Nitrogen (N) derived from symbiotic fixation of atmospheric N2 in wild and cultivated populations of Cyclopia, a woody endemic genus used to make honeybush tea in the Western Cape of South Africa, was quantified by the 15N natural abundance method. Because Cyclopia species are naturally mycorrhizal, non-N2-fixing arbuscular mycorrhizal shrubs of similar phenology to Cyclopia were chosen as reference plants to provide the d15Nvalue of soil-derived N. Isotopic analysis showed that wild populations of Cyclopia were highly dependent on N2 fixation for theirNnutrition, ranging from 70 ± 4%to 100 ± 7% (mean ± SE) at all sites, except for one. Further evidence of the high dependency of wild Cyclopia populations on symbiotic N was provided by their significantly higher foliar N concentrations compared with the non-legume reference plants. However, cultivated Cyclopia exhibited variable amounts of N2 fixation, with Cyclopia genistoides (L.) R. Br., for example, showing low amounts of N2 fixation at Sites P2 and P3 (0 ± 51% and 8 ± 46%, respectively) as a result of low D values (D is defined as the difference between the mean d15N value of the reference plants and the B value of the test Cyclopia species, where B is the d15N of an inoculated test legume grown in an N-free growth medium), whereas at Sites P1, P2, P5 and P6, up to 89 ± 2%, 94 ± 13%, 85 ± 13% and 100 ± 18%, respectively, plant N was derived from atmospheric fixation. The high symbiotic N nutrition observed for wild populations of Cyclopia suggests that these populations are major contributors to the N economy of the nutrientpoor soils of the South African fynbos. These data indicate that by breeding for high N2 fixation rates in Cyclopia cultivars and selecting more efficient rhizobial strains, this legume has the potential to achieve higher N2 fixation rates under cultivation. The low variability in Cyclopia d15N values within sites, however, suggests that genetic variability is not a major factor influencing N2 fixation rates in cultivated Cyclopia, and thatmore benefit may be gained from soil amelioration and the selection of improved rhizobial strains

Transposable elements (TEs) are one major force of genome evolution thanks to theirability to create genetic variation. TEs are ubiquitous and their proportion is variable between species and also populations, suggesting that a tight relationship exists between genomes and TEs. The model system composed of the natural populations of the twin sisters Drosophila melanogaster and D. simulans is interesting to compare host/TE relationship, since both species harbour different amounts of TE copies. The helena element is nearly silenced in D.simulans natural populations despite a very high copy number. Such repression is associated to abundant internally deleted copies suggesting a regulatory mechanism of TEs based on DNA deletion. Another pathway of TE regulation is through epigenetics where the host genome is able to keep intact the DNA sequences of TEs and still silence their activities.Chromatin remodelling is well known in drosophila and specific histone modifications can be associated to specific chromatin domains. We observed an important variation on H3K27me3and H3K9me2, two heterochromatic marks, on TE copies in D. melanogaster and D. simulans natural populations. Also, we show that derepressed lines of D. simulans exist for specific elements, have high TE transcription rates and are highly associated to non constitutive heterochromatic marks. TEs are therefore controlled by the host genome through DNA deletion and a possible chromatin remodelling mechanism. Not only genetic variability is enhanced by TEs but also epigenetic variability, allowing the host genome to be partitioned into chromatin domains. TEs are therefore mandatory to gene network regulation through their ability of "jumping epigenetics".

The fundamental objective of conservation genetics is the identification of the basic units of conservation. Central to this objective is the reconstruction of the adaptive and evolutionary history of populations to evaluate their conservation status. Evolutionary history involves both microevolutionary and macroevolutionary processes and adaptive history is the evolution of specific characters to selective ecological processes in differential heterogeneous environments. Neutral DNA markers such as mitochondrial DNA, minisatellites and microsatellites are most often used for reconstructing history and identifying conservation units. This thesis examined three biological systems: 1) an African cichlid, 2) Canadian moose populations and 3) eastern North American wolves and coyotes to test two hypotheses. Firstly, neutral DNA markers can be used to accurately reconstruct the evolutionary history of populations. Secondly, neutral DNA markers are concordant with adaptive distinctiveness in reconstructing the adaptive history of populations. Few studies have examined these relationships. Lake Magadi tilapia showed discordant patterns between adaptive morphological, physiological and behavioural characters and genetic structure assessed with mitochondrial DNA. I propose this discordance has resulted from selection acting on mitochondrial DNA that has often been assumed to be "neutral". Neutral DNA markers accurately reflected the known history of the moose populations but discordant patterns were observed between neutral and functional loci indicating the former may not accurately reflect adaptive variation. DNA profiles of eastern wolves and coyotes showed a significant conflict in the interpretation of mtDNA and microsatellite data compared to previous genetic studies that examined wolf taxonomy. The data were consistent with the hypothesis of a North American-evolved wolf. Coyote-like mtDNA was not of coyote origin but represented divergent but related sequences of a North American wolf lineage independent of the gray wolf (C. lupus). Under this new model of eastern wolf evolution, we also identified the hybrid origin of eastern coyotes, contrary to previous interpretations, and genetically characterised different wolf "types" within Ontario. These findings could not reject the first hypothesis as neutral markers were used to reconstruct the histories of the three biological systems. However, the findings identified that it is important to ensure the neutrality of DNA markers and that samples are representative of the taxa under investigation. The findings in this thesis did not support the second hypothesis, as neutral DNA markers were not concordant with adaptive characters, i.e. morphology, physiology and functional genetic markers. / Thesis / Candidate in Philosophy

Les éléments transposables (ET) sont une source majeure de variation génétique, ce qui leur confère un rôle essentiel dans l’évolution des génomes. Certes présents dans tous les génomes analysés à ce jour, leurs proportions sont fortement variables entre espèces et aussi entre populations, suggérant une relation unique entre génome hôte et ET. Grâce à un système modèle composé de populations naturelles de deux espèces proches (Drosophila melanogaster et D. simulans) avec des quantités différentes en ET, nous avons pu comparer les relations génome hôte/ET. Nous nous sommes particulièrement interessés à l’élément helena qui, chez D. simulans, montre une activité faible, malgré un nombre de copies élevé.Cette activité moindre est associée à de nombreuses délétions internes des copies, suggérant un mécanisme de régulation d’ET par des délétions de l’ADN. Un autre système de régulation de l’activité des ET utilise le contrôle épigénétique, ce qui permet le maintien des copies d’ET dans le génome mais un blocage de leur activité. Le remodelage de la chromatine est un système épigénétique bien décrit chez la drosophile. Les régions chromatiniennes des génomes sont associées à différents types de modifications d’histone. Nous avons mis en évidence, dans des populations de D. melanogaster et D. simulans, une variation conséquente de modifications d’histones de type hétérochromatique, H3K27me3 et H3K9me2, associées àdes copies de différents ET. De plus, nous avons décrit des populations chez D. simulans dites déréprimées, chez lesquelles certains éléments sont surexprimés et présentent des localisations probablement hétéchromatiques. Les ET sont donc contrôlés par le génome hôte par des délétions internes et probablement par un système épigénétique variable. De plus, dans certaines populations, des copies peuvent échapper à ce contrôle et envahir le génome. Les ET sont donc des grands créateurs de variabilité génétique mais permettent aussi une territorialisation chromatinienne du génome car ils portent des modifications épigénétiques précises et sont capables de les étendre à leurs environnements génomiques. Ceci leur confére la fonction "d'épigénétique mobile". / Transposable elements (TEs) are one major force of genome evolution thanks to theirability to create genetic variation. TEs are ubiquitous and their proportion is variable between species and also populations, suggesting that a tight relationship exists between genomes and TEs. The model system composed of the natural populations of the twin sisters Drosophila melanogaster and D. simulans is interesting to compare host/TE relationship, since both species harbour different amounts of TE copies. The helena element is nearly silenced in D.simulans natural populations despite a very high copy number. Such repression is associated to abundant internally deleted copies suggesting a regulatory mechanism of TEs based on DNA deletion. Another pathway of TE regulation is through epigenetics where the host genome is able to keep intact the DNA sequences of TEs and still silence their activities.Chromatin remodelling is well known in drosophila and specific histone modifications can be associated to specific chromatin domains. We observed an important variation on H3K27me3and H3K9me2, two heterochromatic marks, on TE copies in D. melanogaster and D. simulans natural populations. Also, we show that derepressed lines of D. simulans exist for specific elements, have high TE transcription rates and are highly associated to non constitutive heterochromatic marks. TEs are therefore controlled by the host genome through DNA deletion and a possible chromatin remodelling mechanism. Not only genetic variability is enhanced by TEs but also epigenetic variability, allowing the host genome to be partitioned into chromatin domains. TEs are therefore mandatory to gene network regulation through their ability of “jumping epigenetics”.

The “Free Hormone Hypothesis” posits that only free, unbound hormone is biologically active and available to tissues. Conventional biomedical wisdom proposes that corticosteroid-binding globulin (CBG) normally binds 90-95% of blood glucocorticoid (GC), rendering it unavailable to tissues. Under chronic stress, GC levels greatly exceed binding capacity resulting in impaired bodily function and reduced fitness. However, under normal conditions in northern and southern flying squirrels, less than 10% of GC is bound, presenting a major challenge to the hypothesis. To assess the extent of variation in these properties among vertebrates, I compared all species (88) with known GC and CBG and levels. 92% conform reasonably to known convention. Flying squirrels appear as extreme species, as do New World monkeys, yet both groups evolved from ancestors that followed normal convention. I speculate as to how this state evolved and persisted through time.

The “Free Hormone Hypothesis” posits that only free, unbound hormone is biologically active and available to tissues. Conventional biomedical wisdom proposes that corticosteroid-binding globulin (CBG) normally binds 90-95% of blood glucocorticoid (GC), rendering it unavailable to tissues. Under chronic stress, GC levels greatly exceed binding capacity resulting in impaired bodily function and reduced fitness. However, under normal conditions in northern and southern flying squirrels, less than 10% of GC is bound, presenting a major challenge to the hypothesis. To assess the extent of variation in these properties among vertebrates, I compared all species (88) with known GC and CBG and levels. 92% conform reasonably to known convention. Flying squirrels appear as extreme species, as do New World monkeys, yet both groups evolved from ancestors that followed normal convention. I speculate as to how this state evolved and persisted through time.

Mate choice among males is relatively understudied, despite recent evidence supporting its ubiquity. Theory predicts male mate choice in response to variation in female quality, and male mating strategies that limit polygyny. However empirical research investigating these connections, particularly under natural conditions, is generally lacking. Using the Western black widow spider, Latrodectus hesperus, I investigated male mate choice, male investment, and the advantage of mating with females that differ in their potential to deliver fitness benefits to males via higher fecundity and/or reduced risk of sperm competition. Males were found to be preferentially attracted to larger, high-diet virgin females over females that were smaller due to a lower diet, or having been previously mated, or both. Through a three-year field study on females I found that males likely benefit from selectivity, as smaller, low-diet females often failed to deposit any egg sacs throughout the breeding season, likely due to their shortened web-site tenure and/or lack of bodily resources. I investigated the costs and benefits to male genital breakage in L. hesperus, a strategy typically assumed to result in male sterility while providing paternity protection. I found that genital mutilation in L. hesperus did not cause sterility and that males were capable of inseminating multiple females, likely because of the comparatively low amount of genital damage and the likelihood of the efficient placement of genital fragments. Evidence from double mating trials supported the efficacy of broken genital fragments as plugs, but first male sperm precedence was often maintained in cases where male genital breakage failed or fragments were positioned incorrectly, which occurred frequently. Together, these laboratory and field experiments contribute to a more complete view of mate choice.

Mate choice among males is relatively understudied, despite recent evidence supporting its ubiquity. Theory predicts male mate choice in response to variation in female quality, and male mating strategies that limit polygyny. However empirical research investigating these connections, particularly under natural conditions, is generally lacking. Using the Western black widow spider, Latrodectus hesperus, I investigated male mate choice, male investment, and the advantage of mating with females that differ in their potential to deliver fitness benefits to males via higher fecundity and/or reduced risk of sperm competition. Males were found to be preferentially attracted to larger, high-diet virgin females over females that were smaller due to a lower diet, or having been previously mated, or both. Through a three-year field study on females I found that males likely benefit from selectivity, as smaller, low-diet females often failed to deposit any egg sacs throughout the breeding season, likely due to their shortened web-site tenure and/or lack of bodily resources. I investigated the costs and benefits to male genital breakage in L. hesperus, a strategy typically assumed to result in male sterility while providing paternity protection. I found that genital mutilation in L. hesperus did not cause sterility and that males were capable of inseminating multiple females, likely because of the comparatively low amount of genital damage and the likelihood of the efficient placement of genital fragments. Evidence from double mating trials supported the efficacy of broken genital fragments as plugs, but first male sperm precedence was often maintained in cases where male genital breakage failed or fragments were positioned incorrectly, which occurred frequently. Together, these laboratory and field experiments contribute to a more complete view of mate choice.

Across vertebrates, skeletal shapes are diverse, and much of this variation appears to be adaptive. In contrast, the early developmental programs of these structures are highly conserved across vertebrates. The question then becomes where in the conserved genetic programs of skeletal development does variation lie to direct diversity? In threespine stickleback, rapid changes in head and body shape have been documented during repeated and independent invasions of oceanic fish into freshwater habitats in regions deglaciated approximately 13,000 years ago. However, recent research indicates that similar phenotypic and genetic divergence can occur in decades. A remaining challenge is to link stickleback population genomic variation to causal genes that underlie such rapid phenotypic evolution. Here I use genome wide association studies (GWAS) in natural populations of stickleback to uncover genomic regions that contribute to variation of two dermal bone derived traits, lateral plate number and opercle shape. The decrease of lateral plate body armor and change in opercle bone shape, important for feeding mechanics, are classically associated with freshwater divergence. GWAS has recently begun to be used in natural populations but is still under scrutiny for performance among different populations. Using a population of phenotypically variable stickleback in Oregon, GWAS proved an effective method to uncover genomic regions and genetic variants known to contribute to lateral plate number and opercle shape, as well as new genomic regions and candidate genes not previously implicated in phenotypic variation. Although successful, using similar methods on decades old stickleback populations in Alaska revealed the challenges that accompany controlling population structure created by strong natural selection. Together, I found that although lateral plate number and opercle shape rapidly evolve in a coordinated fashion during adaptation from marine to freshwater environments, phenotypic variation is largely driven by independent genetic architectures. However, in very rapidly evolving populations, despite this independence of genetic architecture, the genetic variants contributing to the traits co-localize to similar genomic regions. This finding could be either biological or methodological which highlights the promise and limitations of using GWAS to identify genetic variation that gives rise to phenotypic diversity. This dissertation includes unpublished co-authored material.

A population of the rare Southern Appalachian endemic species Lilium grayi, (Gray’s lily) Roan Mountain, TN/NC was monitored for 2 years to determine the cause and impact of an early season collapse. High concentrations of the Lilium spp. host-specific fungal phytopathogen, Pseudocercosporella inconspicua (G. Winter) U. Braun were associated with 19/20 symptomatic and 0/30 asymptomatic plants. Strength of the association between pathogen and disease and the replication of disease symptoms in 4/4 healthy hosts showed that P. inconspicua was the causal agent of the disease referred to as lily leaf spot. Disease had a severe impact on the population with 59% of mature and 98% of adolescent plants undergoing early senescence. Only 32% of mature plants produced capsules and they were frequently diseased. A recurring spatiotemporal pattern typical of an infectious disease suggested that the lily leaf spot disease is capable of causing sequential annual epidemics of unknown long-term consequences to the stability of the host population.