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Assessment of the Environmental Fate of Pesticides and Their Assessment of the environmental fate of pesticides and their ecotoxicological impact on soil microorganisms : the case of chlorpyrifos, isoproturon and tebuconazole Véronika Storck To cite this version: Véronika Storck. Assessment of the environmental fate of pesticides and their ecotoxicological impact on soil microorganisms : the case of chlorpyrifos, isoproturon and tebuconazole. Life Sciences [q-bio]. Université de Bourgogne, 2016. English. tel-02801785 HAL Id: tel-02801785 https://hal.inrae.fr/tel-02801785 Submitted on 5 Jun 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Distributed under a Creative Commons Attribution - ShareAlike| 4.0 International License University of Burgundy, Dijon (France) Doctoral School of Environment and Health (ED E2S), Dijon (France) National Institute of Agronomical Research (INRA), Dijon (France) DOCTORAL THESIS by Veronika Storck Assessment of the environmental fate of pesticides and their ecotoxicological impact on soil microorganisms The case of chlorpyrifos, isoproturon and tebuconazole Intended viva: 5th December 2016 Jury members: Supervisor: Dr. Fabrice Martin-Laurent INRA, Dijon, France Co-supervisor: Dr. Marion Devers-Lamrani INRA, Dijon, France Reviewers: Dr. Edward Topp Agriculture and Agri-Food, London, Canada Dr. Kathrin Fenner Eawag, Dübendorf, Switzerland Examiners: Dr. Isabelle Batisson CNRS, Clermont-Ferrand, France Dr. Stéphane Pesce IRSTEA, Lyon, France Jury president: Prof. Jean Guzzo University of Burgundy, Dijon, France The research of this thesis was carried out at the National Institute of Agronomical Research (INRA) in Dijon (France), at the Catholic University of the Sacred Heart in Piacenza (Italy) and at Enoveo srl. in Lyon (France). The research was financially supported through the PhD studies of Veronika Storck at the Doctoral School of Environment and Health (ED E2S) at the University of Burgundy funded by the French Ministry of Education and Research (MESR) (grant number 2013-31), and through the European Marie Curie project ‘Love-to-hate’ (FP7 – PEOPLE – 2012 – IAPP) funded by the European Commission (grant number 324349). INRA of Dijon, France Assessment of the environmental fate of pesticides and their ecotoxicological impact on soil microorganisms The case of chlorpyrifos, isoproturon and tebuconazole Veronika Storck born 5th May 1988 Essen, Germany Supervisor: Dr. Fabrice Martin-Laurent (INRA, Dijon, France) Co-supervisor: Dr. Marion Devers-Lamrani (INRA, Dijon, France) Reviewers: Dr. Edward Topp (Agriculture and Agri-Food, London, Canada) Dr. Kathrin Fenner (Eawag, Dübendorf, Switzerland) Examinants: Dr. Stéphane Pesce (IRSTEA, Lyon, France) Dr. Isabelle Batisson (CNRS, Clermont-Ferrand, France) Jury president: Prof. Jean Guzzo (University of Burgundy, Dijon, France) ‘We cannot solve our problems with the same thinking we used when we created them’ Albert Einstein Contents Thesis outline 10 Chapter 1 General introduction 16 Chapter 2 Towards a better pesticide policy for the European Union 52 Chapter 3 Pesticide dissipation in agricultural soil 82 Chapter 3.1 Dissipation and adsorption of isoproturon, 86 tebuconazole, chlorpyrifos and their main transformation products under laboratory and field conditions Chapter 3.2 Identification and characterization of 118 tebuconazole transformation products in soil by combining suspect screening and molecular typology Chapter 4 Ecotoxicological impact of pesticides on soil microorganisms 158 Chapter 4.1 Lab-to-field evaluation of the ecotoxicological 162 impact of three pesticides on the soil bacterial diversity by Illumina nexy-generation sequencing Chapter 4.2 Impact of repeated treatments of the pesticides 204 chlorpyrifos or tebuconazole on soil microorganisms using alternative methods Chapter 5 Possibilities and limitations of using genes as bioindicators 248 for potential pesticide degradation in soil – the case of pdm- encoded isoproturon degradation Chapter 6 General discussion 274 Summary 306 Acknowledgements 310 List of abbreviations 314 List of authors 320 List of publications 324 List of tables 330 List of figures 336 Thesis outline Preview of this thesis This thesis is divided into six chapters (Thesis outline Figure). Chapter 1 is a general introduction containing background information on the topics that chapters 2 to 5 are based upon. It introduces the pesticide benefit to humanity regarding its important role in modern agriculture, and its downside regarding adverse effects in the environment and for human health. It indicates that the environmental risk assessment procedure of pesticides needs improvements that may be achieved by reforms of the current pesticide policy, and states that especially the impact of pesticides on soil microorganisms important for ecosystem functioning is not sufficiently evaluated during the pesticide authorization process. Moreover, this chapter provides an overview of currently available methods able to evaluate potential pesticide impacts on soil microorganisms, including (i) quantitative and qualitative approaches in analytical chemistry to estimate the exposure of an ecosystem to pesticides and their transformation products (TPs), and (ii) International Standard Organization (ISO) approaches and ‘under-development techniques’ such as Illumina next-generation sequencing to study the response of soil microorganisms to pesticide exposure. Furthermore, this chapter presents the interest of using pesticide degradation-encoding genes as bioindicators sensitive to pesticide exposure. It explains the contribution of this thesis to a European Marie Curie project (‘Love-to-Hate’) and describes the main characteristics of the three chosen model pesticides chlorpyrifos (CHL), isoproturon (IPU) and tebuconazole (TCZ). Finally, the chapter provides the aim of this thesis and the research questions that were aimed to answer by this research. Chapter 2 focuses on pesticide environmental risk assessment. It aims to support the ongoing debate of the society about weaknesses in the current pesticide legislation in Europe. The pesticide authorization process is thoroughly explained and weak points in the pesticide environmental risk assessment are identified. Furthermore, the chapter discusses possibilities for improving the robustness and reliability of the pre- and post- registration regulatory framework of pesticides. Chapter 3 investigates the dissipation of CHL, IPU or TCZ in an agricultural soil (agricultural field in North Italy) and is subdivided into two parts. Chapter 3.1 aims to estimate the level and duration of the exposure of soil microorganisms to the three studied pesticides (applied at three different concentrations). Their dissipation and the Thesis outline 11 formation and dissipation of their main TPs in soil are quantified, as evaluated in a microcosm and a field study. Moreover, the sorption of these molecules to soil is estimated from a batch study. As reference standards of potentially interesting TCZ TPs were not available, their formation, dissipation and soil sorption were not evaluated in this chapter. This lack induced the additional study presented in Chapter 3.2 describing a newly combined approach of (i) suspect screening for the identification of known and unknown (suspected) molecules without the imperative need of reference standards and (ii) molecular typology for the characterization of identified molecules. The development of this approach enabled the comprehensive identification and characterization of TCZ TPs in soil during the field study mentioned before. After defining the pesticide exposure scenario via the research on the environmental dissipation of pesticides, the aim of Chapter 4 is the evaluation of the ecotoxicological impact of pesticides on soil microorganisms. In Chapter 4.1, the impact of a single application of the three pesticides on soil bacteria (lower tier pesticide exposure scenario) is evaluated in the microcosm and field studies described before. To achieve this aim, soil DNA was extracted and bacterial DNA (the hypervariable V3-V4 region of the 16S rDNA) was amplified and then analyzed by Illumina next-generation sequencing. In Chapter 4.2, the impact of the repeated application of CHL and TCZ (higher tier pesticide exposure scenario) on their mineralization pattern in soil, on soil buffering functions for other molecules, and on soil bacteria (the latter evaluated by Illumina next-generation sequencing) was assessed. In Chapter 5, the possibility of using pesticide degradation-encoding genes as bioindicators for the pesticide degradation potential of a soil was tested by quantifying the abundance of pdmA and pdmB genes (involved in IPU degradation) as bioindicators for IPU degradation in two Central Eastern French soils in a microcosm and a field study. Chapter 6 is a general discussion critically analyzing the findings of this thesis. It describes the feasible changes of the current pesticide policy that would be necessary to make the pesticide authorization process scientifically more reliable. Moreover, forces and
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