Rivulus marmoratus (Cyprinodont; Aplochelidae) is the only vertebrate known to reproduce by obligate internal self-fertilization. Selfing is the ultimate form of inbreeding and leads to virtual homozygosity in approximately ten generations, so that this mode of reproduction is effectively clonal. Populations of this species consist of arrays of homozygous clones, with high clonal diversity and low representation of each clone. Males occur at low frequency (<1%) in most populations, with a few notable exceptions. Females have never been recorded.High frequencies of males have been reported on two occasions; high numbers of males were reported on the island of Curacao in the 1970's, and males comprised up to 24% of the populations sampled on several Belize Cays during 1990-91. Hermaphrodites collected from one island, Twin Cays, proved to be heterozygous at mini-satellite loci when progeny tested and DNA fingerprinted. This was evidence that recent outcrossing had occurred.Earlier experiments with Floridian clonal lines had suggested that sex in this species was thermolabile; incubation of embryos at low temperatures (19° C)) produced up to 100% males. It was suggested that this might be part of an environmental sex determination (ESD) system in this species.If the induction of males is indeed part of and ESD in this species, then it may be that the temperature at which males are induced in a particular clone will be related to the temperature regime that clone encounters in nature. Therefore, it was hypothesized that the temperature at which males are induced would be related to the geographic origin of each clone, so that clones from the cooler extremes of the range would produce males at a lower threshold temperature than clones from the more equatorial center of the range. This study used laboratory reared descendants of clones from Vero Beach, Florida, Rio de Janiero, Brazil (the extreme north and south of the range), and from several Belize barrier islands and the Belize mainland (near the center of the range) in an effort to detect differences in the production of males between clonal lines based on their geographic origin. Eggs were collected from these hermaphrodites and incubated at 26° C)C, 22.5° C), or 19 ° C), representing normal, and below average temperatures for this species. These embryos were hatched and reared to sexual maturity at which time their sex was evaluated.Incubation at 26° C) resulted in 2-17% males, incubation at 22.5° C) resulted in 6-53% males, and incubation at 19° C) resulted in 47-74% males. However, the variation in male induction between clonal lines did not correspond to a geographic hypothesis.Also, significant numbers of deformed individuals were noted, particularly among individuals incubated at the lower two temperatures. Incubation at 26° C) resulted in 0-24% deformed, incubation at 22.5° C) resulted in 6-66% deformed, and incubation at 19° C) resulted in 36-87% deformed. It is clear that development at these lower temperatures is difficult for this species, and it may be that low temperature developmental stress is not a general challenge to this species. / Master of Science

xi, 103 p. : ill. A print copy of this thesis is available through the UO Libraries. Search the library catalog for the location and call number. / Although outcrossing is the most widespread mating system among animals and plants, the reason for this prevalence is not fully understood. Evolutionary theory has classified the potential selective pressures driving the evolution and maintenance of outcrossing into two broad categories: deleterious mutations and changing ecological conditions. Despite the inherent advantages of self-fertilization, exposure to either or both of these selective pressures is predicted to favor outcrossing over self-fertilization. I tested these predictions using experimental evolution in populations of Caenorhabditis elegans with genetically modified rates of outcrossing and selfing. I found that outcrossing reduces the fixation of deleterious mutations under mutation influx and that outcrossing expedites adaptation to a bacterial pathogen. Further, I identified facultative outcrossing, a novel life history characteristic, in specific C. elegans strains that predominantly reproduce by selfing but engage in outcrossing when stressed. The shift from a primarily selfing mating system to a predominantly outcrossing system is similar to the environmentally induced facultative sex observed in asexual species, which is thought to enable more rapid adaptation. Facultative outcrossing, although not previously documented, may play a major role in the life histories of many highly selfing species. Finally, most mutations are deleterious and therefore elevated mutation rates are generally thought to produce progressively larger reductions in fitness. Using the chemical mutagen ethylmethanesulfonate, I found the surprising result that populations exposed to a mutation rate at least fifty times greater than natural rates exhibited significantly greater fitness than populations exposed to substantially lower mutation rates. This unexpected fitness optimum may be the result of a volatile balance between the influx of deleterious mutations and compensatory mutations. This work confirms the predictions of several long-standing evolutionary theories by identifying both deleterious mutations and changing ecological conditions as selective pressures capable of driving the evolution and maintenance of outcrossing. These selective pressures, which are ubiquitous in nature, may explain the prevalence of outcrossing relative to selling. This dissertation includes previously published and co-authored materials. / Committee in charge: Barbara Roy, Chairperson, Biology; Patrick Phillips, Advisor, Biology; Karen Guillemin, Member, Biology; William Bradshaw, Member, Biology; Douglas Kennett, Outside Member, Anthropology

The ability of natural selection to promote and preserve genetic variation at Major Histocompatibility Complex (MHC) class I loci was examined in two fish species known to have low genetic variation either as a consequence of their breeding system or population structure. The tempo and modes of molecular evolution acting on these loci has also been discussed. The marine killifish Rivulus marmoratus is the only vertebrate known to exist in nature in homozygous form. The findings of this study suggest that MHC class I genes at one locus in R. marmoratus have rapidly accumulated variation, particularly in region of the gene encoding functional important domains. This rapid accumulation of variation is likely due to the increased intensity of natural selection acting on these genes resulting from the homozygosity of the species. The contention that the variation characterized at one MHC locus is of recent origin is further supported by an analysis of the mitochondrial control region if R. marmoratus, which suggests that the individuals included in this study are recently diverged. The variation characterized here is the first evidence of genetic variation at coding loci in this species, and may be evidence of the cost of homozygosity for R. marmoratus. Variation at a single MHC class I locus was also characterized in several members of the Death Valley pupfish species complex. Two of these species, Cyprinodon diabolis and Cyprinodon salinus have undergone intense historical or ongoing population bottlenecks. Despite the action of random genetic drift acting on these species, both maintain multiple MHC class I alleles at one locus. The alleles characterized in C. diabolis and C. salinus were also recovered from other members of the Death Valley complex. This suggests that overdominant natural selection has acted to maintain ancestral genetic variation at this locus in both species. This result has important implications for the design of conservation programs for these endangered species, and may have broader implications for the design of captive breeding programs of species with highly reduced population sizes in general. / Ph. D.

Organisms capable of self-fertilization typically exhibit two evolutionary syndromes uniting high inbreeding depression with low levels of selfing, or low inbreeding depression and high levels of selfing. This study tests for inbreeding depression in an apparent self-compatible, hermaphroditic marine nemertean worm, Prosorhochmus americanus. Fecundity and timing to first reproduction were assessed in isolated and paired worms. Isolated worms produced significantly more offspring than paired worms and did not show inbreeding avoidance. The selfing rate of natural populations was evaluated using species-specific microsatellites and is consistent with preferential selfing (mean: 0.801), though some outcrossing appears to take place. Population genetic structure indicates populations are disjunct and characterized by low levels of gene flow. Our results reveal P. americanus exhibits an interesting suite of life-history traits, uniting high dispersal potential through self-fertilization and high fecundity, with the lack of a dispersive larval stage and low levels of gene flow.

Part I: Coral reefs are essential to coastal economies, protecting coastlines from storms, and harboring high biodiversity. However, reefs are declining due to local anthropogenic stressors and ocean warming. Sedimentation, a local stressor, aggravates the impacts of warming on corals and hinders their survival and growth. Therefore, it is important to investigate whether sedimentation and temperature have a synergistic effect on vulnerable coral species, especially during earlier stages of development. To quantify these effects, survival and growth of newly settled Acropora cervicornis corals were measured at two temperatures (29 and 31°C, representing current and predicted for 2050 Summer temperatures) and three sediment concentrations (30, 60 and 120 mg.cm-2, representing a range from natural sedimentation to dredging conditions). The intent of this study was to mix multiple genotypes to test temperature and sedimentation among genotypic unique individuals. However, only 20% of colonies spawned, and spawning was asynchronous by genotype. Therefore, individuals were produced from self-fertilization. The overall high mortality seen in this study suggests that self-fertilization in A. cervicornis does not produce viable juveniles. Although temperature did not have a significant effect on the survival of self-fertilized juveniles, sediment concentration did. The lowest sediment concentration led to the highest juvenile survival in both ambient and heated conditions. The growth of A. cervicornis selfed individuals was not significantly affected by temperature or sedimentation. These results suggest that reducing sedimentation in dredging and coastal construction areas around coral reefs facilitates the survival of Acropora cervicornis juveniles and may help to ensure their persistence in the future. Because self-fertilized larvae were used, these results need to be interpreted with caution, and this research needs to be repeated with outcrossed A. cervicornis. What is clear is that genotypic diversity is needed for A. cervicornis population growth and resilience. Part II: Chimeras occur when two or more genetically unique individuals of the same species fuse together. The presence of chimerism can aid in the survival and evolution of organisms. This study investigated whether the prevalence of chimerism differs between coral species of different reproduction modes and growth rates. To fulfill this goal, the surface area of egg and/or larvae of three coral species, Montastraea cavernosa, Acropora cervicornis, and Porites astreoides, were measured and compared with the respective surface areas of the newly settled juveniles. This comparison suggested that M. cavernosa displayed a greater tendency to form chimeras than A. cervicornis and P. astreoides. Observations during embryogenesis confirmed this prediction. Montastraea cavernosa is a slow grower and has the smallest eggs of all three study species. Chimerism during embryogenesis may increase this species’ tendency to start the sessile stage at a slightly bigger size and thus increases its competitive abilities for reef space. In contrast, A. cervicornis is a broadcast spawner, fast grower and has a relatively larger egg size, possibly explaining the reduced chimeric tendency during embryogenesis. It is possible that P. astreoides formed chimeras during embryogenesis within the polyp, but they did not form them in the swimming planulae stage. The lack of chimerism during P. astreoides planulae development may be attributed to its brooding reproductive mode, directly releasing large competent larvae that have large initial sizes at settlement. Therefore, the ability to form chimeras in an early developmental stage might provide an ecological advantage to M. cavernosa that contributes to its abundance in Broward County: the greater size at settlement caused by chimerism during embryogenesis may provide this species a competitive advantage for reef space.

Genealogy reconstruction is widely used in biology when relationships among entities are studied. Phylogenies, or evolutionary trees, show the differences between species. They are of profound importance because they help to obtain better understandings of evolutionary processes. Pedigrees, or family trees, on the other hand visualize the relatedness between individuals in a population. The reconstruction of pedigrees and the inference of parentage in general is now a cornerstone in molecular ecology. Applications include the direct infer- ence of gene flow, estimation of the effective population size and parameters describing the population’s mating behaviour such as rates of inbreeding. In the first part of this thesis, we construct genealogies of various types of cancer. Histopatho- logical classification of human tumors relies in part on the degree of differentiation of the tumor sample. To date, there is no objective systematic method to categorize tumor subtypes by maturation. We introduce a novel algorithm to rank tumor subtypes according to the dis- similarity of their gene expression from that of stem cells and fully differentiated tissue, and thereby construct a phylogenetic tree of cancer. We validate our methodology with expression data of leukemia and liposarcoma subtypes and then apply it to a broader group of sarcomas and of breast cancer subtypes. This ranking of tumor subtypes resulting from the application of our methodology allows the identification of genes correlated with differentiation and may help to identify novel therapeutic targets. Our algorithm represents the first phylogeny-based tool to analyze the differentiation status of human tumors. In contrast to asexually reproducing cancer cell populations, pedigrees of sexually reproduc- ing populations cannot be represented by phylogenetic trees. Pedigrees are directed acyclic graphs (DAGs) and therefore resemble more phylogenetic networks where reticulate events are indicated by vertices with two incoming arcs. We present a software package for pedigree reconstruction in natural populations using co-dominant genomic markers such as microsatel- lites and single nucleotide polymorphism (SNPs) in the second part of the thesis. If available, the algorithm makes use of prior information such as known relationships (sub-pedigrees) or the age and sex of individuals. Statistical confidence is estimated by Markov chain Monte Carlo (MCMC) sampling. The accuracy of the algorithm is demonstrated for simulated data as well as an empirical data set with known pedigree. The parentage inference is robust even in the presence of genotyping errors. We further demonstrate the accuracy of the algorithm on simulated clonal populations. We show that the joint estimation of parameters of inter- est such as the rate of self-fertilization or clonality is possible with high accuracy even with marker panels of moderate power. Classical methods can only assign a very limited number of statistically significant parentages in this case and would therefore fail. The method is implemented in a fast and easy to use open source software that scales to large datasets with many thousand individuals.

De nombreuses populations voient leurs conditions de vie modifiées par les changements globaux. Au-delà de leurs conséquences écologiques, les changements globaux peuvent également modifier les régimes de sélection des populations. Le déclin récent des pollinisateurs pourrait altérer fortement le succès reproducteur de nombreuses populations de plantes à fleurs. Cependant, ses conséquences évolutives n'ont pas été étudiées jusqu'ici. Cette thèse traite de la possibilité d'adaptation des stratégies de reproduction des plantes face aux changements globaux et plus spécifiquement face au déclin des pollinisateurs. Cette question a été abordée en deux temps. Premièrement, l'analyse de l'abondante littérature théorique et empirique sur les systèmes de reproduction des plantes et dans un moindre mesure la construction d'un modèle d'évolution des traits floraux d'attraction ont permis de clarifier des scénarios d'évolution à court terme. Deuxièmement, une approche empirique originale a été menée, consistant à comparer directement les traits de populations ancestrales et descendantes de trois espèces annuelles, à partir de la culture en jardin commun de graines anciennes et récentes en provenance de régions où des indices de déclin des pollinisateurs existent. Cette approche permet de mettre clairement en évidence l'évolution génétique. Les données existantes indiquent que le déclin des pollinisateurs peut accentuer la limitation pollinique et par conséquent augmenter la sélection sur les traits floraux. Par ailleurs, la variation génétique substantielle dans les populations suggère que l'évolution rapide des populations de plantes est possible. L'analyse de la littérature et notre étude théorique suggèrent que l'accroissement de la capacité d'autofécondation autonome ou l'accroissement de l'attraction des pollinisateurs sont deux scénarios d'évolution possibles. Deux types de réponses évolutives ressortent de nos travaux expérimentaux. D'abord, une avancée du calendrier de floraison a été retrouvée chez les trois espèces étudiées. Ce résultat souligne le rôle de l'évolution génétique, et pas seulement de la plasticité phénotypique, dans les avancées de phénologies de printemps que l'on retrouve chez de très nombreux organismes. Ensuite, contrairement aux traits de phénologie, les traits floraux ont évolué dans des directions opposées selon les espèces. Ainsi, des traits floraux a priori plus attractifs ont évolué chez une des espèces tandis que pour une autre, des traits floraux a priori moins attractifs ont évolué, mais s'accompagnent d'une meilleure capacité d'autofécondation autonome. Cette étude confirme que les traits de reproduction des plantes peuvent évoluer en quelques décennies seulement. La possibilité du sauvetage évolutif des populations, par l'évolution rapide des traits et des stratégies de reproduction des plantes, est une perspective de recherche qui découle de ces résultats. / Global change alters life conditions of numerous populations. Beyond ecological consequences, global change can also modify selection regimes in populations. While the recent pollinator decline may specifically affect the reproductive success of flowering plants, its evolutionary consequences have not been studied yet. This thesis deals with the possibility of adaptation of plant reproductive strategies under global change and more specifically under pollinator decline. This question was addressed in two steps. First, the analysis of the extensive literature on plant mating systems, and, to a lesser extent, the construction of a model for the evolution of attractive floral traits, allowed us to clarify evolutionary scenarios at short-time scales. Second, we conducted an original empirical approach, consisting in the direct comparison of ancestral and descendant populations by re-growing old and recent seeds under identical conditions. This approach allowed us to test whether genetic evolution of reproductive traits occurred in the context of pollinator decline for three annual plant species. Data from the literature indicates that pollinator decline likely increases pollen limitation and thus selection on floral traits. Moreover, the substantial genetic variation within populations suggests that rapid evolution is possible. Increased autonomous selfing or increased pollinator attraction are two possible routes of plant adaptation to pollinator decline emerging from the analysis of the literature and from our theoretical study. Our empirical work brings out two types of evolutionary trends. Firstly, earlier flowering phenology was found in all three studied species. This result shows that genetic evolution, not only phenotypic plasticity; certainly contribute to the spring phenological advancements reported for numerous species. Secondly, unlike phenological traits, floral traits evolved in opposite directions depending on the species. Showy floral traits evolved in a species while joint evolution of autonomous selfing with a reduction of floral attractiveness seemed to evolve in another species. This study shows that plant reproductive traits can evolve in a few decades. Whether or not rapid evolution of plant reproductive traits can act as an evolutionary rescue for threatened populations is a research question that arises from these results.

Genealogy reconstruction is widely used in biology when relationships among entities are studied. Phylogenies, or evolutionary trees, show the differences between species. They are of profound importance because they help to obtain better understandings of evolutionary processes. Pedigrees, or family trees, on the other hand visualize the relatedness between individuals in a population. The reconstruction of pedigrees and the inference of parentage in general is now a cornerstone in molecular ecology. Applications include the direct infer- ence of gene flow, estimation of the effective population size and parameters describing the population’s mating behaviour such as rates of inbreeding. In the first part of this thesis, we construct genealogies of various types of cancer. Histopatho- logical classification of human tumors relies in part on the degree of differentiation of the tumor sample. To date, there is no objective systematic method to categorize tumor subtypes by maturation. We introduce a novel algorithm to rank tumor subtypes according to the dis- similarity of their gene expression from that of stem cells and fully differentiated tissue, and thereby construct a phylogenetic tree of cancer. We validate our methodology with expression data of leukemia and liposarcoma subtypes and then apply it to a broader group of sarcomas and of breast cancer subtypes. This ranking of tumor subtypes resulting from the application of our methodology allows the identification of genes correlated with differentiation and may help to identify novel therapeutic targets. Our algorithm represents the first phylogeny-based tool to analyze the differentiation status of human tumors. In contrast to asexually reproducing cancer cell populations, pedigrees of sexually reproduc- ing populations cannot be represented by phylogenetic trees. Pedigrees are directed acyclic graphs (DAGs) and therefore resemble more phylogenetic networks where reticulate events are indicated by vertices with two incoming arcs. We present a software package for pedigree reconstruction in natural populations using co-dominant genomic markers such as microsatel- lites and single nucleotide polymorphism (SNPs) in the second part of the thesis. If available, the algorithm makes use of prior information such as known relationships (sub-pedigrees) or the age and sex of individuals. Statistical confidence is estimated by Markov chain Monte Carlo (MCMC) sampling. The accuracy of the algorithm is demonstrated for simulated data as well as an empirical data set with known pedigree. The parentage inference is robust even in the presence of genotyping errors. We further demonstrate the accuracy of the algorithm on simulated clonal populations. We show that the joint estimation of parameters of inter- est such as the rate of self-fertilization or clonality is possible with high accuracy even with marker panels of moderate power. Classical methods can only assign a very limited number of statistically significant parentages in this case and would therefore fail. The method is implemented in a fast and easy to use open source software that scales to large datasets with many thousand individuals.:Abstract v Acknowledgments vii 1 Introduction 1 2 Cancer Phylogenies 7 2.1 Introduction..................................... 7 2.2 Background..................................... 9 2.2.1 PhylogeneticTrees............................. 9 2.2.2 Microarrays................................. 10 2.3 Methods....................................... 11 2.3.1 Datasetcompilation ............................ 11 2.3.2 Statistical Methods and Analysis..................... 13 2.3.3 Comparison of our methodology to other methods . . . . . . . . . . . 15 2.4 Results........................................ 16 2.4.1 Phylogenetic tree reconstruction method. . . . . . . . . . . . . . . . . 16 2.4.2 Comparison of tree reconstruction methods to other algorithms . . . . 28 2.4.3 Systematic analysis of methods and parameters . . . . . . . . . . . . . 30 2.5 Discussion...................................... 32 3 Wild Pedigrees 35 3.1 Introduction..................................... 35 3.2 The molecular ecologist’s tools of the trade ................... 36 3.2.1 3.2.2 3.2.3 3.2.1 Sibship inference and parental reconstruction . . . . . . . . . . . . . . 37 3.2.2 Parentage and paternity inference .................... 39 3.2.3 Multigenerational pedigree reconstruction . . . . . . . . . . . . . . . . 40 3.3 Background..................................... 40 3.3.1 Pedigrees .................................. 40 3.3.2 Genotypes.................................. 41 3.3.3 Mendelian segregation probability .................... 41 3.3.4 LOD Scores................................. 43 3.3.5 Genotyping Errors ............................. 43 3.3.6 IBD coefficients............................... 45 3.3.7 Bayesian MCMC.............................. 46 3.4 Methods....................................... 47 3.4.1 Likelihood Model.............................. 47 3.4.2 Efficient Likelihood Calculation...................... 49 3.4.3 Maximum Likelihood Pedigree ...................... 51 3.4.4 Full siblings................................. 52 3.4.5 Algorithm.................................. 53 3.4.6 Missing Values ............................... 56 3.4.7 Allelefrequencies.............................. 58 3.4.8 Rates of Self-fertilization.......................... 60 3.4.9 Rates of Clonality ............................. 60 3.5 Results........................................ 61 3.5.1 Real Microsatellite Data.......................... 61 3.5.2 Simulated Human Population....................... 62 3.5.3 SimulatedClonalPlantPopulation.................... 64 3.6 Discussion...................................... 71 4 Conclusions 77 A FRANz 79 A.1 Availability ..................................... 79 A.2 Input files...................................... 79 A.2.1 Maininputfile ............................... 79 A.2.2 Knownrelationships ............................ 80 A.2.3 Allele frequencies.............................. 81 A.2.4 Sampling locations............................. 82 A.3 Output files..................................... 83 A.4 Web 2.0 Interface.................................. 86 List of Figures 87 List of Tables 88 List Abbreviations 90 Bibliography 92 Curriculum Vitae I

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