Freshwater mussel species in the genera Fusconaia and Pleurobema are particularly challenging to identify in the field. In this study, mussels from these genera were collected from the Green River, Kentucky for genetic and morphological analyses. I used molecular markers to detect any cryptic species within these genera and to test for genetic differentiation between two closely related nominal taxa P. rubrum and P. sintoxia using both mitochondrial (ND1, COI, 16S rRNA) and nuclear (ITS1) DNA sequences. After species identification, I used microsatellite DNA markers to estimate genetic diversity and effective population sizes (Ne) of species of Pleurobema. I used microsatellite primers that were developed for P. clava and P. pyriforme in previous studies, as well as microsatellites that I developed for P. plenum. Finally, I assessed morphological variation in my study species and developed dichotomous keys for the identification of both live mussels and shells. My results suggest that P. rubrum and P. sintoxia are the same species based on the mitochondrial DNA analyses, as there were few genetic differences between them. My results showed phylogenetically distinct lineages for F. flava, F. subrotunda, P. cordatum and P. plenum but no cryptic species were detected in the Green River. Current and contemporary Ne showed that these species have large population sizes that should allow for avoiding inbreeding and maintaining their evolutionary potential. Large genetic diversity as well as long-term effective population size could be the result of these species historically occurring as much larger assemblages that extended into the Ohio River and its numerous tributaries. The last objective was to assess morphometrical differences among these species. Using Canonical Variate Analysis, I found discernable morphological differences between the investigated species of Fusconaia and Pleurobema. The two Fusconaia species were morphologically different from the Pleurobema species. However, the Canonical Variate Analysis did not show differences among the Pleurobema species. I used decision tree analysis to develop a dichotomous tree, and random forest analysis was used to aid in the development of a dichotomous key by finding the most important diagnostic characters to distinguish these mussels. I then used the less subjective and easier to identify characters for the development of my dichotomous keys for live mussels and shells. However, both keys need to be tested in the field to determine their effectiveness. I could not separate P. rubrum and P. sintoxia mussels for morphometric analysis due to the lack of genetic differentiation and the inconsistent identification by the experts. However, I did describe a few individuals that look like P. rubrum and P. sintoxia to the eye of the experts. The description of these individuals matched previous descriptions of these mussels. Future studies need to assess taxonomic relationships among these species using genomics approaches, which might result in better node resolution. High genetic diversity and large effective population numbers for Pleurobema species suggest that these species' populations are genetically healthy. However, these results need to be interpreted carefully, and I therefore recommend additional studies to assess life history, habitat, host-fish availability, and current reproduction of these mussels in the Green River. / Doctor of Philosophy / Freshwater mussels offer important ecosystem services for humans to include water purification, nutrient storage and recycling, and mussels are part of the aquatic food web. In addition, freshwater mussels are indicators of ecosystem health. Because they rely on fish hosts to complete their complex life cycle, conservation of freshwater mussel species is particularly challenging. In this study, I focused my attention on freshwater mussel species commonly known as "pigtoes" which belong to the genera Fusconaia and Pleurobema. These species are difficult to distinguish morphologically even by experts. Hence, my study used molecular genetic markers to first identify these species. To assess the size and genetic health of these populations, I estimated genetic diversity and effective population number (Ne). Finally, I developed dichotomous keys to identify live mussel specimens and shells of molecularly identified mussels. My genetic results showed that there are five species of pigtoes in the Green River, which include Rough Pigtoe (Pleurobema plenum), Ohio Pigtoe (P. cordatum), Pink Pigtoe/Round Pigtoe (P. sintoxia/rubrum), Long-Solid (Fusconaia subrotunda), and Wabash Pigtoe (F. flava). A sixth pigtoe species, the endangered clubshell (Pleurobema clava), also is extant in the upper Green River but was not included in the analyses due to its rarity in the river. My results suggest that these mussel populations are large and healthy enough to survive and to adapt over time. The morphometric analysis using Canonical Variate Analysis (CVA) resulted in differentiation between F. flava and F. subrotunda. These two Fusconaia species also were morphologically different from the investigated species of Pleurobema. Results of this analysis suggest that species in the Genus Pleurobema remain difficult to differentiate from each other. The lack of genetic differentiation and the inconsistency in the experts' identification of P. rubrum and P. sintoxia, did not provide enough information to separate these two putative species for morphometric analysis. However, I was able to describe a few individuals that looked like typical examples of these two species to the eyes of the experts. Future taxonomic studies should use next-generation sequencing, which would likely result in a better resolution of evolutionary relationships. Large Ne values for each species suggest that these populations are genetically healthy. However, these results need to be considered together with results of future studies on life history, habitats, abundance, and reproduction of these species in the wild. Finally, the dichotomous mussel identification keys are intended to support more accurate identification of these species in the Green River, KY but need to be field-tested by mussel biologists

The family Xystodesmidae (Polydesmida) includes 521 species with a center of diversity concentrated in the Appalachian Mountains. Within this family, the genus Cherokia, a monotypic taxon with the type species Cherokia georgiana, is divided into three subspecies. The last revision of this genus was made by Richard Hoffman in 1960. Here, I used morphological and molecular data sets to review the genus, and evaluate whether it is a monophyletic group. I included material from literature records and three natural history collections. Newly collected samples were obtained through a citizen science project. Morphological characters such as the shape of the paranota, body size, and coloration were evaluated. Seven gene loci were used to estimate a molecular phylogeny of the genus, and a species delimitation analysis was used to evaluate the status of the subspecies. The geographical range of Cherokia was expanded to include a newly reported state (Virginia) and ca. 160 new localities compared to the previously known range. Morphological characters such as the shape of the paranota and body size that were historically used to establish subspecies, showed a direct relation with geographical distribution and elevation (clinal variation), but not with the phylogeny. Coloration was variable and did not accord with geography or phylogeny. The phylogeny recovered a monophyletic lineage, and the species delimitation test supports a single species. The molecular and morphological evidence showed that Cherokia is a monotypic genus with the sole species Cherokia georgiana being geographically widespread and highly variable in its morphology / Master of Science in Life Sciences / Millipedes are a mega-diverse group of soil dwelling animals that feed on leaf litter. The Appalachian Mountains has a huge diversity of millipedes, in particular those in the family Xystodesmidae. Within this family, I studied the genus Cherokia, commonly known as "Georgia flat-backed millipedes". The single species in this group, Cherokia georgiana, is divided into three subspecies. The last thorough study of this genus was done by Richard Hoffman in 1960, so a modern analysis with DNA sequencing was needed to test subspecies boundaries. Here, I used hundreds of specimens from three natural history museums, and fresh specimens obtained for DNA sequencing with the help of citizen scientists. I measured the shape and size of the body and coloration patterns to determine if they were related to the geographical distribution of Cherokia. I used DNA sequencing to make an evolutionary tree of the genus. I found Cherokia individuals in Virginia for the first time and found ca. 160 new sites or locations not reported previously. The shape and size of the body was related to millipede location and elevation. Coloration was not related to geography or phylogeny, and in some localities, multiple color patterns co-existed. The genetic information from DNA sequencing indicated that all Cherokia were more closely related to each other than to any other millipede genus. In conclusion, I found that the genus Cherokia is a single species, Cherokia georgiana, that has a wide geographical distribution and a considerable diversity of body shape and color. Diversity of shape and color does not reflect subspecies boundaries but instead reflects intra-population and geographic variation.

Background Bartonella bacilliformis is the etiological agent of Carrion’s disease, a neglected tropical poverty-linked illness. This infection is endemic of Andean regions and it is estimated that approximately 1.7 million of South Americans are at risk. This bacterium is a fastidious slow growing microorganism, which is difficult and cumbersome to isolate from clinical sources, thereby hindering the availability of phylogenetic relationship of clinical samples. The aim of this study was to perform Multi Locus Sequence Typing of B. bacilliformis directly in blood from patients diagnosed with Oroya fever during an outbreak in Northern Peru. Methodology/Principal Findings DNA extracted among blood samples from patients diagnosed with Oroya’s fever were analyzed with MLST, with the amplification of 7 genetic loci (ftsZ, flaA, ribC, rnpB, rpoB, bvrR and groEL) and a phylogenetic analysis of the different Sequence Types (ST) was performed. A total of 4 different ST were identified. The most frequently found was ST1 present in 66% of samples. Additionally, two samples presented a new allelic profile, belonging to new STs (ST 9 and ST 10), which were closely related to ST1. Conclusions/Significance The present data demonstrate that B. bacilliformis MLST studies may be possible directly from blood samples, being a promising approach for epidemiological studies. During the outbreak the STs of B. bacilliformis were found to be heterogeneous, albeit closely related, probably reflecting the evolution from a common ancestor colonizing the area. Additional studies including new samples and areas are needed, in order to obtain better knowledge of phylogenetic scenario B. bacilliformis

Herein we tested the repeatability of RAD-seq phylogeographic construction by creating a second, independent phylogeography of the pitcher-plant mosquito, Wyeomyia smithii. We sampled 25 populations drawn from different localities nearby previous collection sites and used these new data to construct a second, independent phylogeography to test the reproducibility of phylogenetic patterns. Our previous phylogeography was based on 3,741 phylogenetically informative markers from 21 populations and rooted with mitochondrial COI. The present phylogeography was based on 16,858 informative markers and rooted with RAD-seq. We found correspondence between clades at the extremes of W. smithii's distribution; however, there were several discrepancies between the trees, including the refugium that gave rise to all post-glacial populations. We observed that combining all 46 populations resolved these discrepancies and, equally importantly, that extensive taxon sampling in areas of historical importance is more valuable than increasing the number of informative sites in establishing an accurate, robust phylogeography. This thesis includes unpublished co-authored material.

The phylogeny of Eumalacostraca (Crustacea) has remained elusive, despite over a century of morphological and more recently molecular studies. Prior to this, no large scale combined evidence analyses of all eumalacostracan orders has been carried out. Evidence from four nuclear ribosomal and mitochondrial loci (18S rRNA, 28S rRNA, 16S rRNA, and cytochrome c oxidase subunit I) were combined with a newly synthesized morphological dataset to examine the utility of available data. With an aim to resolve conflict in the morphological dataset, fossils were added to the morphological dataset and their effects examined. The stratigraphic congruence of crustaceans was examined to determine the quality of existing trees. A modified protocol was developed to generate new molecular markers for the investigation of eumalacostracan phylogeny. Finally the effects of the new data on existing datasets were examined in a combined evidence approach. Significant conflict was detected between data partitions, especially between morphology and molecules. The addition of fossil data revealed the morphological dataset to be very sensitive to taxon sampling. Stratigraphic congruence was found to be poor for the five crustacean groups examined. Histone 3 and alanyl-tRNA synthetase were shown to be capable of successfully recovering relationships at the level of genus and above. Glutamyl-prolyl-tRNA synthetase failed to recover monophyletic groupings when analysed alone. Combining all molecular data produced well-supported phylogenies, but significant conflict between data partitions was identified and trees were very sensitive to taxon sampling. Rate heterogeneity and conflict between data partitions mean that the total sum of molecular and morphological evidence as presented here is currently unable to resolve a wellsupported eumalacostracan phylogeny. We recommend additional taxon sampling and further sampling of novel markers using the modified protocol developed here as well as the addition of fossil data and the exploitation of next generation sequencing technologies.

Liverworts (Phylum Marchantiophyta) form the earliest diverging lineage of extant land plants and hold a key position in the evolutionary history of land plants. The family Cleveaceae, first defined in 1881 by Leitgeb by its complex thallus anatomy and star-shaped epidermal pores with strongly thickened radial walls, is now included within Marchantiopsida (Complex Thalloid Liverworts). The family is widely distributed in warm temperate to arctic regions throughout the Northern Hemisphere, but very rare in South America and absent from Australia and New Zealand. Since Cleveaceae was described, the number and names of genera and species recognized within the family have changed many times, with three genera currently accepted: Athalamia (12 to 15 species), Sauteria (2 to 5 species) and Peltolepis (1 or 2 species). However, no robust phylogenetic frame work or taxonomic monographs of the family have previously been undertaken and the species diversity within the genera remains uncertain. The study presents the first robust phylogenetic framework for the family, based on three chloroplast genes (psbA, rpoC1 and rps4) and one nuclear ribosomal region (26S) sequenced for 54 ingroup accessions representing a substantial part of the geographic range of the family (except South America) and a large part of its morphological diversity. Samples sequenced include the type species of the three currently accepted genera as well as those of previously synonymised genera; Clevea, Spathysia, Gollaniella and Sauchia. The current circumscription and relationships of the genera were tested using Maximum Parsimony, Bayesian and Likelihood inferences. Cleveaceae was strongly supported as monophyletic with four main lineages resolved within the family, two corresponding to Peltolepis and Sauteria and two composing a polyphyletic Athalamia. The latter is therefore divided into a more narrowly defined Athalamia and a fourth genus, for which Clevea is the earliest published name. Inferred evolutions of twelve morphological characters, that are useful or have been used in classifications of Cleveaceae, were reconstructed using maximum parsimony and maximum likelihood methods. The misleading influence of homoplasy on past morphological classifications and phylogenies is discussed. The study also presents the first worldwide taxonomic revision of Cleveaceae based on around 2300 specimens studied, including all available type specimens. The monographic work is based on morphological and anatomical characters reviewed using dissecting, compound and scanning electron microscopes including both herbarium and fresh material collected on field excursions during the study. Ventral scale morphology, position and aggregation of androecia and shape of receptacle were found to be valuable characters in generic and species delimitation. The traditional definition of genera of Cleveaceae based mainly on two characters (carpocephalum stalk position and anatomy) was rejected. A total of seven species were recognized from 25 formerly described names including one new necessary combination: Clevea hyalina (Sommerf.) Lindb., C. spathysii (Lindenb.) Müll. Frib., C. pusilla (Steph.) Rubasinghe & D.G.Long comb. nov., Athalamia pinguis Falc., Sauteria alpina (Nees) Nees, S. spongiosa (Kashyap) S.Hatt., and Peltolepis quadrata (Saut.) Müll. Frib. Nine excluded and doubtful species are listed along with reasons for their exclusion. Generic and species keys are presented based on vegetative, reproductive and spore morphological characters. For each genus and species, descriptions, illustrations, and distribution maps are given along with ecological, taxonomic and nomenclatural notes.

Senna comprises c.300-350 species and is one of the most species-rich and widely distributed genera in the family Leguminosae (Fabaceae). It is most diverse in tropical America, with secondary centres of diversity in tropical Africa, Madagascar and Australia. Only a few species are found in South Eastern Asia and in the Pacific Islands. Several Senna species are used as purgatives or laxatives in medicine, and some species are cultivated for their attractive flowers and foliage. The taxonomic revision of Senna species from Arabia presented here is based principally on herbarium specimens from the Royal Botanic Garden Edinburgh and the Royal Botanic Gardens, Kew and on field observations made in the Kingdom of Saudi Arabia, the Sultanate of Oman and the Republic of Yemen. These materials and observations also formed the basis for a review of the morphological features of the Senna species in Arabia, which was undertaken in the context of the morphological variability of the whole genus. The taxonomic revision recognises nine species that occur in Arabia: Senna alexandrina Mill., S. didymobotrya (Fresen.) H.S. Irwin & Barneby, S. holosericea (Fresen.) Greuter, S. hookeriana Batka, S. italica Mill., S. obtusifolia (L.) H.S. Irwin & Barneby, S. occidentalis (L.) Link, S. sophera (L.) Roxb. and S. tora (L.) Roxb. The study demonstrated that a putative tenth species, S. socotrana (Serrato) Lock, previously thought to be endemic to the region, is morphologically indistinguishable from the widespread Senna sophera. Consequently the study presents S. socotrana (Serrato) Lock as a syn. nov. Four taxa, S. alata (L.) Roxb., S. artemisioides (DC.) Randell subsp. artemisioides, S. auriculata Roxb., and S. multiglandulosa (Jacq.) H.S. Irwin & Barneby, are recorded as or cultivated in the Arabian Peninsula. A phylogeny is presented for the genus Senna, including all nine Arabian species, based upon the chloroplast DNA regions matK and rps16 and analysed in a parsimony and Bayesian framework. Sampling new to this study included 118 accessions of Arabian species representing substantial parts of their geographic range within the Arabian Peninsula. Sequence data newly generated from a further 13 accessions were added to 104 generated in previous studies bringing the number of accessions representing other Senna species from across the range of the genus to 117. The phylogeny is useful in a taxonomic context and is also suggestive of the phylogenetic utility of some features, especially the morphology of the pods. Reconstruction of the morphological diversification of extrafloral nectaries in the context of the phylogeny suggests that it may have been more complex than outlined in previously published work. The phylogeny shows Senna species found in the Arabian Peninsula are placed in three separate clades, suggesting that Senna arrived in the Arabian Peninsula at least three times independently. Dating the phylogeny using a relaxed molecular clock in a Bayesian approach indicates that these arrivals happened at different times over the past c. 20 Mya, suggesting that a common geological explanation is unlikely and that dispersal must have played a key role in developing these biogeographic patterns.

Borealosuchus is a significant genus as it is considered either a basal crocodylian or close outgroup. A new species identified from the early Campanian Mooreville Chalk of Alabama is one of the oldest known crocodylians and extends the stratigraphic range of Borealosuchus from the Maastrichtian minimally into the early Campanian. It co-occurs with the giant alligatoroid Deinosuchus in deposits formed in a marginal marine setting with fluvial input. Its age, along with the phylogenetic position of Borealosuchus, renders this material critical for understanding the morphological conditions and relationships at the root of Crocodylia. The most complete specimen consists of partial cranial, mandibular, and postcranial remains preserving a unique combination of characters including a short dentary symphysis, splenial participation in the symphysis, lack of discrete concavity on the angular dorsal margin for an external mandibular fenestra, and bipartite ventral osteoderms. The external mandibular fenestra was either absent or very small, raising questions about the ancestral condition for this structure in Crocodylia. The new species is, surprisingly, closer phylogenetically to more derived species of Borealosuchus, including B. wilsoni, B. threeensis, and B. acutidentatus, than to B. sternbergii and B. formidabilis. This creates many range extensions and ghost lineages in the clade and suggests substantial unsampled diversity within Borealosuchus. Additional mandibular material from the Santonian Eutaw Formation may be referable to this new species, further increasing its stratigraphic range. A specimen consisting of a partial post-cranial skeleton from the Mooreville Chalk is significantly larger than the holotype of the new species, and is referable to Borealosuchus sp., but is too incomplete for more precise referral. These newly described specimens provide additional information about the faunal assemblages of Alabama during the Late Cretaceous and have broader implications because they are some of the earliest crocodylian material known. Indeed, the Eutaw material appears to be the oldest known occurrence of Crocodylia.

Evolutionary transitions between marine and freshwater habitats are rare events that can have profound impacts on aquatic biodiversity. The main goal of my thesis is determining the processes involved in transitions between marine and freshwater biomes, and the resulting patterns of diversity using phylogenetic approaches. To test hypotheses regarding the geography, timing, frequency, and mechanisms regulating biome transitions I generated multi-locus time-calibrated molecular phylogenies for groups of fishes that include both exclusively marine and freshwater species. My analysis of anchovies demonstrated that Neotropical freshwater anchovies represent a monophyletic radiation with a single origin in South American freshwaters. I used a phylogeny of herring and allies (Clupeiformes) to investigate the evolution of diadromy, a migratory behavior in which individuals move between oceans and freshwater habitats for reproduction and feeding. These analyses do not support the hypothesis that differences in productivity between marine and freshwater explain the origins of diadromous lineages. Diadromy has been considered an evolutionary pathway for permanent biome transitions, however I found that diadromy almost never produces a fully marine or freshwater clade. Marine lineages often invade continental freshwaters during episodes of marine incursion. In South America, the rich diversity of marine derived fish lineages invaded during Eocene marine incursions from either the Pacific or the Caribbean, and Oligocene marine incursions from the Caribbean. I falsified the highly cited Miocene marine incursion hypothesis, but found that the Pebas Mega-Wetland catalyzed diversification in some marine derived lineages. Using diversification analyses, I investigated the evolutionary processes that have generated disparate patterns of diversity between continents and oceans. I found that freshwater silversides have higher speciation and extinction rates than marine silversides. Lineages accumulation plots suggest ecological limits are not regulating clade growth in either marine or freshwater biomes. Overall, biome conservatism is a widespread pattern among fishes, and this pattern is largely driven by competition in clades that are physiologically capable of biome transitions. Biome transitions are facilitated by rare paleogeographic events, such as marine incursions. Finally, a difference in net diversification rate is the macroevolutionary mechanism that best explains the difference in diversity between continents and oceans.

The genus Fucus (Phaeophyceae) has nine commonly accepted species, six of which inhabit North American shores: F. gardneri, F. serratus, F. distichus, F. evanescens, F. vesiculosus, and F. spiralis. Fucus inhabit the intertidal zones of the Atlantic, Pacific and Arctic Oceans and due to their highly plastic and morphologically simple phenotypes, are valuable candidates for molecular phylogenetic research. Furthermore, Fucus species are valuable for biogeographic investigations due to their relatively slow migration, limited dispersal range, and easily collected distribution. The closest ancestral species to the genus is a Pacific endemic Hesperophycus californicus supporting a North Pacific origin of the species (Serrão et al. 1999). However, a closer relationship between Atlantic and Arctic Fucus has been described casting doubt on the Pacific origins of the genus (Lindstrom 2001, Dunton 1992). The investigation of Fucus across this area is necessary to elucidate the apparent contradiction in the origin of the genus. Samples were collected from 79 locations across North American waters. The amplification of the divergent mtDNA spacer region was performed to assess taxonomic placement of 55 collected species and reconstruct the biogeography of North American Fucus species. Morphological analysis was also completed based on 21 measured characteristics in an attempt to support molecular analysis. Results of the study indicate two distinct lineages among collected samples. One lineage (F. spiralis and F. vesiculosus) with a distinct Atlantic origin and the second lineage (F. distichus and F. serratus) with a widely distributed F. distichus, and the Atlantic F. serratus. Morphological analysis and statistical support for these lineages was determined through discriminant analysis of the collected samples. Overall results determined biogeographic influences in the F. distichus species complex across Arctic, Pacific, and Atlantic regions with little apparent biogeographical influence within F. vesiculosus, F. serratus and F. spiralis. Closer phylogenetic relationships between Arctic and Atlantic samples were apparent as suggested by Lindstrom (2001) and Dunton (1992).