV. The samples were pre-concentrated using DLLME prior to analysis with flame atomic absorption spectroscopy. Dithizone, chloroform and methanol were used as chelating agent, extraction solvent and dispersion solvent respectively during the DLLME technique. The pH of the sample was adjusted to around 8 using sulphuric acid or sodium hydroxide solution. The influential DLLME parameters, such as pH volume and type of extraction solvent, and voume of disperser solvent, were optimized prior to the application of the developed method to real samples (roadside dust, fruits and vegetables). In Paper V, chromium speciation in fruits and vegetables was studied. The chromium in fruit and vegetable sample juices was pre-concentrated using DLLME prior to analysis with flame atomic absorption spectroscopy. Diphenylcarbazide (DPC) was used as a chelating agent in this study, and salting out of the complex from the aqueous medium into the organic phase was effected using sodium acetate. Chloroform and methanol were used as extraction and dispersion solvents respectively in the DLLME method for the determination of chromium (VI). For total chromium, the trivalent chromium was oxidised using acidified KMnO4 to hexavalent chromium before performing the DLLME technique. The concentration of chromium (III) was determined by finding the difference between total chromium and concentration of chromium (VI). The important parameters that influence the efficiency of the DLLME technique were also optimized using the univariate approach. After optimization, the developed method was applied to real samples. In Paper VI, the concentration of malathion pesticide in fruits was determined using QuECHERS for pre-concentration and UV-Vis spectrophotometry for instrumental analysis. Acetonitrile was used as the extraction solvent and Z-sep+/PSA sorbent combination was used for sample clean-up. The acetonitrile extract from QuEChERS was then hydrolysed using KOH followed by reaction with acidified potassium bromate for colour development. The coloured product formed was then analysed using UV-Vis spectrophotometry. Among the fruits analysed, Oranges had no malathion residue in them. However, trace amounts of malathion, below WHO maximum allowable limits, were found in pears and apples. / NRF

En 2000, la Directive Cadre sur l’Eau a exigé le retour au bon état chimique et écologique des cours d’eau. Elle a notamment classé 45 substances comme étant prioritaires pour cette évaluation (directive 2013/39/UE), dont en grande partie des pesticides. En effet, en raison de leur utilisation massive, on retrouve aujourd’hui ces contaminants dans tous les compartiments de l’environnement. Par ailleurs, au vu de sa capacité à intégrer les contaminations, le biofilm est considéré comme un excellent bioindicateur pour l’évaluation de la qualité de l’eau. Celui-ci est à la base de la chaine trophique dans les milieux aquatiques et se compose de microorganismes (microalgues, bactéries, champignons, etc…) enchâssés dans une matrice de substances polymériques extracellulaires (EPS).Cette thèse porte sur l’étude des mécanismes de transfert et de distribution des pesticides dans les biofilms en lien avec les effets toxiques associés. Ces travaux ont été réalisés avec une approche toxicocinétique par laquelle la bioaccumulation du diuron (un herbicide inhibiteur de la photosynthèse) a été suivie dans les différents compartiments du biofilm. Ce suivi a été réalisé sous plusieurs conditions abiotiques (vitesses de courant, températures et photopériodes) pour différentes durées d’exposition. En parallèle des descripteurs fonctionnels et structuraux ont été mesurés comme des activités photosynthétique (pour les communautés autotrophes) et enzymatiques (pour les communautés hétérotrophes), ainsi que la biomasse totale, la production de protéines et de polysaccharides.L’ensemble des expérimentations menées au cours de cette thèse a permis de décrire les mécanismes de sorption du diuron dans les biofilms, à savoir des processus d’absorption par les cellules et d’adsorption par la matrice EPS. Puis l’influence des différents paramètres environnementaux étudiés a pu être mise en avant. Ces travaux démontrent la pertinence de l’approche toxicocinétique-toxicodynamique pour l’étude de l’impact des pesticides sur les biofilms fluviaux. / In 2000, the Water Framework Directive required the return of rivers to good chemical and ecological status. In particular, it has classified 45 substances as priority for this assessment (Directive 2013/39/EU), including a large proportion of pesticides. Indeed, due to their massive use, these contaminants are now found in all compartments of the environment. In addition, given its ability to integrate contamination, biofilm is considered an excellent bioindicator for water quality assessment. It is at the base of the trophic chain in aquatic environments and is composed of microorganisms (microalgae, bacteria, fungi, etc...) embedded in a matrix of extracellular polymeric substances (EPS).This thesis focused on the analysis of mechanisms of pesticide transfer and distribution in biofilms related to toxic impacts. This work was carried out using a toxicokinetic approach whereby the bioaccumulation of diuron (a photosynthesis inhibitor herbicide) was monitored in the different compartments of the biofilm. This assessment was carried out under several abiotic conditions (flow velocity, temperatures and photoperiods) under varying exposure durations. In parallel, functional and structural descriptors were measured as photosynthetic (for autotrophic communities) and enzymatic (for heterotrophic communities) activities, as well as biofilm biomass and protein and polysaccharide production.All the experiments performed during this thesis made it possible to highlight the sorption mechanisms of diuron in biofilm, i.e. absorption processes by cells and adsorption phenomenon within the EPS matrix. Then the influence of the different environmental parameters studied was emphasized. This work demonstrates the relevance of the toxicokinetic-toxicodynamic approach to the study of the impact of pesticides on fluvial biofilms.