Genetic and Phenotypic Patterns of Variabilities in Arenaria Grandiflora L
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Genetic and phenotypic patterns of variabilities in Arenaria grandiflora L. species complex (Caryophyllaceae) : new elements for taxonomy and conservation Marwa Daoud To cite this version: Marwa Daoud. Genetic and phenotypic patterns of variabilities in Arenaria grandiflora L. species complex (Caryophyllaceae) : new elements for taxonomy and conservation. Biodiversity and Ecology. Museum national d’histoire naturelle - MNHN PARIS, 2017. English. NNT : 2017MNHN0013. tel- 01948885 HAL Id: tel-01948885 https://tel.archives-ouvertes.fr/tel-01948885 Submitted on 9 Dec 2018 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. MUSEUM NATIONAL D’HISTOIRE NATURELLE Ecole Doctorale Sciences de la Nature et de l’Homme – ED 227 Année 2017 N° attribué par la bibliothèque |_|_|_|_|_|_|_|_|_|_|_|_| THESE Pour obtenir le grade de DOCTEUR DU MUSEUM NATIONAL D’HISTOIRE NATURELLE Spécialité : Ecologie et Systématique Présentée et soutenue publiquement par Marwa DAOUD Le 08/12/2017 Genetic and phenotypic patterns of variabilities in Arenaria grandiflora L. species complex (Caryophyllaceae) New elements for taxonomy and conservation Sous la direction de : MACHON Nathalie et SILJAK-YAKOVLEV Sonja Composition du jury : Nathalie MACHON Professeur, Muséum National d'Histoire Naturelle Directrice de Thèse Sonja SILJAK-YAKOVLEV Directrice de Recherche, CNRS Directrice de Thèse Magda BOU DAGHER-KHARRAT Professeur, Université Saint-Joseph de Beyrouth Rapporteur Joan VALLES Professeur, Université de Barcelone Rapporteur Errol VELA Maître de Conférences, Université de Montpellier Examinateur I THESE Pour obtenir le grade de DOCTEUR DU MUSEUM NATIONAL D’HISTOIRE NATURELLE Spécialité : Ecologie et Systématique Présentée et soutenue publiquement par Marwa DAOUD Le 08/12 2017 Genetic and Phenotypic Patterns of Variabilities in Arenaria grandiflora L. species complex (Caryophyllaceae) New Elements for Taxonomy and Conservation Sous la direction de : MACHON Nathalie et SILJAK-YAKOVLEV Sonja Composition du jury : Nathalie MACHON Professeur, Muséum National d'Histoire Naturelle Directrice de Thèse Sonja SILJAK-YAKOVLEV Directrice de Recherche, CNRS Directrice de Thèse Magda BOU DAGHER-KHARRAT Professeur, Université Saint-Joseph de Beyrouth Rapporteur Joan VALLES Professeur, Université de Barcelone Rapporteur Errol VELA Maître de Conférences, Université de Montpellier Examinateur II III Dedicated to: My country (Syria) My parents (Hayat and Louay) My husband (Moutasem) And my cute daughter (Ame) My brother (Nebras) My sisters (Mhar and Oshine) . for their unconditional support, their wishes, their fortitude, their understanding, their help and guidance in every walk of life. IV V Acknowledgements Many inspiring people have been involved in the work leading to my PhD thesis. I will like to acknowledge everyone for their contributions to the studies conducted during my time as a PhD student and I am especially thankful to: Josie LAMBOURDIÈRE and Jawad ABDELKRIM for their technical assistance in microsatellites analysis at SSM in the French National Museum of Natural History, Paris, France Chantal GRIVEAU for her assistance in preparations of clones at National Botanical Conservatory of the Paris Basin (CBNBP), Paris, France Mickael BOURGE and Mario GOMEZ PACHECO for their technical assistance in cytometry at IBiSA Cytometry Platform of the Imagif, Gif-sur-Yvette, France Sandrine FONTAINE and Christian RAQUIN, for their both technical assistance and guidance in culture in vitro at ESE in University of Paris-Sud Saclay. Raphaël CORNETTE and Florian JABBOUR, for their both assistance and support in morphometrics analysis, in the French National Museum of Natural History, Paris, France Nélida PADILLA GARCIA, Manuel CARTEREAU, Bernard PASQUIER and Bruno DURAND for their assistance in collection of material Myriam GAUDEUL for her assistance in sampling from Paris’ herbarium in the French National Museum of Natural History, Paris, France Sophie NADOT and Thierry ROBERT for their constructive advices from University of Paris- Sud Saclay, Orsay, France VI Preface Five years ago, I started this project as a thesis project, but it turned out to be long battle against a large- flowered Sandwort from Caryophyllaceae (Arenaria grandiflora L.). This thesis is the report of this long process, but it cannot express the long days spent in both the field and the lab, battling shoulder to shoulder with my fellow scientists. It cannot also express the stress, the sadness and the tiredness because of the war and the victims in my country (Syria). The first year was only just unsuccessful attempts to detect both the nuclear microsatellite markers and the chloroplast markers (we used 4 pairs of universal primers (MatK 390F / 1440R, TrnK-MatK 3914F / 1110R, TrnL-trnF e / f and Trnt-L La / Lb), in dry and old herbarium’ specimens (Paris’ herbarium, Montpellier’ herbarium, Geneva’ herbarium and Barcelona’ herbarium). A new and different scheme to search was established in the second year. This work is to the best of my knowledge original except where references are made to previous work. This thesis has been writing based on experiments conducted in the scientific group of both two laboratories Center for Ecology and Conservation Sciences (CESCO; MNHN) and Ecology, Systematics and Evolution (ESE, CNRS, University of Paris-Sud Saclay). I would like to thank my supervisors Nathalie MACHON and Sonja SILJAK-YAKOVLEV for their excellent guidance and support during this process. It is great to have finally completed my thesis. Within, this interesting subject, I have gained much knowledge and discovered a domain, where I like to work in the future. At the end, thanks to you, reader, if you are reading this line after the others, you at least read one page of my thesis. Thank You VII Summary of the thesis Population-level conservation is being extremely required to restrain the biodiversity loss within a species. So, the assessment of the variability within the species complex is being renowned as an important first step to well implement the future conservation settings for threatened species. The species complex of Arenaria grandiflora is a short-lived perennial herbaceous and a threatened taxon in certain of sites of its distribution areas in Europe, with unresolved genetics and taxonomy, which lead to potential problems in the conservation and utilization of the resource. A differentiation among populations of the species complex of A. grandiflora is presented in this study based on the genetic, cytogenetic and phenotypic patterns. Intraspecific ploidy level variation is an important aspect of numerous species, so, the present study explores this phenomenon within the A. grandiflora species complex in some type of populations (27 natural populations). To infer the intraspecific genetic and cytogenetic patterns of variability among the studied natural populations of the investigated species complex (A. grandiflora), three methods were used: nuclear microsatellite markers, cytogenetic and flow cytometry approaches. Moreover, the phenotypic patterns of variation among both the stock of seeds and the herbarium materials of A. grandiflora were defined. These patterns were detected using three methods of seed germination (in vitro culture, filter papers and potting soil) and morphometric approaches. A significant differentiation among populations’ patterns of molecular, cytogenetic and phenotypic variation was detected within the A. grandiflora species complex. Presence of two closely related cytotypes (diploids 2n=2x=22 and tetraploids 2n=4x=44) was detected using both classical and more recent methods (chromosome number count and flow cytometry respectively). The species complex of A. grandiflora exhibits high variation in 2C-DNA value, the genome size ranges from 2.11±0.74 pg to 2.70±0.11 pg for the diploid populations and from 4.30±1.51 pg to 5.27±0.14 pg for the tetraploid populations. Moreover, the seeds of tetraploids germinate well and in high proportion than the seeds of the diploid ones. In addition, both acicular and linear leaves from the diploid populations differ significantly within the diploids and with the lanceolate leaves of the tetraploid ones. New protocol of seed germination for the tetraploids by in vitro culture after scarifying was described for the first time. The affected factors on seed germination percentages were determined by an explanatory model of six predictors (altitude, longitude, latitude, ploidy levels, both period and condition of seed storage). VIII Consequently, all these findings are fundamental for the determination of the evolutionarily significant units (ESUs) within A. grandiflora species complex and thus the definition of efficient restoration plans in the future. This study would consider as the preliminary signal for necessary revision for the intraspecific taxonomic keys problematic for this species complex. IX Table of Contents 1. Chapter I: General introduction 1.1 Polyploidy in plants – a brief history 1.2 Evolutionarily significant units (ESUs)– a confuse