Recombination is suppressed and variability reduced in a nascent Y chromosome.

Several hypotheses have been elaborated to account for the evolutionary decay commonly observed in full-fledged Y chromosomes. Enhanced drift, background selection and selective sweeps, which are expected to result from reduced recombination, may all share responsibilities in the initial decay of proto-Y chromosomes, but little empirical information has been gathered so far. Here we take advantage of three markers that amplify on both of the morphologically undifferentiated sex chromosomes of the European tree frog (Hyla arborea) to show that recombination is suppressed in males (the heterogametic sex) but not in females. Accordingly, genetic variability is reduced on the Y, but in a way that can be accounted for by merely the number of chromosome copies per breeding pair, without the need to invoke background selection or selective sweeps.

ADME pharmacogenetics: investigation of the pharmacokinetics of the antiretroviral agent lopinavir coformulated with ritonavir.

BACKGROUND: An ADME (absorption, distribution, metabolism and excretion)-pharmacogenetics association study may identify functional variants relevant to the pharmacokinetics of lopinavir co-formulated with ritonavir (LPV/r), a first-line anti-HIV agent. METHODS: An extensive search of literature and web resources helped select ADME genes and single nucleotide polymorphisms (SNPs, functional and HapMap tagging SNPs) with a proven or potentially relevant role in LPV/r pharmacokinetics. The study followed a two-stage design. Stage 1 (discovery) considered a Caucasian population (n=638) receiving LPV/r, where we selected 117 individuals with low LPV clearance (cases) and 90 individuals with high clearance (controls). Genotyping was performed by a 1536-SNP customized GoldenGate Illumina BeadArray. Stage 2 (confirmation) represented a replication study of candidate SNPs from the stage 1 in 148 individuals receiving LPV/r. The analysis led to formal population pharmacokinetic-pharmacogenetic modeling of demographic, environmental and candidate SNP effects. RESULTS: One thousand three hundred and eighty SNPs were successfully genotyped. Nine SNPs prioritized by the stage 1 analysis were brought to replication. Stage 2 confirmed the contribution of two functional SNPs in SLCO1B1, one functional SNP in ABCC2 and a tag SNP of the CYP3A locus in addition to body weight effect and ritonavir coadministration. According to the population pharmacokinetic-pharmacogenetic model, genetic variants explained 5% of LPV variability. Individuals homozygous rs11045819 (SLCO1B1*4) had a clearance of 12.6 l/h, compared with 5.4 l/h in the reference group, and 3.9 l/h in individuals with two or more variant alleles of rs4149056 (SLCO1B1*5), rs717620 (ABCC2) or rs6945984 (CYP3A). A subanalysis confirmed that although a significant part of the variance in LPV clearance was attributed to fluctuation in ritonavir levels, genetic variants had an additional effect on LPV clearance. CONCLUSION: The two-stage strategy successfully identified genetic variants affecting LPV/r pharmacokinetics. Such a general approach of ADME pharmacogenetics should be generalized to other drugs.

Chromosomal evolution and zoogeographic origin of southeast Asian shrews (genus Crocidura).

To the origins and evolution of Indomalayan shrews, we investigated the chromosomal variations of 14 species of Crocidura from SE Asia. Intraspecific polymorphism was mainly due to variation in the number of short chromosomal arms but C. lepidura and C. hutanis showed a polymorphism due to a centric fusion. The undifferentially stained karyotypes were similar in 9 species, all possessing 2n = 38 and FN = 54-56 (68); C. fuliginosa had 2n = 40 and FN = 54-58. These karyotypes are close to the presumed ancestral state for the genus Crocidura. Four species from Sulawesi had a reduced diploid number (2n = 30-34), a trend not observed among other SE Asian species but present in few Palaearctic taxa. Compared to the apparent stasis of karyotypic evolution observed among other SE Asian species, the high degree of interspecific differences reported among Sulawesian shrews is unusual and needs further investigation. Stasis and reduction in diploid number found in both Indomalayan and Palaeractic species suggest that these two groups share a common ancestry. This is in sharp contrast to most Afrotropical species which evolved towards higher diploid and fundamental numbers. The zoogeographical implications of these results are discussed.

Changes in reproductive investment with altitude in an alpine plant

Aims: In perennial species, the allocation of resources to reproduction results in a reduction of allocation to vegetative growth and, therefore, impacts future reproductive success. As a consequence, variation in this trade-off is among the most important driving forces in the life-history evolution of perennial plants and can lead to locally adapted genotypes. In addition to genetic variation, phenotypic plasticity might also contribute to local adaptation of plants to local conditions by mediating changes in reproductive allocation. Knowledge on the importance of genetic and environmental effects on the trade-off between reproduction and vegetative growth is therefore essential to understand how plants may respond to environmental changes. Methods: We conducted a transplant experiment along an altitudinal gradient from 425 m to 1921 m in the front range of the Western Alps of Switzerland to assess the influence of both altitudinal origin of populations and altitude of growing site on growth, reproductive investment and local adaptation in Poa alpina. Important findings: In our study, the investment in reproduction increased with plant size. Plant growth and the relative importance of reproductive investment decreased in populations originating from higher altitudes compared to populations originating from lower altitudes. The changes in reproductive investment were mainly explained by differences in plant size. In contrast to genetic effects, phenotypic plasticity of all traits measured was low and not related to altitude. As a result, the population from the lowest altitude of origin performed best at all sites. Our results indicate that in P. alpina genetic differences in growth and reproductive investment are related to local conditions affecting growth, i.e. interspecific competition and soil moisture content.

Inbreeding and selection on sex ratio in the bark beetle Xylosandrus germanus.

ABSTRACT: BACKGROUND: Local Mate Competition (LMC) theory predicts a female should produce a more female-biased sex ratio if her sons compete with each other for mates. Because it provides quantitative predictions that can be experimentally tested, LMC is a textbook example of the predictive power of evolutionary theory. A limitation of many earlier studies in the field is that the population structure and mating system of the studied species are often estimated only indirectly. Here we use microsatellites to characterize the levels of inbreeding of the bark beetle Xylosandrus germanus, a species where the level of LMC is expected to be high. RESULTS: For three populations studied, genetic variation for our genetic markers was very low, indicative of an extremely high level of inbreeding (FIS = 0.88). There was also strong linkage disequilibrium between microsatellite loci and a very strong genetic differentiation between populations. The data suggest that matings among non-siblings are very rare (3%), although sex ratios from X. germanus in both the field and the laboratory have suggested more matings between non-sibs, and so less intense LMC. CONCLUSIONS: Our results confirm that caution is needed when inferring mating systems from sex ratio data, especially when a lack of biological detail means the use of overly simple forms of the model of interest.

Molecular evidence for ancient asexuality in timema stick insects.

Asexuality is rare in animals in spite of its apparent advantage relative to sexual reproduction, indicating that it must be associated with profound costs [1-9]. One expectation is that reproductive advantages gained by new asexual lineages will be quickly eroded over time [3, 5-7]. Ancient asexual taxa that have evolved and adapted without sex would be "scandalous" exceptions to this rule, but it is often difficult to exclude the possibility that putative asexuals deploy some form of "cryptic" sex, or have abandoned sex more recently than estimated from divergence times to sexual relatives [10]. Here we provide evidence, from high intraspecific divergence of mitochondrial sequence and nuclear allele divergence patterns, that several independently derived Timema stick-insect lineages have persisted without recombination for more than a million generations. Nuclear alleles in the asexual lineages displayed significantly higher intraindividual divergences than in related sexual species. In addition, within two asexuals, nuclear allele phylogenies suggested the presence of two clades, with sequences from the same individual appearing in both clades. These data strongly support ancient asexuality in Timema and validate the genus as an exceptional opportunity to attack the question of how asexual reproduction can be maintained over long periods of evolutionary time.

Pharmacogenetics of Antiviral Treatment

RESUME - FRANÇAISRésuméDans ce travail de thèse, l'importance de la pharmacogénétique des traitements antiviraux a été évaluée en déterminant, au moyen de trois différentes approches, l'impact de variations génétiques sur la pharmacocinétique de deux traitements antirétroviraux (à savoir l'efavirenz et le lopinavir) ainsi que sur la capacité de pouvoir éliminer le virus de l'hépatite C de façon naturelle ou suite à un traitement médicamenteux.L'influence des variations génétiques sur les taux plasmatiques de l'efavirenz et de ses métabolites primaires a été évaluée par l'analyse d'un seul gène candidat : le cytochrome P450 (CYP) 2A6, impliqué dans une voie métabolique accessoire de l'efavirenz. Cette étude a permis de démontrer que le génotype du CYP2A6 devient cliniquement déterminant en l'absence de fonction du CYP2B6, impliqué dans la voie métabolique principale, et que la perte simultanée des voies métaboliques principales et accessoires entraine une augmen¬tation du risque d'interruption du traitement, soulignant la valeur prédictive du génotypage.L'influence de la génétique sur la clairance du lopinavir a été évaluée par l'analyse à grande échelle de gènes candidats, à savoir les gènes potentiellement impliqués dans l'absorption, le métabolisme, la distribution et l'élimination d'un médicament. Cette étude a permis l'identification de 4 polymorphismes, dans des transporteurs et des enzymes métaboliques, associés à la clairance du lopinavir et expliquant 5% de la variabilité inter¬individuelle de ce phénotype.L'influence de la génétique sur la capacité d'éliminer le virus de l'hépatite C, de façon naturelle ou à la suite d'un traitement, a été évaluée par l'analyse du génome entier. Cette étude a permis l'identification d'un polymorphisme situé à proximité de l'interféron-X3. Quatre variations génétiques potentiellement causales ont ensuite pu être identifiées par reséquencage. Finalement, la contribution nette de ce gène sur l'élimination du virus a pu être évaluée dans une cohorte infectée par une seule et même source, permettant ainsi de contrôler l'effet de la diversité virale, du genre et de la présence de co-infections.Cette thèse a permis de mettre en évidence les diverses méthodes disponibles pour la recherche en pharmacogénétique, ainsi que l'importance du reséquencage pour l'identification de variations génétiques causales.SUMMARY - ENGLISHSummaryIn this thesis work the relevance of pharmacogenetics of antiviral treatment has been assessed by investigating, through three different approaches, the impact of host genetic variation on antiretroviral drug disposition (namely efavirenz and lopinavir) and on natural or treatment-induced clearance of hepatitis C virus.The influence of host genetic variation on efavirenz and its primary metabolite plasma levels was assessed by single candidate gene approach, through comprehensive analysis of cytochrome P450 (CYP) 2A6 - involved in efavirenz accessory metabolic pathway. The study could demonstrate that CYP2A6 genotype became increasingly relevant in the setting of limited CYP2B6 function - involved in efavirenz main metabolic pathway - and that individuals with both main and accessory metabolic pathways impaired were at higher risk for treatment discontinuation, overall emphasizing the predictive power of genotyping.The influence of host genetic variation on lopinavir clearance was assessed by large scale candidate gene approach, through analysis of genes involved in the absorption, distribution, metabolism and elimination. The study identified four genetic variants in drug transporters and metabolizing enzymes that explained 5% of the interindividual variability in lopinavir clearance.The influence of host genetic variation on hepatitis C virus (HCV) natural or treatment- induced clearance was assessed through genome-wide association study approach. This study identified an intergenic polymorphism, part of a linkage disequilibrium block encompassing the interferon-3 gene, as highly associated with treatment-induced and spontaneous HCV clearance. Resequencing and recombinant mapping lead to the identification of four potentially causal genetic variants. Finally, we could assess the net contribution of genetic variants in interferon-3 to clearance by controlling for viral diversity, gender and co-infection status in a single source infected cohort.This thesis highlights the various genetic tools available to pharmacogenetic discovery (candidate gene, pathway or and genome-wide approaches), and the importance of resequencing for mapping of causal variants.

Demogenetics in metapopulations

SUMMARY : Human-induced habitat fragmentation constitutes a major threat to biodiversity. Small and isolated populations suffer from increased stochasticity and from limited rescue effects. These two factors may be sufficient to cause local extinctions but fragmentation induces some genetic consequences that can also contribute significantly to extinction risks. Increased genetic drift reduces the effectiveness of selection against deleterious mutations, leading to their progressive accumulation. Drift also decreases both the standing genetic variation and the rate of fixation of beneficial mutations, limiting the evolutionary potential of isolated populations. Demography and genetics further interact and feed back on each other, progressively driving fragmented populations into "extinction vortices". The aim of the thesis was to better understand the processes occurring in fragmented populations. For this, I combined simulation studies and empirical data from three species that live in structured habitats. Chapter 1 and 2 investigate the demography of two shrew species in fragmented habitats. I showed that connectivity and habitat quality strongly affect the demography of the greater white-tooted shrew, although demographic stochasticity was extremely high. I also demonstrated that habitat fragmentation is one of the leading factors allowing the local coexistence of two competing shrew species. Chapter 3 and 4 focus on measuring connectivity in fragmented populations based on genetic data. In particular, I showed that genetic data can be used to detect the landscape elements impeding dispersal. In Chapter 5 that deals with the accumulation of deleterious mutations in fragmented populations, I demonstrated that mutation accumulation, as well a time to extinction, can be predicted from simple demographic and genetic measures. In the last two chapters, I monitored individual reproductive success in an isolated tree frogs population. These data allowed quantifying the effective population size, a measure closely linked to population evolutionary potential. To conclude, this thesis brings some new insights into the processes occurring in fragmented populations, and I hope it will contribute to the improvement of the management and conservation of fragmented populations.

Local adaptation and life history traits evolution in heterogeneous environment : an empirical example in the freshwater snail, Galba truncatula

Abstract: To understand the processes of evolution, biologists are interested in the ability of a population to respond to natural or artificial selection. The amount of genetic variation is often viewed as the main factor allowing a species to answer to selection. Many theories have thus focused on the maintenance of genetic variability. Ecologists and population geneticists have long-suspected that the structure of the environment is connected to the maintenance of diversity. Theorists have shown that diversity can be permanently and stably maintained in temporal and spatial varying environment in certain conditions. Moreover, varying environments have been also theoretically demonstrated to cause the evolution of divergent life history strategies in the different niches constituting the environment. Although there is a huge number of theoretical studies selection and on life history evolution in heterogeneous environments, there is a clear lack of empirical studies. The purpose of this thesis was to. empirically study the evolutionary consequences of a heterogeneous environment in a freshwater snail Galba truncatula. Indeed, G. truncatula lives in two habitat types according the water availability. First, it can be found in streams or ponds which never completely dry out: a permanent habitat. Second, G. truncatula can be found in pools that freeze during winter and dry during summer: a temporary habitat. Using a common garden approach, we empirically demonstrated local adaptation of G. truncatula to temporary and permanent habitats. We used at first a comparison of molecular (FST) vs. quantitative (QST) genetic differentiation between temporary and permanent habitats. To confirm the pattern QST> FST between habitats suggesting local adaptation, we then tested the desiccation resistance of individuals from temporary and permanent habitats. This study confirmed that drought resistance seemed to be the main factor selected between habitats, and life history traits linked to the desiccation resistance were thus found divergent between habitats. However, despite this evidence of selection acting on mean values of traits between habitats, drift was suggested to be the main factor responsible of variation in variances-covariances between populations. At last, we found life history traits variation of individuals in a heterogeneous environment varying in parasite prevalence. This thesis empirically demonstrated the importance of heterogeneous environments in local adaptation and life history evolution and suggested that more experimental studies are needed to investigate this topic. Résumé: Les biologistes se sont depuis toujours intéressés en l'aptitude d'une population à répondre à la sélection naturelle. Cette réponse dépend de la quantité de variabilité génétique présente dans cette population. Plus particulièrement, les théoriciens se sont penchés sur la question du maintient de la variabilité génétique au sein d'environnements hétérogènes. Ils ont alors démontré que, sous certaines conditions, la diversité génétique peut se maintenir de manière stable et permanente dans des environnements variant au niveau spatial et temporel. De plus, ces environments variables ont été démontrés comme responsable de divergence de traits d'histoire de vie au sein des différentes niches constituant l'environnement. Cependant, malgré ce nombre important d'études théoriques portant sur la sélection et l'évolution des traits d'histoire de vie en environnement hétérogène, les études empiriques sont plus rares. Le but de cette thèse était donc d'étudier les conséquences évolutives d'un environnement hétérogène chez un esgarcot d'eau douce Galba truncatula. En effet, G. truncatula est trouvé dans deux types d'habitats qui diffèrent par leur niveau d'eau. Le premier, l'habitat temporaire, est constitué de flaques d'eau qui peuvent s'assécher pendant l'été et geler pendant l'hiver. Le second, l'habitat permanent, correspond à des marres ou à des ruisseaux qui ont un niveau d'eau constant durant toute l'année. Utilisant une approche expérimentale de type "jardin commun", nous avons démontré l'adaptation locale des individus à leur type d'habitat, permanent ou temporaire. Nous avons utilisé l'approche Fsr/QsT qui compare la différentiation génétique moléculaire avec la différentiation génétique quantitative entre les 2 habitats. Le phénomène d'adapation locale démontré par QsT > FsT, a été testé experimentalement en mesurant la résistance à la dessiccation d'individus d'habitat temporaire et permanent. Cette étude confirma que la résistance à la sécheresse a été sélectionné entre habitats et que les traits responsables de cette resistance sont différents entre habitats. Cependant si la sélection agit sur la valeur moyenne des traits entre habitats, la dérive génétique semble être le responsable majeur de la différence de variances-covariances entre populations. Pour finir, une variation de traits d'histoire de vie a été trouvée au sein d'un environnement hétérogène constitué de populations variants au niveau de leur taux de parasitisme. Pour conclure, cette thèse a donc démontré l'importance d'un environnement hétérogène sur l'adaptation locale et l'évolution des traits d'histoire de vie et suggère que plus d'études empiriques sur le sujet sont nécessaires.

Evidence of recombination in putative ancient asexuals.

Ancient asexuals have been considered to be a contradiction of the basic tenets of evolutionary theory. Barred from rearranging genetic variation by recombination, their reduced number of gene arrangements is thought to hamper their response to changing environments. For the same reason, it should be difficult for them to avoid the build-up of deleterious mutations. Several groups of taxonomically diverse organisms are thought to be ancient asexuals, although clear evidence for or against the existence of recombination events is scarce. Several methods have recently been developed for predicting recombination events by analyzing aligned sequences of a given region of DNA that all originate from one species. The methods are based on phylogenetic, substitution, and compatibility analyses. Here we present the results of analyses of sequence data from different loci studied in several groups of evolutionarily distant species that are considered to be ancient asexuals, using seven different types of analysis. The groups of organisms were the arbuscular mycorrhizal fungi (Glomales), Darwinula stevensoni (Darwinuloidea crustacean ostracods) and the bdelloid rotifers (Bdelloidea), which are thought to have been asexual for the last 400, 25-100, and 35-40 Myr, respectively. The seven different analytical methods evaluated the evolutionary relationships among haplotypes, and these methods had previously been shown to be reliable for predicting the occurrence of recombination events. Despite the different degree of genetic variation among the different groups of organisms, at least some evidence for recombination was found in all species groups. In particular, predictions of recombination events in the arbuscular mycorrhizal fungi were frequent. Predictions of recombination were also found for sequence data that have previously been used to infer the absence of recombination in bdelloid rotifers. Although our results have to be taken with some caution because they could signal very ancient recombination events or possibly other genetic variation of nonrecombinant origin, they suggest that some cryptic recombination events may exist in these organisms.

Variability of the nucleolar organizer activity in human lymphocytes via Ag-staining.

Chromosomes with Ag staining that varies from one metaphase to the other can be distinguished from those with an Ag-staining that is the same in all metaphases. The intercellular variation of an Ag-NOR can be attributed to many different factors. Whatever the importance of technical factors, they do not seem to account for the large variations in Ag-staining which were observed for each ac. This suggests the existence of a natural intercellular variability of the NOR's activity. The variation of the Ag-stainability of a given NOR, the diversity of Ag-stainings observed on the ten ac of one individual and the differences that exist between individuals raise the question of the existence of a compensation of activity between nucleolar organizers. The study, for each individual, of the mean sum of staining per metaphase reveals that this value is not absolutely constant from one individual to another; in the carriers of Robertsonian fusions it is smaller than in chromosomally normal individuals. The analysis of the transmission shows that inactive NORs remain inactive and that active NORs present a variation in the activity from one generation to the next.

Insights into the biogeographical history of the Lower Guinea Forest Domain: evidence for the role of refugia in the intraspecific differentiation of Aucoumea klaineana.

Determining the biogeographical histories of rainforests is central to our understanding of the present distribution of tropical biodiversity. Ice age fragmentation of central African rainforests strongly influenced species distributions. Elevated areas characterized by higher species richness and endemism have been postulated to be Pleistocene forest refugia. However, it is often difficult to separate the effects of history and of present-day ecological conditions on diversity patterns at the interspecific level. Intraspecific genetic variation could yield new insights into history, because refugia hypotheses predict patterns not expected on the basis of contemporary environmental dynamics. Here, we test geographically explicit hypotheses of vicariance associated with the presence of putative refugia and provide clues about their location. We intensively sampled populations of Aucoumea klaineana, a forest tree sensitive to forest fragmentation, throughout its geographical range. Characterizing variation at 10 nuclear microsatellite loci, we were able to obtain phylogeographic data of unprecedented detail for this region. Using Bayesian clustering approaches, we demonstrated the presence of four differentiated genetic units. Their distribution matched that of forest refugia postulated from patterns of species richness and endemism. Our data also show differences in diversity dynamics at leading and trailing edges of the species' shifting distribution. Our results confirm predictions based on refugia hypotheses and cannot be explained on the basis of present-day ecological conditions.

A female melanin ornament signals offspring fluctuating asymmetry in the barn owl.

Sexual selection theory predicts that males advertise quality by displaying extravagant ornaments. By contrast, whether phenotypic variation in females has a signalling function remains an open question. Here, to our knowledge, we provide the first evidence that a female plumage trait can signal fluctuating asymmetry in the offspring. We experimentally demonstrate in wild barn owls (Tyto alba) that the extent to which females display black spots on their plumage does not only signal offspring parasite resistance as shown in a previous study but also developmental homeostasis in the offspring. A greater number of spotted females produced offspring that had more symmetrical feathers during the period of growth. Males, that pair non-randomly with respect to female plumage spottiness therefore appear to gain substantial benefits by mating with heavily spotted females. Genetic variation in plumage spottiness is nevertheless maintained as the covariation between offspring body mass and mother plumage spottiness varies annually depending on environmental conditions.

Phylogéographie des principales vipères européennes (Vipera ammodytes, V. aspis et V. berus), structuration génétique et multipaternité chez Vipera berus

Résumé: Les vipères du genre Vipera sont des serpents venimeux distribués dans la totalité du Paléarctique. Malgré cette répartition considérable, elles sont extrêmement menacées, leur déclin étant principalement dû à la destruction et à la fragmentation de leur habitat ainsi qu'à la persécution humaine. Afin d'apporter de nouveaux éléments dans le contexte de la protection de ce groupe de reptiles, nous avons utilisé durant ce travail de thèse différents marqueurs moléculaires pour étudier la structuration génétique à petite et à large échelle chez trois espèces appartenant au genre Vipera. La première étude, une phylogéographie moléculaire de la vipère ammodytes (Vipera ammodytes), a montré dans l'ensemble de l'aire de répartition une forte structuration génétique provenant d'isolements antérieures au Pléistocène. La présence d'un nombre important de clades dans le centre des Balkans suggère que cette région a fourni de nombreux refuges isolés durant les glaciations. Ces dernières ont également eu un impact considérable sur la diversité génétique au sein de la majorité des clades, suite à d'importants goulots d'étranglement durant le Pléistocène. L'étude de la phylogéographie de la vipère aspic (Vipera aspis) a montré une différenciation génétique entre les populations présentes de chaque côté des Alpes, mais également une forte structuration interne avec la mise en évidence d'un refuge en France. Cette étude est la première à établir clairement l'utilisation d'un refuge français pour un vertébré terrestre. La troisième partie de cette thèse a étudié la phylogéographie de la vipère péliade (Vipera berus), espèce cible de ce travail. En plus de la mise en évidence d'un groupe génétique inattendu (localisé dans le nord de l'Italie, le sud de l'Autriche, le nord de la Slovénie et l'extrême sud-est de la Suisse), la variabilité génétique au sein du groupe nordique (comprenant les animaux de l'entier de l'aire de répartition de l'espèce à l'exception des individus du groupe italien et les animaux provenant des Balkans) est suffisamment importante pour conclure à l'utilisation de refuges glaciaires nordiques durant les dernières glaciations, en complément des refuges habituellement décrits pour la majorité des espèces animales (soit les péninsules ibérique, italienne et balakanique). Ces résultats nous ont conduit à effectuer une étude morphologique (quatrième partie) comparant les vipères péliades du "clade italien" et du "clade nordique" décrits ci-dessus. Seules de petites différences morphologiques ont pu être mises en évidence, malgré une séparation de ces groupes estimée à plus d'un million d'années. Une étude à plus petite échelle, centrée sur le Massif jurassien et certaines populations alpines et françaises, a été entreprise afin d'estimer leur diversité génétique et d'évaluer la structuration génétique entre les populations à l'aide de marqueurs microsatellites (cinquième partie). Une importante structuration a été observée entre les populations distantes de plus de 3 kilomètres, la structuration entre les populations plus proches étant plus limitée. De plus, une diversité génétique plus faible dans les populations jurassiennes et alpines comparativement aux populations du massif central et de la côte atlantique a été constatée, probablement due à une perte de diversité génétique lors de la recolonisation post-glaciaire. La sixième étude s'est intéressée au succès reproducteur des mâles de vipères péliades en conditions naturelles. Une corrélation entre la taille des mâles et leur succès reproducteur a été relevée, les individus de plus grande taille ayant un succès reproducteur plus élevé. Le taux de multipaternité a aussi été investigué, démontrant que la proportion de pontes issues de plusieurs pères est élevée (69%) malgré la faible densité de vipères observée sur le site étudié. Finalement, aucun lien entre le nombre de pères au sein d'une ponte et la mortalité des jeunes à la naissance n'a pu être mis en évidence, contrastant avec des travaux précédents. En conclusion, l'observation de la structuration très marquée chez les vipères péliades devrait permettre d'affiner les méthodes de protection de l'espèce dans le massif jurassien. A plus large échelle, l'importante structuration génétique observée chez les vipères ammodytes, aspic et péliade résultant de l'utilisation de nombreux refuges glaciaires, complémentaires aux refuges habituellement utilisés par les espèces animales, démontre l'intérêt de l'analyse phylogéographique des reptiles pour la compréhension des phénomènes de colonisation et d' extinction des populations durant la fin du Tertiaire et le Quaternaire. La mise en évidence chez les différentes espèces de vipères étudiées de nombreux groupes génétiques distincts (ESUs) devrait conduire à des modifications de la taxonomie ainsi qu'au statut de protection de ces espèces. Abstract: The vipers of the genus Vipera are venomous snakes widespread throughout the Palaearctic regions. Despite a large distribution area, several species are extremely threatened, especially due to the destruction and fragmentation of their habitats, as well as by human persecution. In order to increase the knowledge on these species and to improve their protection, several molecular markers have been used to investigate the genetic structure on small and large scales, within three species of the genus Vipera. The first study, a molecular phylogeography of the nose-horned viper (Vipera ammodytes), showed a considerable structuring throughout the distribution area, due to isolation into refugia before the Pleistocene. A high number of clades in the centre of the Balkans suggests that this region harboured numerous isolated glacial refugia during the last glaciation. Moreover, low genetic diversity within several clades implies that most populations of nose-horned vipers have suffered bottlenecks during the Pleistocene. The study of the phylogeography of the asp viper (Vipera aspis) showed genetic differentiation between populations on each side of the Alps, as well as considerable internal genetic structure, suggesting the use of a glacial refugium in France. This study is the first to establish firmly the occurrence of a French refugia for a terrestrial vertebrate. The third part of this work involved a phylogeographic study of the adder (Vipera berus), the target species of this thesis. Three clades were revealed: a Balkan clade (corresponding to the subspecies V. b. sachalinensis), an unexpected Italian clade (limited to northern Italy, southern Austria, northern Slovenia and southeasternmost corner of Switzerland) and a Northern clade clade (including adders of the whole distribution area excepted animals from the Balkan and the Italian clades). The genetic variability within the Northern clade is sufficiently high to conclude that a northern glacial refugia during the last glaciation, in addition to those refugia already described for the main species (Iberian, Italian and Balkan peninsula). These results motivated a morphological study (part four) comparing the adders from the Italian and the Northern clades describe above. Only small morphological differences have been found, despite the split between these two clades have taken place more than 1 million years ago. A study on a local scale, focused on the Jura Mountains, on a few populations in the Alps and France was, performed to estimate the genetic diversity and the genetic structure between populations using microsatellite markers (part five). Considerable structure was observed between populations separated by more than 3 kilometres, whereas the structure between closer populations is less marked. Moreover, lower genetic diversity in the populations from Jura Mountains and Alps was noticed compared to populations from Massif Central of Atlantic coast. Such loss of genetic variation probably followed post-glacial recolonisation. The sixth study focused on the reproductive success of male adders in the wild. A positive correlation between body length and reproductive success was observed. Multiple paternity was also observed in most of clutches (69%) despite the low density of adders in the study area. Finally, no relationship was found between the number of fathers in a clutch and the survival of offspring at birth, contradicting previous studies. To conclude, the observation of a significant genetic structure in Vipera berus will enable recommendations to be made to improve protection of this species in the Jura Mountain. On a larger scale, the considerable genetic structure found within Vipera ammdoytes, V. aspis and V. berus, resulting from isolation in additional glacial refugia to those already described for other species, demonstrates the relevance of phylogeographic studies of reptiles to better understand the colonisation and disappearance during the last Tertiary and the Quaternary. The observation of several groups of evolutionary significant units (ESUs) within the three studied species might lead to a revision of the taxonomy, as well as their conservation status.

Linking the population genetics and ecology of arbuscular mycorrhizal fungi

Résumé Les champignons endomycorhiziens arbusculaires (CEA) ont co-évolué avec les plantes terrestres depuis plus de 400 millions d'années. De nos jours, les CEA forment une symbiose avec les racines de la majorité des plantes terrestres. Les CEA sont écologiquement importants parce qu'ils influencent non seulement la croissance des plantes, mais aussi leur diversité. Les CEA sont des biotrophes obligatoires qui reçoivent leur énergie sous forme de glucides issus de la photosynthèse des plantes. En contrepartie, les CEA apportent à leurs hôtes du phospore. Les CEA croissent et se reproduisent clonalement en formant des hyphes et des spores. De plus, les CEA sont coenocytiques et multigénomiques; le cytoplasme d'un CEA contient des noyeaux génétiquement différents. De nombreuses études ont démontré que différentes espèces de CEA agissent différentiellement sur la croissance des plantes. Malgré une conscience de plus en plus forte de l'existence d'une variabilité intraspécifique, la question de savoir si les populations de CEA sont génétiquement variables a été largement négligée. Dans le Chapitre 2, j'ai cherché à savoir si une population de CEA provenant d'un seul champ possède une diversité génétique. Cette étude a mis en évidence une importante variation génétique et phénotypique au sein d'individus de la même population. Des différences au niveau de traits de croissance, héritables et liés à la valeur sélective, indiquent que la variation génétique observée entre isolats n'est pas entièrement neutre. Dans le Chapitre 3, je montre que les différences génétiques entre isolats de CEA d'une population provoquent de la variation dans la croissance des plantes. L'effet des isolats dépend des conditions environnementales et varie de bénéfique à parasitique. Dans le Chapitre 4, je montre que des traits de croissance de CEA varient significativement dans des environnements contrastés. J'ai détecté de fortes interactions entre différents génotypes de CEA et différentes espèces de plantes. Ceci suggère que dans un environnement hétérogène, la sélection pourrait localement favoriser différents génotypes de CEA, maintenant ainsi la diversité génétique dans la population. Les résultats de ce travail aident à mieux comprendre l'importance écologique de la variation intraspécifique des CEA. La possibilité de pouvoir cultiver des individus d'une population de CEA au laboratoire nous a permis une meilleure compréhension de la génétique de ces champignons. De plus, ce travail est une base pour de futures expériences visant à comprendre l'importance évolutive de la diversité intraspécifique des CEA. Abstract Arbuscular mycorrhizal fungi (A1VIF) have co-evolved with land plants -for over 400 million years. Today, AMF form symbioses with roots of most land plants and are ecologically important because they alter plant growth and affect plant diversity. AMF are obligate biotrophs, obtaining their energy in form of plant-derived photosynthates. In return,- they supply their host plants with phosphorous. These fungi grow and reproduce clonally by hyphae and spores. They are coenocytic and multigenomic, harbouring genetically different nuclei in a common cytoplasm. Many studies have shown different AMF species differentially alter plant growth. Despite the increasing awareness of intraspecific variability the question whether there is any genetic variation among different individuals of the same population has been largely neglected. In Chapter 2, we investigated whether there is genetic diversity in a field population of the AMF G. intraradices. This work revealed that large genetic and heritable phenotypic variation exists in this AMF population. Differences in fitness-related growth traits among isolates suggest that some of the observed genetic variation is not selectively neutral. In Chapter 3, we show that genetic differences among isolates from the same population also cause variation in plant growth. The isolate effects on plant growth depended on the environmental conditions and varied from beneficial to detrimental. In Chapter 4, fitnessrelated growth traits of genetically different isolates were significantly altered in contrasting environments. we detected strong AMF isolate by host species interacfions which suggests that in a heterogeneous environment selection could locally favour different AMF genotypes, thereby maintaining high genetic diversity in the population. The results of this work contribute to the understanding of the ecological importance of intraspecific diversity in AMF. The possibility of culturing individuals of an AMF field population under laboratory condition gave new insights into AMF genetics and lays a foundation for future studies to analyse the evolutionary significance of intraspecific genetic diversity in AMF.