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Approche Mécanistique De La Réponse De La Palourde Japonaise

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Approche Mécanistique De La Réponse De La Palourde Japonaise Approche mécanistique de la réponse de la palourde japonaise, Ruditapes philippinarum, exposée à la bactérie Vibrio tapetis : influence de la température et du régime algal Gaëlle Richard To cite this version: Gaëlle Richard. Approche mécanistique de la réponse de la palourde japonaise, Ruditapes philip- pinarum, exposée à la bactérie Vibrio tapetis : influence de la température et du régime algal. Bi- ologie animale. Université de Bretagne occidentale - Brest, 2015. Français. NNT : 2015BRES0103. tel-02272897 HAL Id: tel-02272897 https://tel.archives-ouvertes.fr/tel-02272897 Submitted on 28 Aug 2019 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. THÈSE / UNIVERSITÉ DE BRETAGNE OCCIDENTALE présentée par sous le sceau de l’Université européenne de Bretagne Gaëlle Richard pour obtenir le titre de DOCTEUR DE L’UNIVERSITÉ DE BRETAGNE OCCIDENTALE Préparée à l'Institut Universitaire Européen de la Mention : biologie marine Mer (IUEM), au Laboratoire des Sciences de École Doctorale des Sciences de la Mer l'Environnement Marin (LEMAR, UMR 6539) Approche mécanistique de Soutenance prévue le 17 décembre 2015 devant le jury composé de : Réjean TREMBLAY la réponse de la palourde Professeur, Institut des Sciences de la Mer de Rimouski / Rapporteur Ingrid ARNAUDIN japonaise, Ruditapes Maitre de Conférences, Université de la Rochelle / Rapporteur Marie-Agnès TRAVERS philippinarum, exposée à la Chercheur IFREMER, La Tremblade / Examinateur Fabrice PERNET bactérie Vibrio tapetis : Chercheur IFREMER, Brest / Examinateur Christine PAILLARD Directrice de recherche, CNRS, Brest / Co-directrice de thèse influence de la température 1 Fabienne GUERARD Professeur, UBO, Brest / Directrice de thèse et du régime algal Sommaire Introduction ................................................................................................................................ 1 Contexte général .................................................................................................................... 2 Problématique et objectifs de la thèse .................................................................................. 6 Etat de l’art ................................................................................................................................. 9 1. La palourde japonaise, Ruditapes philippinarum ......................................................... 10 1.1. Systématique et principales caractéristiques ........................................................ 10 1.2. Biologie de la palourde japonaise .......................................................................... 11 2. Mécanismes de la réponse des invertébrés marins exposés à des stresseurs environnementaux .............................................................................................. 19 2.1. Qu’est-ce que le stress ? ........................................................................................ 19 2.2. Retour sur les réponses générales à des stresseurs chez les vertébrés et les invertébrés ....................................................................................................................... 20 2.3. La réponse immunitaire chez les invertébrés marins ............................................ 25 2.4. Modulation de la réponse immunitaire par l’environnement............................... 50 2.5. Le métabolisme énergétique chez les invertébrés marins .................................... 58 3. Un modèle d’interaction hôte-pathogène-environnement : la maladie de l’anneau brun (MAB) ........................................................................................................................... 64 3.1. La MAB : signes cliniques ....................................................................................... 64 3.2. Caractérisation du pathogène associé à la MAB: Vibrio tapetis ............................ 67 3.3. Les différents modes d’infection de R. philippinarum par V. tapetis en milieu contrôlé ............................................................................................................................ 70 3.4. Réponses de R. philippinarum à l’infection par V. tapetis ..................................... 71 3.5. Influence de l’environnement sur l’interaction R. philippinarum / V. tapetis ....... 76 Matériel et méthodes .............................................................................................................. 78 1. Matériel biologique et conditionnement ..................................................................... 79 1.1. Expérimentation préliminaire : existe-t-il des variations endogènes des activités SOD, CAT et PO chez Ruditapes philippinarum ? ............................................................. 79 1.2. Expérimentation 1 : effet de la température sur l’interaction R. philippinarum – V. tapetis ............................................................................................................................... 80 1.3. Expérimentation 2 : effet du régime algal et de la maturité sexuelle sur l’interaction R. philippinarum – V. tapetis ....................................................................... 81 2. Design expérimentaux et plans d’échantillonnage ...................................................... 85 2.1. Expérimentation préliminaire ................................................................................ 85 2.2. Expérimentation 1 .................................................................................................. 86 2.3. Expérimentation 2 : étude individuelle ................................................................. 87 2.4. Expérimentation 2 : sous-échantillonnage et étude par pool ............................... 88 3. Prélèvements ................................................................................................................ 90 3.1. Prélèvement des fluides......................................................................................... 90 3.2. Prélèvement des organes ...................................................................................... 91 4. Indices de condition ..................................................................................................... 92 5. Cytométrie en flux ........................................................................................................ 92 5.1. Concentration, viabilité, taille et complexité des hémocytes .................................. 92 5.2. Taux de phagocytose ................................................................................................ 93 5.3. Production d’ERO ..................................................................................................... 93 6. Histologie : sexage et détermination des stades de maturité ..................................... 94 6.1. Préparation des échantillons ................................................................................. 94 6.2. Colorations et montage ......................................................................................... 95 6.3. Analyse qualitative au microscope optique ........................................................... 96 7. Diagnostic de la maladie de l’anneau brun .................................................................. 97 7.1. Qualitativement : via l’observation des signes cliniques et l’utilisation d’une clé de détermination.............................................................................................................. 97 7.2. Quantitativement : via l’analyse d’images ............................................................ 98 7.3. Prévalence et intensité de la MAB ......................................................................... 99 8. Dosages biochimiques .................................................................................................. 99 8.1. Protéines ................................................................................................................ 99 8.2. Dosages enzymatiques ......................................................................................... 101 8.3. Lipides .................................................................................................................. 112 8.4. Glucides ................................................................................................................ 114 9. Quantification de V. tapetis dans les fluides extrapalléaux ....................................... 114 10. Analyses statistiques .............................................................................................. 115 10.1. Analyse de coordonnées principales de matrices de voisinage et partition de la variance 115 10.2. Analyses de la variance : plan d’expérience en parcelles divisées .................. 117 10.3. Analyse de variance multivariée par permutations ......................................... 117 10.4. Modélisation Linéaire Mixte ............................................................................ 118 10.5. Modélisation Linéaire Généralisée Mixte : régression logistique ...................
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