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Morphological and Molecular Approaches to the Diversity of Digenean Parasites in Two Sparid Fishes, Diplodus Vulgaris (G, 1817) and Sparus Aurata L

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Morphological and Molecular Approaches to the Diversity of Digenean Parasites in Two Sparid Fishes, Diplodus Vulgaris (G, 1817) and Sparus Aurata L REPUBLIQUE ALGERIENNE DEMOCRATIQUE ET POPULAIRE MINISTERE DE L’ENSEIGNEMENT SUPERIEUR ET DE LA RECHERCHE SCIENTIFIQUE UNIVERSITE ORAN 1 FACULTE DES SCIENCES DE LA NATURE ET DE LA VIE DEPARTEMENT DE BIOLOGIE Laboratoire Réseaux de Surveillance Environnementale THÈSE Présenté par Mohammed RIMA Pour l’obtention du diplôme de DOCTORAT 3ème cycle LMD En Biologie Option : Sciences de la Mer et du Littoral Morphological and molecular approaches to the diversity of Digenean parasites in two sparid fishes, Diplodus vulgaris (G, 1817) and Sparus aurata L. 1758 along the Algerian coasts of the western Mediterranean. Soutenance le : / / 2018. Devant le jury composé de : PRESIDENT : Mme Farida MESLI Professeur, Université Oran 1 Ahmed Ben Bella EXAMINATEUR : Mme Amaria MAATALLAH-BOUTIBA Maitre de conférences A, Université Oran 1 Ahmed Ben Bella EXAMINATEUR : Mr Abdellah BOUDJEMAA Professeur, Université Mohamed Boudiaf (USTO) EXAMINATEUR : Mr Tomáš SCHOLZ Professeur, Institut of Parasitology, République Tchèque EXAMINATEUR : Mme Kheira SENOUCI Professeur, Université Oran 1 Ahmed Ben Bella RAPPORTEUR : Mme Douniazed MARZOUG Maitre de conférences A, Université Oran 1 Ahmed Ben Bella INVITE : Mr Francisco Esteban MONTERO ROYO Institue Cavanilles, Université de Valencia, Spain Année universitaire 2017-2018 TABLE OF CONTENTS RÉSUMÉ i ABSTRACT ii iii الملخص LIST OF FIGURES iv LIST OF TABLES viii REMERCIEMENTS ix DÉDICACES xii List of abbreviations xiii CHAPTER 1. General Introduction 1 CHAPTER 2. literature Review 11 CHAPTER 3. General Materials and Methods 21 CHAPTER 4. New molecular and morphological data for opecoelid digeneans in two 38 Mediterranean sparid fishes with descriptions of Macvicaria gibsoni n. sp. And M. crassigula (Linton, 1910) (sensu stricto) CONCLUSION AND FUTURE PERSPECTIVES REFERENCES. APPENDICES RÉSUMÉ La présente étude porte sur l'application d'une approche intégrative à l'identification des parasites trématodes retrouvés chez deux espèces de poissons sparidés, le Diplodus vulgaris (Geoffroy Saint-Hilaire, 1817) et Sparus aurata L. le long des côtes algériennes de la Méditerranée occidentale. Une zone non explorée sur la diversité des helminthes de poisson. Un total de 810 poissons (390 D. vulgaris et 420 S. aurata) a été examiné dans trois localités (au large de Bouzedjar, d'Alger et d'Annaba). Bénéficiant d'échantillons de grande taille et d'une collaboration fructueuse, nous avons généré une base de données de séquences pour le gène cluster mitochondrial cox1 et nucléaire ITS1-5.8S-ITS2 et / ou les séquences d'ADNr 28S partielles pour un total de 15 espèces de trématodes. Une caractérisation morphologique détaillée est fournie pour ces espèces représentant 11 familles : Opecoelidae (5 espèces de 2 genres); Acanthocolpidae, Aephnidiogenidae, Aporocotylidae, Derogenidae, Hemiuridae, Heterophyidae, Lepocreadiidae, Monorchiidae, Strigeidae (une espèce chacune), et une seule espèce de l'Aspidogastridae (Sous-classe Aspidogastrea). Trois nouvelles espèces pour la science ont été décrites à partir du matériel étudié : Macvicaria gibsoni, Rima, Marzoug, Pérez-del-Olmo, Kostadinova, Bouderbala et Georgieva, 2017; une nouvelle espèce de Cardicola; et une nouvelle espèce putative de Monorchis. Les données de séquence de liaison avec les évaluations morphologiques détaillées ont aidé à clarifier la délinéation des espèces au sein des complexes de deux groupes, à savoir « crassigula », le complexe d'espèces de Macvicaria et « Monorchis parvus », le complexe d'espèces de Monorchis. Des aperçus morphologiques et moléculaires sur la diversité des espèces de poissons marins sont fournis à la lumière des phylogénies pour les Opecoelidae, Aporocotylidae, Hemiuroidea, et une phylogénie largement construite pour les Digènes. Le grand nombre d'espèces de trématodes récupérées dans les deux poissons hôtes et la découverte de trois nouvelles espèces pour la science donnent à penser que la diversité de ces parasites dans la région, peut-être plus élevé qu’elle ne le connaît actuellement. Les nouvelles données sur les séquences recueillies au cours de la présente étude feront progresser d'autres études sur la diversité, les aires de répartition des hôtes et la répartition de ces parasites importants. Ces résultats mettent en évidence l'importance de l'application des méthodes morphologiques et moléculaires dans l'évaluation de la diversité des parasites en Méditerranée. Mots Clés : Sparidae, Digène, trématode diversité, Méditerranée, phylogénie moléculaire, cox1, ITS1-5.8S-ITS2, 28S rDNA, Bouzedjar, Alger, Annaba i ABSTRACT The present study focuses on the application of an integrative approach to the identification of trematode parasites recovered in two sparid fish species, Diplodus vulgaris (Geoffroy Saint- Hilaire, 1817) and Sparus aurata L. along the Algerian coasts of the Western Mediterranean, an unexplored area regarding fish helminth diversity. A total of 810 fishes (390 D. vulgaris and 420 S. aurata), were examined at three localities (off Bouzedjar, Algiers and Annaba). Profiting from large sample sizes and fruitful collaboration, we have generated a sequence database for the mitochondrial cox1 and nuclear ITS1-5.8S-ITS2 gene cluster and/or partial 28S rDNA sequences for a total of 15 trematode species. Detailed morphological characterisation is provided for these species representing 11 families: Opecoelidae (5 species of 2 genera); Acanthocolpidae, Aephnidiogenidae, Aporocotylidae, Derogenidae, Hemiuridae, Heterophyidae, Lepocreadiidae, Monorchiidae, Strigeidae (one species each), and a single species of the Aspidogastridae (Subclass Aspidogastrea). Three species new to science were described from the studied material: Macvicaria gibsoni, Rima, Marzoug, Pérez-del-Olmo, Kostadinova, Bouderbala & Georgieva, 2017; a new species of Cardicola; and a putative new species of Monorchis. Linking sequence data with detailed morphological assessments helped clarify species delineation within the species complexes of two groups, i.e. the "crassigula" species complex of Macvicaria and "Monorchis parvus" species complex of Monorchis. Morphological and molecular insights into the diversity of marine fish digeneans are provided in the light of phylogenies for the Opecoelidae, Aporocotylidae, Hemiuroidea, and a broadly built phylogeny for the Digenea. The large number of trematode species recovered in the two fish hosts and the finding of three species new to science suggest that the diversity of these parasites in the region may be higher than currently known. The novel sequence data gathered during the present study will advance further studies on the diversity, host ranges and distribution of these important parasites. The present results highlight the importance of the application of morphological and molecular methods in the assessment of parasite diversity in the Mediterranean. Keywords: Sparidae, Digenea, Sparus aurata, Diplodus vulgaris, Mediterranean, molecular phylogeny, cox1, ITS1-5.8S-ITS2, 28S rDNA, Bouzedjar, Algiers, Annaba ii الملخص الهدف الرئيسي من هدا العمل هو استخدام التقنيات الجزئية لتحديد ومعرفة الطفيليات الداخلية في تنائي الجينات ودلك على اثنين من اسماك فصيلة اﻻسبور (.S. aurata L و (D. vulgaris (G, 1847)على طول الساحل الجزائري من منطقة غرب البحر اﻷبيض المتوسط، التي تعتبر منطقة من المناطق الغير المستكشفة في تنوع الديدان الطفيلية ثنائية الجينات. ويحتل هدين النوعين من اﻷسماك موردا بيولوجيا هاما في السلسلة الغذائية البحرية ولسكان المنطقة. تم جمع العينات على مدى عامين(2014-2015) وبمشاركة فعالة تقدر ب 800 من اﻷسماك على التوالي: S. aurata 410 وD. 390 vulgaris في ثﻻثة مناطق مختلفة على طول الساحل الجزائري: في الغرب مناء بوزجار, في الوسط مناء الجزائر و في الشرق مناء عنابة. من خﻻل اﻻستفادة من أحجام العينات الكبيرة والتعاون المثمر، قمنا بتوليد قاعدة بيانات تسلسلية لكتلة الميتوكوندريا Cox1 والنواة النووية ITS1-5.8S-ITS2 و / أو متواليات rDNA 28S الجزئية ل 15 نو ًعا من تنائي الجينات. البيانات المورفولوجية الجديدة المفصلة لهده لﻷنواع التي تمثل 11 عائلة: Opecoelidae (5 أنواع لجنسين) ; Hemiuridae ،Derogenidae، Aporocotylidae ،Aephnidiogenidae ،Acanthocolpidae, Strigeidae ،Monorchiidae ،Heterophyida )نوع واحد لكل منها( ، ونوع واحد من Aspidogastridae )صنف Aspidogastrea(. ان التحاليل النشؤية المستندة على تسلسل الحمض النووي للمناطق التالية: 28S , ITS1-5.8S-ITS والتسلسل الجزئي لجين الميتوكوندريا كوكس1, قدمت بيانات جديدة لﻷنواع التي تنتمي الى اسر جديدة لثنائية الجينات: Macvicaria gibsoni, Rima, Marzoug, Pérez-del-Olmo, Kostadinova, Bouderbala & Georgieva, 2017; نوع جديد من Cardicola، وأنواع جديدة مفترضة من Monorchis. ساعد ربط البيانات المتسلسلة مع التقييمات المورفولوجية التفصيلية في توضيح وتحديد اﻷنواع الى مجموعتين: نوع ال "crassigula" في النوع المعقد Macvicaria ونوع " Monorchis parvus" في النوع المعقد Monorchis. يشير العدد الكبير من أنواع الديدان التي تم احصائها في النوعين المضيفين لﻷسماك التي تم فحصهم وإيجاد ثﻻثة أنواع جديدة للعلم إلى أن تنوع هذه الطفيليات في المنطقة قد يكون أعلى مما هو معروف حاليًا. فإن تسلسل البيانات الجديدة التي تم جمعها خﻻل هذه الدراسة تقدم المزيد من الدراسات على التنوع وتوزيع هذه الطفيليات الهامة. تسلط النتائج الحالية الضوء على أهمية تطبيق الطرق المورفولوجيا والبيانات الجزيئية في تقييم تنوع الطفيليات في البحر المتوسط. الكامات المفتاحية: فصيلة اﻷسبور, تنائي الجينات, Sparus aurata, Diplodus vulgaris ,الجزائر,البحر اﻷبيض المتوسط, ITS1-5.8S-ITS2 ,كوكس 1 ,التك ُّون الجزيئي ,28S rDNA, بوزجار ,الجزائر ,عنابة iii LIST OF FIGURES CHAPTER I Page Fig. 1.1 Distribution map of Sparus aurata L. (the gilthead seabream) Froese, R. 04 and D. Pauly. Editors. 2016. FishBase.www.fishbase.org, version (10/2016). Fig. 1.2 Distribution map
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