Broad-scale population genetics can reveal population structure across an organism’s entire range, which can enable us to determine the most efficient population-wide management strategy depending on levels of connectivity. Genetic variation and differences in genetic diversity on small-scales have been reported in anemones, but nothing is known about their broad-scale population structure, including that of “host” anemone species, which are increasingly being targeted in the aquarium trade. In this study, microsatellite markers were used as a tool to determine the population structure of a sessile, host anemone species, Heteractis magnifica, across the Indo-Pacific region. In addition, two rDNA markers were used to identify Symbiodinium from the samples, and phylogenetic analyses were used to measure diversity and geographic distribution of Symbiodinium across the region. Significant population structure was identified in H. magnifica across the Indo-Pacific, with at least three genetic breaks, possibly the result of factors such as geographic distance, geographic isolation and environmental variation. Symbiodinium associations were also affected by environmental variation and supported the geographic isolation of some regions. These results suggests that management of H. magnifica must be implemented on a local scale, due to the lack of connectivity between clusters. This study also provides further evidence for the combined effects of geographic distance and environmental distance in explaining genetic variance.

In a recent global study of whale shark population genetics, aggregations were found to belong to either the Indo-Pacific or Atlantic population. This overview included an aggregation found within the Red Sea near Al Lith, Saudi Arabia, however the Mafia Island, Tanzania, aggregation was not part of the study. Both aggregations have unique aspects with the Saudi Arabian individuals showing sexual parity with no segregation, while recent acoustic results have revealed cryptic residency at Mafia Island. Genetic analysis using 11 microsatellite markers was performed on whale sharks from both locations. A combination of primers sourced from previous studies and newly designed primers were used to compare both aggregations and the individuals within. Samples were collected in the Red Sea for 5 seasons spanning 6 years, and for 2 seasons in Tanzania. Analysis with STRUCTURE showed a lack of significant genetic differences between the two aggregations, confirming that whale sharks in Tanzania are part of the Indo-Pacific population. Kinship analysis using COLONY found two potential pairs of full siblings in Tanzania. One pair had a high probability (.993) of being a full sibling dyad while the other had a lower probability (.357). There were no sibling pairs identified from the Red Sea aggregation. Genetic diversity was investigated using allelic richness over the 6 seasons at Al Lith, with values showing no significant change. This is in contrast to results that showed a decline in genetic diversity at Western Australia’s Ningaloo reef. These differences, however, only highlight the need for genetic diversity studies over longer time periods and at other aggregations within the Indo-Pacific.

Genetic, ecological, and environmental variation within and among populations of hosts and pathogens can influence local susceptibility of hosts and transmission rates of pathogens. These complex interactions can lead to geographic variation in the prevalence of pathogens. The interaction between leprosy and nine-banded armadillos in the southeastern U.S. provides an opportunity to examine the genetic, ecological, and environmental factors contributing to the variation in pathogen prevalence on a large geographic scale. Using genotypic data for loci associated with the immune response to leprosy and for anonymous loci, we have assessed the population structure of armadillo populations across the southern U.S. Additional statistical tests were used to assess any significant differences between functional and anonymous loci that indicate selection. Our results suggest that local adaptation does not influence the geographic distribution of leprosy in armadillo populations and that very little genetic variation can be attributed to differences among host populations.

The ability of populations to continually respond to directional selection even after many generations instead of reaching response plateaus suggests the presence of mechanisms for rapidly generating novel adaptive variation within organismal genomes. The contributions of cis regulation are now being widely studied. This study details the contributions of one such mechanism capable of generating adaptive genetic variation through transcribed microsatellite mutation. Microsatellites are abundant in eukaryotic genomes, exhibit one of the highest known mutation rates; and mutations involve indels that are reversible. These features make them excellent candidates for generating variation in populations. This study explores the functional roles of transcribed microsatellites in Helianthus annuus (common sunflower). More specifically, I explored the role of microsatellites as agents of rapid change that act as “tuning knobs” of phenotypic variation by influencing gene expression in a stepwise manner by expansions and contractions of the microsatellite tract. A bioinformatic study suggests that selection has favored expansion and maintenance of transcriptomic microsatellites. This inference is based on the non-random distribution of microsatellites, prevalence of motifs associated with gene regulation in untranslated regions, and the enrichment of microsatellites in Gene Ontologies representing plant response to stress and stimulus. A population genetics study provides support for selection on these transcribed microsatellites when compared to anonymous microsatellites that were assumed to evolve neutrally. The natural populations utilized in this study show greater similarity in allele frequencies, mean length, and variance in lengths at the transcribed microsatellites relative to that observed at anonymous microsatellite loci. This finding is indicative of balancing selection, and provides evidence that allele lengths are under selection. This finding provides support for the tuning knob hypothesis. The findings of a functional genomic study with regard to the tuning knob hypothesis are ambiguous. No correlation between allele lengths and gene expression was detected at any of three loci investigated. However, the loci utilized exhibited narrow ranges in length. The tuning knob hypothesis implies that similar allele lengths are likely to exhibit similar gene expression levels. Hence, variation in the populations studied may be tracking the optimal gene expression levels.

The genetic mechanisms by which natural populations maintain abundant phenotypic variation and adapt to their local environments remains a controversial topic in evolutionary biology. An intriguing mechanism involving highly mutable microsatellites follows the “tuning knob” model which proposes that stepwise changes in microsatellite allele lengths reciprocally generate phenotypic variation in a stepwise manner. In this study, I explored the predictions of the tuning knob model focusing specifically on transcribed microsatellites within and among natural populations of common sunflower (Helianthus annuus L.) transecting a latitudinal cline. An RNA-Seq experiment was conducted on 95 individuals from Kansas and Oklahoma grown in a common garden. To explore the potential role that microsatellites play in gene expression divergence in common sunflower, enrichment of microsatellites within differentially expressed (DE) genes was assessed. The results showed that A and AG repeat-containing microsatellites are enriched within DE genes and that 83.5% of these microsatellites are located within untranslated regions (UTRs). This finding is consistent with a role for transcribed microsatellites in gene expression divergence. RNA-Seq data were then used to assess microsatellite allele length effects on gene expression. Of all the microsatellites characterized in a reference transcriptome, 3,325 were consistently genotyped. The study identified 479 microsatellites at which allele length significantly correlated with gene expression (eSTRs). When irregular allele sizes were removed from the analysis, the number of eSTRs rose to 2379. eSTRs were most abundant within UTRs (70.4%) which suggests that they are well-positioned as cis-regulatory elements. A population genetic study conducted with 672 individuals across 17 sunflower populations from Saskatchewan to Oklahoma revealed strong signatures of directional selection acting on 13 eSTRs compared to 19 anonymous microsatellites assumed to evolve in a neutral fashion. This demonstrates that longer or shorter alleles may be favored in more extreme environments to that considered in the focal study. A second common garden experiment conducted with populations further north and south of focal populations revealed consistent patterns of correlation between microsatellite allele length and gene expression at some eSTRs. This study provides evidence that a substantial number of transcribed microsatellites function as “tuning knobs” of adaptation in common sunflower by modulating gene expression in a stepwise manner. These findings imply that the genomes of natural populations may include hundreds of active tuning knobs that can facilitate rapid evolution.

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Advances in aquaculture technologies are being investigated to support the replenishment of local fisheries, develop marine food fish farming opportunities and to increase seafood production globally. In order to promote the expansion and development of aquaculture technologies required to raise new finfish species, a number of key bottlenecks restricting commercial-scale culture need to be addressed, including the ability to control fish reproduction in captivity and to produce high quality seeds. One candidate species for large-scale production, and the focus of this work, is common snook. Prized as a food fish in Mexico, Central and South America and as a popular game fish along the Gulf coast of the United States; common snook are economically important having both a high market value and recreational demand. Despite recent advances in captive spawning, a number of reproductive bottlenecks still need to be addressed such as lack of spontaneous spawning in captivity, poor fertilization rates and inconsistent production of high quality eggs and larvae. Therefore, the overall aim of this thesis was to better understand the reproductive biology of common snook in order to develop protocols to improve the reliability of captive spawning in closed recirculating aquaculture systems and the quality of eggs produced as a basis for commercial scale cultivation. First, this PhD project described oocyte development in common snook and validated a non-invasive method for assessing reproductive condition in wild and captive stocks (Chapter 2). This was done by using a tiered and adaptable staging scheme to compare the wet mount technique with histological preparations of ovarian biopsies. When compared with histology, the wet mount provided an immediate and precise method for determining whether female broodstock were candidates for hormonal induction. In fishery biology, an understanding of fish reproductive success and population reproductive potential is critical for designing and implementing effective fisheries management strategies. The wet mount technique provides a tool for non-lethal, low-cost determination of reproductive status in wild fish stocks. The next research chapter focused on spawning induction of captive snook populations. The first trial compared the effects of slow and regular release GnRHa implants whereas the second trial investigated the effects of GnRHa, alone or in combination with the dopamine antagonist, pimozide (PIM), on milt characteristics and plasma steroid levels in captive male common snook broodstock (Chapter 3). In an effort to better enable reliable control of reproduction under captive conditions, the annual plasma sex steroid profile of captive male and female broodstock maintained under natural photo-thermal conditions was also examined. When possible, milt samples were collected pre and post implantation; sperm density, sperm motility and spermatocrit were documented among individual males. The assigned treatments appeared to have no or little effects on milt production in male broodstock although plasma steroid levels were found to be significantly elevated in individuals treated with GnRHa in combination with the dopamine antagonist, pimozide. At the time this work was performed, no data on spawning dynamics, including individual spawning performance, had been reported for common snook in captivity. Mass spawning tanks are complex systems where fish are left to spawn naturally and fertilized eggs are collected with little or no control over the mating of the animals. Therefore, the third part of this thesis explored the potential of DNA profiling for monitoring mating outcomes in captive broodstock by employing eight microsatellite markers to detect and quantify individual parental contributions for 2,154 larvae obtained from the three broodstock tanks (Chapter 4). The panel of loci was generally robust and allowed unambiguous assignment of 89% of larvae to a single family. Overall, spawn contribution data 1) provided a confirmation of GnRHa treatment efficacy in female snook with a minimum stage of oogenesis (late secondary growth-SGl) required for successful spawning, 2) identified a potential impact of handling on maturation and spawning of captive broodstock and 3) confirmed that, through photothermal conditioning, captive broodstock can spawn over consecutive days and several times per year including outside of their natural spawning season. The exogenous cues that tropical species use to synchronize key life events like reproduction remain largely unstudied, therefore, my PhD project also investigated the influence of tidal cycle on reproductive activity in common snook (Chapter 5). Real-time quantitative RT-PCR assays were developed and validated to measure the temporal expression patterns of gonadotropin genes (fshβ and lhβ) during the reproductive cycle in males and females. These were evaluated in relation to sex steroid production, LH blood plasma levels, gonadal development and tidal cycle. The phylogenetic analysis of the deduced amino acid sequence of common snook for fshβ and lhβ revealed strong identity with other teleosts (75-90%). Additionally, the mRNA profiles of fshβ and lhβ in the pituitary of females displayed a clear pattern of expression concomitant with histological changes in oocyte development. Histological observations of gonads suggested a circa-tidal rhythm of follicular development. The findings, as a whole, provided new information supporting the role of tidal cycle on the entrainment of gametogenesis allowing for a better understanding of the environmental control of reproduction in common snook. Although the primary research emphasis in this PhD was on broodstock spawning and gamete quality, the final chapter focuses on larval ontogeny. The goal of this research was to gain improve understanding of the early life history characteristics of common snook in order to improve larval culture technologies. To do so, a combination of digital photography and histological techniques were used to document the embryonic and early larval development (0 to 14 days post hatch-DPH) of hatchery-reared individuals (Chapter 6). Larvae hatched 15 h after fertilization at 28°C, lacked pigmentation, had a rudimentary digestive tract and undeveloped visual system. Development was rapid and by 3 DPH larvae had almost doubled in length, the yolk sac was nearly exhausted, the mouth was open and eyes were pigmented with a well-structured retinal layer. The alimentary canal was differentiated into three distinct sections including the foregut, midgut and hindgut. Food was observed in the gut (rotifers) and structural epithelium organelles, such as the nucleus, mitochondria, and dark vesicles, were all present in high numbers. The swim bladder was formed and inflated. In summary, understanding early ontogenetic development in common snook can help provide information needed to address key bottlenecks seen in captive cultivation, such as the high incidence of larval mortality observed during the transition from endogenous to exogenous feeding. Overall, this doctoral work 1) validated molecular and endocrine analytical tools for future studies of common snook reproductive physiology, 2) provided a better understanding of both broodfish requirements in tank systems as well as the endocrine control of reproduction and spawning at the level of the brain-pituitary-gonadal axis, 3) increased our knowledge in genetic management of captive broodstock, in terms of parentage assignment and 4) offered new insight into wild population reproductive strategy as well as how reproduction is entrained through environmental cues and the pathways leading to oocyte recruitment and maturation. The new information presented here can be used to conserve wild snook stocks through production of farm raised individuals as a sustainable source of seafood and for fisheries enhancement.

La famille des Fagacées regroupe des espèces présentant un intérêt économique, écologique et social non négligeable. Par ailleurs, ces espèces, et plus particulièrement celles du genre Quercus que l’on retrouve dans des milieux extrêmement diversifiés, constituent de bons modèles d’étude de l’adaptation des arbres à leur environnement. Pour comprendre l’architecture génétique des caractères liés à l’adaptation chez le chêne, des cartes génétiques ont été établies essentiellement sur la base de marqueurs moléculaires dominants. Le travail qui a fait l’objet de cette thèse, a consisté à développer une carte génétique de seconde génération à partir des ressources génomiques disponibles chez cette espèce. Dans un premier temps, nous avons recherché des motifs microsatellites (SSR, simple sequence repeats) au sein des séquences exprimées (EST) assemblées sous la forme d’un unigène de 28 000 éléments non redondant. Un jeu de 748 marqueurs a été développé et 255 d’entre eux ont été localisés sur la carte génétique du chêne pédonculé (Q. robur L.) en utilisant une approche dite de « bin mapping ». Leur transférabilité a été testée chez le châtaignier européen (Castanea sativa Mill.) et le hêtre commun (Fagus sylvatica L.), deux espèces phylogénétiquement proche du chêne. Un taux de transférabilité de 28% a été observé pour le hêtre et de 56,6% pour le châtaigner. Une carte génétique a alors été établie pour le châtaigner en utilisant les marqueurs SSR localisés sur la carte du chêne. La comparaison des cartes de liaison du chêne et du châtaignier a mis en évidence une bonne conservation de la macro synténie et de la macro colinéarité entre les deux espèces, ce qui ouvre des perspectives intéressantes pour le transfert d’informations génétiques (QTL par exemple) d’une espèce à l’autre. Cette étude sera prochainement enrichie par la cartographie de marqueurs orthologues dérivés de polymorphismes ponctuels (SNP), ce qui permettra de comprendre l’évolution conjointe des trois espèces majeures de la famille des Fagacées. / The Fagaceae family comprises species of economic, ecological and social importance. In addition, these species and particularly those belonging to the Quercus genus that are present in very diverse ecological niches, constitute good models to study the adaptation of forest trees to their natural environment. To understand the genetic architecture of adaptive traits in oak, genetic linkage maps have been previously established based on dominant markers. In this thesis, we developed a second generation genetic map using the genomic resources that were available in this species. First, we bioinformatically screened an expressed sequence tags catalog assembled into a 28 000 unigene elements, for simple sequence repeats (SSRs). A set of 748 markers was developed and 255 were localized on the pedunculate oak (Q. robur L.) linkage map using a bin mapping approach. Their transferability was tested in the European chestnut (Castanea sativa Mill.) and common beech (Fagus sylvatica L.), two phylogenetically related species to oak. Transferability rates of 28% and 56.6% were observed for beech and chestnut, respectively. A genetic map was then established for chestnut on the basis of orthologous SSRs already mapped in oak. The comparison between both maps clearly showed that the macro-synteny and the macro-colinearity were conserved across genus, opening interesting perspectives in respect to the transfer of genetic information (eg. QTLs, quantitative trait loci) from one species to another. This study will be soon completed by the mapping of orthologous markers derived from single nucleotide polymorphisms (SNPs). This will made it possible to better understand the evolution of the genome of these three major species of the Fagaceae family.