Subtribe Alopecurinae (Poeae, Poaceae) sensu lato‘s seven genera share interesting morphological similarities (dense spicate panicles and one-flowered spikelets) that were widely thought to have a common origin. However, recent molecular evidence for three of the genera has suggested that the subtribe may be polyphyletic. To test this, five DNA regions were sequenced and analyzed using phylogenetic methods. Results confirm that Alopecurinae s.l. as presently treated is polyphyletic and should be dissolved. Additionally, the genus Cornucopiae may be just another Alopecurus. Limnas and Pseudophleum are not closely allied to Alopecurus or each other, and are even further from Phleum. Phleum is a distinct lineage that is not closely allied to any other included Alopecurinae genus. Evidence for revising infrageneric classifications of Alopecurus and Phleum is presented, as is evidence for separating A. magellanicus into two or more subspecies.

Les coumarines et furocoumarines sont des phytoalexines synthétisées par certaines familles de plantes (ex : Rutacées dont font partie les agrumes), pour se défendre contre les bioagresseurs. Les furocoumarines peuvent être toxiques pour l’homme, lorsqu’elles sont combinées à certains médicaments : c’est l’effet pomelo. Aujourd’hui, la plupart des cytochromes P450 impliqués dans la synthèse des furocoumarines chez les Apiacées, ont déjà été caractérisés. En revanche, malgré l’importance économique des agrumes, nous en savons très peu sur la voie de biosynthèse des coumarines et furocoumarines chez ces plantes. Dans ce travail, nous avons créé, optimisé et validé une méthode d’analyse en chromatographie liquide à ultra haute performance couplée à un spectromètre de masse (UPLC-MS), pour identifier et quantifier 28 coumarines et furocoumarines dans la peau et la pulpe d’agrumes. Cette méthode nous a permis de chémotyper 62 variétés d’agrumes, distinguées par leur faible ou forte capacité de production de ces composés. En parallèle, un travail de bioinformatique sur des banques publiques d’ADN génomique d’agrumes, a permis d’identifier sept gènes présentant de fortes homologies de séquences avec ceux intervenant dans la synthèse des furocoumarines chez Pastinaca sativa (CYP71) et chez Arabidopsis thaliana (CYP82). Une analyse quantitative de leur niveau d’expression chez des agrumes, a montré que quatre d’entre eux étaient plus fortement exprimés chez les fruits fortement producteurs de coumarines et furocoumarines. Le clonage de ces gènes et leur expression hétérologue chez la levure, a révélé la fonction de CYP82D64 de pomelo et de Combava, qui hydroxyle la xanthotoxine pour donner la 5-hydroxy-xanthotoxine. La synthèse des coumarines et furocoumarines chez les agrumes, ainsi mieux appréhendée, nous a permis de proposer un schéma de sélection variétale visant à abaisser les taux de ces composés chez les Citrus. Nous avons aussi montré l’évolution convergente des CYP71 et CYP82 dans leur synthèse chez les Apiacées et les Rutacées respectivement. La découverte du premier cytochrome P450 de Citrus intervenant dans la production de ces composés, ouvre de nouvelles perspectives quant à l’élucidation de leur voie de biosynthèse chez les agrumes / Coumarins and furanocoumarins are phytoalexines synthesized by some plant families (e.g. Rutaceæ that include citrus), to defend themselves against bioaggressors. Furanocoumarins can be toxic for humans, when combined with some drugs: this is the grapefruit juice effect. Nowadays, most of the cytochrome P450s involved in the furanocoumarin synthesis in Apiaceæ, have already been characterized. However, despite the economical importance of citrus, a little is known about the coumarins and furanocoumarins pathway in these plants. In this work, we created, optimized and validated an analytical method by ultra high performance liquid chromatography coupled with mass spectrometry (UPLC-MS), to identify and quantitate 28 coumarins and furanocoumarins in citrus peel and pulp. This method allowed us to chemotype 62 citrus varieties, distinguished by their low or high capacity to produce these compounds. In parallel, a bioinformatic work on public banks of genomic DNA from citrus, allowed to identify seven genes with high sequence homologies with those involved in the synthesis of furanocoumarins in Pastinaca sativa (CYP71) and in Arabidopsis thaliana (CYP82). A quantitative analysis of their expression level in citrus showed that four of them were more expressed in high coumarins and furanocoumarins producing fruits. The cloning of these genes and their heterologous expression in yeast, revealed the function of grapefruit and Combava CYP82D64, which catalyzes the hydroxylation of xanthotoxin in 5-hydroxy-xanthotoxin. The synthesis of coumarins and furanocoumarins in citrus, then better apprehended, allowed us to propose a breeding scheme aiming at decreasing the levels of these compounds in Citrus. We also showed the convergent evolution of CYP71 and CYP82 in their synthesis in Apiaceæ and in Rutaceæ respectively. The discovery of the first cytochrome P450 from Citrus involved in the production of these compounds, opens up new prospects for the elucidation of their biosynthetic pathway in citrus

Subtribe Alopecurinae (Poeae, Poaceae) sensu lato‘s seven genera share interesting morphological similarities (dense spicate panicles and one-flowered spikelets) that were widely thought to have a common origin. However, recent molecular evidence for three of the genera has suggested that the subtribe may be polyphyletic. To test this, five DNA regions were sequenced and analyzed using phylogenetic methods. Results confirm that Alopecurinae s.l. as presently treated is polyphyletic and should be dissolved. Additionally, the genus Cornucopiae may be just another Alopecurus. Limnas and Pseudophleum are not closely allied to Alopecurus or each other, and are even further from Phleum. Phleum is a distinct lineage that is not closely allied to any other included Alopecurinae genus. Evidence for revising infrageneric classifications of Alopecurus and Phleum is presented, as is evidence for separating A. magellanicus into two or more subspecies.

Les interactions intimes et répétées entre hôtes et parasites peuvent engendrer la spécialisation d’un parasite à son hôte, grâce à des adaptations comportementales, morphologiques et/ou génétiques, combinées avec un flux de gènes limité. C’est un processus clef car il participe à l’évolution de la biodiversité parasitaire et peut ainsi permettre de mieux comprendre l’émergence d’organismes pathogènes. Encore peu étudié, une spécialisation d’hôtes a néanmoins été démontré lors de précédentes études chez deux espèces de tiques nidicoles : chez Ixodes uriae une tique dure, parasite des oiseaux marins coloniaux en zone arctique, et dans un complexe de tiques molles Ornithodoros capensis sensu lato, parasitant aussi de nombreuses espèces d’oiseaux marins, mais cette fois-ci en zones tempérées et tropicales. Ces deux espèces sont vectrices d’une grande diversité d’agents pathogènes incluant des virus, des bactéries et des protozoaires. Cependant, les facteurs impliqués dans le phénomène de spécialisation d’hôte restent inconnus. Dans ce cadre, le but de ma thèse était donc de déterminer 1) si l’évolution des divergences en fonction des hôtes est toujours accompagnée par les mêmes changements phénotypiques et 2) si ces changements pourraient permettre d’identifier les facteurs de sélection sous-jacents. Dans ce contexte, des campagnes d’échantillonnage de tiques ont été menées durant la période de reproduction des hôtes oiseaux dans les différentes zones de leur répartition et nous avons réalisé des analyses morphométriques, basées sur l’utilisation de landmarks et de contours sur chaque individu tique et des analyses phylogénétiques et génétiques des populations sur les mêmes individus. L’ensemble de ces résultats suggère la présence de convergences morphologiques au sein de ces systèmes et souligne un rôle de la sélection dans ce processus de divergence. En effet, les caractéristiques écologiques des hôtes mais aussi le micro-habitat exercent des pressions sélectives importantes dans ces deux systèmes pouvant être à l’origine de la divergence observée entre les populations. De plus, les caractéristiques biologiques de chaque espèce de tiques, telle que la capacité de dispersion, entrent également en jeu et peuvent fortement modifier l’épidémiologie des agents infectieux dont elles sont vectrices.Mots clés : Argasidae, écologie de la transmission, évolution convergente, interactions hôte-parasite, Ixodidae, oiseaux marins. / Intimate and repeated interactions between hosts and parasites can lead to parasite specialization to a given host via behavioral, morphological and/or genetic adaptations that act in combination with restricted gene flow. Specialization is a key process leading to the generation of parasite biodiversity and can help us understand the emergence of pathogenic organisms. Although little studied, host specialization has already been demonstrated to occur in previous studies of two nidicolous tick species: Ixodes uriae a hard tick parasitizing colonial seabirds in polar regions, and soft ticks of the complex Ornithodoros capensis sensu lato, that also exploit colonial seabirds, but this time in temperate and tropical zones. Both of these species act as vector to a wide variety of pathogenic organisms, including viruses, bacteria and protozoa. However, the factors involved in host specialization remain unknown. In this context, the aim of my thesis was to determine 1) whether the evolution of host specialization is always accompanied by the same phenotypic changes and 2) whether these changes could help to identify the selective factors that influence this phenomenon. In this context, tick collections were conducted during the breeding period of the host birds in different areas of their distribution and morphometric analyses, based on landmark and contour methods, were performed on each individual tick. Phylogenetic and population genetic analyses were also carried out using the same individuals. Overall, the results demonstrate that morphological convergence occurs within these systems, highlighting the role of selection in the divergence process. Indeed, the ecological characteristics of the hosts, but also their micro-habitat, may exert significant selective pressures on ticks and may cause the observed divergence among populations. Likewise, the biological characteristics of each tick species, particularly in relation to dispersal capacity, may also come into play and will greatly modify the epidemiology of associated infectious agents.Keywords: Argasidae, convergent evolution, host-parasite interactions, Ixodidae, transmission ecology, seabirds.

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Background: Heat-shock proteins of the 70 kDa family (Hsp70s) are essential chaperones required for key cellular functions. In eukaryotes, four subfamilies can be distinguished according to their function and localisation in different cellular compartments: cytosol, endoplasmic reticulum, mitochondria and chloroplasts. Generally, multiple cytosol-type Hsp70s can be found in metazoans that show either constitutive expression and/or stress-inducibility, arguing for the evolution of different tasks and functions. Information about the hsp70 copy number and diversity in microbial eukaryotes is, however, scarce, and detailed knowledge about the differential gene expression in most protists is lacking. Therefore, we have characterised the Hsp70 gene family of Paramecium caudatum to gain insight into the evolution and differential heat stress response of the distinct family members in protists and to investigate the diversification of eukaryotic hsp70s focusing on the evolution of heat-inducibility. Results: Eleven putative hsp70 genes could be detected in P. caudatum comprising homologs of three major Hsp70-subfamilies. Phylogenetic analyses revealed five evolutionarily distinct Hsp70-groups, each with a closer relationship to orthologous sequences of Paramecium tetraurelia than to another P. caudatum Hsp70-group. These highly diverse, paralogous groups resulted from duplications preceding Paramecium speciation, underwent divergent evolution and were subject to purifying selection. Heat-shock treatments were performed to test for differential expression patterns among the five Hsp70-groups as well as for a functional conservation within Paramecium. These treatments induced exceptionally high mRNA up-regulations in one cytosolic group with a low basal expression, indicative for the major heat inducible hsp70s. All other groups showed comparatively high basal expression levels and moderate heat-inducibility, signifying constitutively expressed genes. Comparative EST analyses for P. tetraurelia hsp70s unveiled a corresponding expression pattern, which supports a functionally conserved evolution of the Hsp70 gene family in Paramecium. Conclusions: Our analyses suggest an independent evolution of the heat-inducible cytosol-type hsp70s in Paramecium and in its close relative Tetrahymena, as well as within higher eukaryotes. This result indicates convergent evolution during hsp70 subfunctionalization and implies that heat-inducibility evolved several times during the course of eukaryotic evolution.

Background: Heat-shock proteins of the 70 kDa family (Hsp70s) are essential chaperones required for key cellular functions. In eukaryotes, four subfamilies can be distinguished according to their function and localisation in different cellular compartments: cytosol, endoplasmic reticulum, mitochondria and chloroplasts. Generally, multiple cytosol-type Hsp70s can be found in metazoans that show either constitutive expression and/or stress-inducibility, arguing for the evolution of different tasks and functions. Information about the hsp70 copy number and diversity in microbial eukaryotes is, however, scarce, and detailed knowledge about the differential gene expression in most protists is lacking. Therefore, we have characterised the Hsp70 gene family of Paramecium caudatum to gain insight into the evolution and differential heat stress response of the distinct family members in protists and to investigate the diversification of eukaryotic hsp70s focusing on the evolution of heat-inducibility. Results: Eleven putative hsp70 genes could be detected in P. caudatum comprising homologs of three major Hsp70-subfamilies. Phylogenetic analyses revealed five evolutionarily distinct Hsp70-groups, each with a closer relationship to orthologous sequences of Paramecium tetraurelia than to another P. caudatum Hsp70-group. These highly diverse, paralogous groups resulted from duplications preceding Paramecium speciation, underwent divergent evolution and were subject to purifying selection. Heat-shock treatments were performed to test for differential expression patterns among the five Hsp70-groups as well as for a functional conservation within Paramecium. These treatments induced exceptionally high mRNA up-regulations in one cytosolic group with a low basal expression, indicative for the major heat inducible hsp70s. All other groups showed comparatively high basal expression levels and moderate heat-inducibility, signifying constitutively expressed genes. Comparative EST analyses for P. tetraurelia hsp70s unveiled a corresponding expression pattern, which supports a functionally conserved evolution of the Hsp70 gene family in Paramecium. Conclusions: Our analyses suggest an independent evolution of the heat-inducible cytosol-type hsp70s in Paramecium and in its close relative Tetrahymena, as well as within higher eukaryotes. This result indicates convergent evolution during hsp70 subfunctionalization and implies that heat-inducibility evolved several times during the course of eukaryotic evolution.