Gene trees enable the detailed study of the evolution of genes. These trees can be additive, based purely on mutational counts, or ultrametric, where the branches represent the time elapsed. The purpose of this thesis will be to relate evolutionary rate changes to gene function by examining the effects of a tree transformation on gene trees. The study will focus on the coffee genomes C. arabica, C. eugenioides and C. canephora. Three gene family trees will be studied: N-methyltransferases, FAD2 enzymes and terpenes. An additive tree can be transformed into an ultrametric tree using a transformation method. By taking each additive tree branch length and dividing the branch by the ultrametric branch length we can get a branch mutation rate. To quantify functional divergence, we compared aligned sequences of genes from groups of interest to calculate the ratio of non-synonymous mutations to synonymous mutations (Kn/Ks). Kn/Ks was found to correlate with the branch mutation rate in some but not all groups of interest. In groups in which correlation was present, there was also a correlation between Kn/Ks and branch length. The branch mutation rate can be used in some cases to further prove that functional divergence is present.

Phylogenetic analysis is a central tool in studies of comparative genomics. When a new region of DNA is isolated and sequenced, researchers are often forced to throw away months of computation on an existing phylogeny of homologous sequences in order to incorporate this new sequence. The previously constructed trees are often discarded, and the researcher begins the search again from scratch. The jumpstarting algorithm uses trees from the prior search as a starting point for a new phylogenetic search. This technique drastically decreases search time for large data sets. This kind of analysis is necessary as researchers analyze tree of life size data sets.

No description available.

Xenarthra form the least diversified major clade of placental mammals, being comprised of 31 described species of sloth, armadillo, and anteater. The past decade has seen a growth in the amount of xenarthran genetic data available, including the recent publication of a phylogenetic framework based on mitochondrial genomes, but more is required to aid in conservation assessments and to elucidate the evolutionary history of this unique order. We aimed to expand upon this by generating a framework based on nuclear genes. Using molecular baits, we enriched nuclear DNA from all extant and a selection of extinct ancient Xenarthrans for 74 phenotypically relevant genes. We aim to build phylogenetic trees based on each successfully enriched loci. Comparing trees against the previous mitochondrial framework will provide a measure of robustness. By seeing how estimated divergence times and rates vary across nuclear genes, we were able to examine how particular xenarthran phenotypes were selected upon in different points of their evolutionary history. While the sequencing of certain genes and species encountered mixed success rates, we have a solid framework for further study and can confirm that nuclear and mitochondrial information yields very similar evolutionary histories. / Thesis / Master of Science (MSc) / Xenarthrans - sloths, armadillos, and anteaters - have a long and complicated evolutionary history. In recent years a growth of new genetic information has made it easier to answer questions about their relation to each other and to other species. By examining many new gene sequences across all living Xenarthra, plus some extinct species, we aim to bolster our understanding of these relationships and the importance of particular traits.

Protists are defined as eukaryotic organisms that are not animals, land plants, or fungi and make up the vast majority of eukaryotic diversity. Unfortunately, they are often overlooked and under sampled. Some rationales behind this are that they are often microscopic, most rarely the cause of human disease, and are often difficult to get into culture. Molecular phylogenetics has revolutionized our understanding of the evolutionary history of eukaryotes and without a clear picture of protist evolution our understanding would be severely hindered. Phylofisher is an easy-to-use phylogenomic software package, with an accompanying database and various utilities, intended for deep phylogenomic analyses. Here I present thorough documentation for the use of PhyloFisher. I also developed a PhyloFisher utility, nucl_matrix_constructor.py, to allow the software package to build nucleotide phylogenomic matrices. This will allow users to address more shallow evolutionary questions. I developed a metabarcoding pipeline based on Oxford Nanopore Technology’s long read sequencing platform to explore the diversity of eukaryotic organisms in environmental samples. I characterized a novel discosean amoeba. Lastly, I predicted the proteomes from publicly available, but non-annotated genomes for 19 Acanthamoeba spp. and two Balamuthia spp. and used the PhyloFisher tools documented and developed to gain a better understanding into the evolutionary history of the clade as well as provide key resources in the future research of the clade.

In evolutionary biology, genetic sequences carry with them a trace of the underlying tree that describes their evolution from a common ancestral sequence. Inferring this underlying tree is challenging. We investigate some curious cases in which different methods like Maximum Parsimony, Maximum Likelihood and distance-based methods lead to different trees. Moreover, we state that in some cases, ancestral sequences can be more reliably reconstructed when some of the leaves of the tree are ignored - even if these leaves are close to the root. While all these findings show problems inherent to either the assumed model or the applied method, sometimes an inaccurate tree reconstruction is simply due to insufficient data. This is particularly problematic when a rapid divergence event occurred in the distant past. We analyze an idealized form of this problem and determine a tight lower bound on the growth rate for the sequence length required to resolve the tree (independent of any particular branch length). Finally, we investigate the problem of intermediates in the fossil record. The extent of ‘gaps’ (missing transitional stages) has been used to argue against gradual evolution from a common ancestor. We take an analytical approach and demonstrate why, under certain sampling conditions, we may not expect intermediates to be found.

The evolution of multicellularity is an example of an evolutionary transition in individuality, in which a group of lower-level biological units (cells, in this case) emerges as a higher-level unit (the multicellular organism) with its own fitness, heritability and individuality. The volvocine green algae are a model system for the transition to multicellularity and for the evolution of cellular differentiation. Some of the developmental changes that collectively make up this transition have occurred more than once in the volvocine lineage; others have reverted from derived to ancestral states. The transition from cells to multicellular organisms began over 200 million years ago in this lineage, and the subsequent changes have been sporadic, with several important changes occurring early in the transition and some body plans remaining largely unchanged over long evolutionary time scales. Two suites of characters that differ among species within the genus Volvox have each evolved convergently or in parallel in lineages that diverged at least 175 million years ago. This complex history suggests that other origins of multicellularity may have involved important roles for cooperation, conflict and conflict mediation; parallel evolution of some traits; sporadic rather than constant change; and long-term coexistence of forms with different levels of complexity. Data from one species, Pleodorina starrii, support motility as a major selective pressure driving the the origins of cellular differentiation. Optimization of the proportion of soma in this species appears to be prevented by a constraint that prevents independent change in colonies with different numbers of cells. Finally, P. starrii presents an exceptionally high level of phenotypic variability, suggesting that the genotype-phenotype map has not completely shifted from the cell to the colony and that the transition to a new, higher-level individual in this species is incomplete.

This dissertation comprises a series of studies designed to explore the associations between plants and the endophytic fungi they harbor in their above-ground tissues. By viewing endophyte diversity in ecologically and economically important hosts through the lenses of phylogenetic biology, microbiology, and biotechnology, this body of work links plant ecology with newly discovered symbiotic units comprised of endophytic fungi and the bacteria that inhabit them.This work begins with a large-scale survey of endophytic fungi from native and non-native Cupressaceae in Arizona and North Carolina. After isolating over 400 strains of endophytes, I inferred the evolutionary relationships among these fungi using both Bayesian and parsimony analyses. In addition to showing that native and introduced plants contained different endophytes, I found that the endophytes themselves harbor additional microbial symbionts, recovering members of the beta- and gamma-proteobacterial orders Burkholderiales, Xanthomonadales, and Enterobacteriales and numerous novel, previously uncultured bacteria. This work finds that phylogenetically diverse bacterial endosymbionts occur within living hyphae of multiple major lineages of ascomycetous endophytes.A focus on 29 fungal/bacterial associations revealed that bacterial and fungal phylogenies are incongruent with each other and did not reflect the phylogenetic relationships of host plants. Instead, both endophyte and bacterial assemblages were strongly structured by geography, consistent with local horizontal transmission. Endophytes could be cured of their bacterial endosymbionts using antibiotics, providing a tractable experimental system for comparisons of growth and metabolite production under varying conditions. Studies of seven focal fungal/bacterial pairs showed that bacteria could significantly alter growth of fungi at different nutrient and temperature levels in vitro, and that different members of the same bacterial lineages interact with different fungi in different ways.Focusing on one isolate, I then describe for the first time the production of indole-3-acetic acid (IAA) by a non-pathogenic, foliar endophytic fungus (Pestalotiopsis neglecta), suggesting a potential benefit to the host plant harboring this fungus. I show that this fungus is inhabited by an endohyphal bacterium (Luteibacter sp.) and demonstrate that mycelium containing this bacterium produces significantly more IAA in vitro than the fungus alone. I predict that the general biochemical pathway used by the fungal-endohyphal complex is L-tryptophan-dependent and measure effects of IAA production in vivo, focusing on root and shoot growth in tomato seedlings.

The New World Junonia butterflies include well-studied model organisms yet their phylogeny remains unresolved by traditional cox1 DNA barcodes. Sixteen Junonia mitochondrial genomes were sequenced using next generation MiSeq technology. Junonia lemonias, an Old World species, has mitochondrial genome features typical of Ditrysian Lepidoptera, and synteny is maintained throughout Junonia. Analysis of Junonia mitogenomes produced a robust phylogeny that was used with biogeographic information to infer that Junonia crossed the Pacific Ocean to invade the New World on 3 separate occasions. Junonia vestina, a high elevation species from the Andes Mountains, shows high altitude adaptation in the mitochondrial protein coding loci atp6, atp8, cox1, cob, nad1, and nad2, with the strongest effects seen in cox1 and nad1. There is some overlap between these genes with human loci that have disease associations with the same amino acid positions which could help elucidate the function of high elevation mutations in J. vestina. / February 2016

The eastern hemlock [Tsuga canadensis (L.) Carrière] is one of the native dominant forest components of northeastern US. At present, these valuable stands face an alarming decline, in part due to the Fiorinia externa, elongate hemlock scale (EHS), (Hemiptera: Coccoidea: Diaspididae). The armored shield of F. externa provides an excellent defense against insecticides, natural enemies and adverse conditions. Chemical and classical biocontrol methods have been unable to stop the spread of this pest. Recently, the occurrence of an epizootic within the F. externa population in the Mianus River Gorge Preserve in Bedford, NY revealed a promising opportunity for control of this scale. Entomopathogenic fungi represent a valuable, although under-utilized, group of organisms with unique capabilities for self-sustaining pest management. Given the significant impact of this epizootic on F. externa, we have conducted extensive research on the biology, genetics and biological control potential of this epizootic. We molecularly identified a complex of entomopathogenic, phytopathogenic, and endophytic fungi associated with the epizootic in 36 localities within the states of New York, Pennsylvania, Connecticut and New Jersey. One fungus, Colletotrichum sp., was the most commonly isolated organism in populations of F. externa within areas of the epizootic. The host range of this Colletotrichum species comprised both insects and plants, although diverse life cycles occured in the different hosts. Endophytic growth was observed in 28 species of plants comprising 18 families (52% of the sampling), whereas in F. externa biotrophic and necotrophic growth was detected. Colletotrichum is a widely known phytopathogenic genus and reports of entomopathogenic activity are extremely rare. In order to understand the biological processes involved in the host-pathogen interactions we quantified the pathogenicity and virulence of this Colletotrichum sp. to four insect families and six plants families as well as the occurrence of sexual recombination in this Colletotrichum sp., both in vitro and in planta. We observed that this Colletotrichum sp. displays a propensy to induce rapid disease and mortality in F. externa hosts. Phylogenetic analysis comprising six of the most commonly studied nuclear genes in molecular phylogenetics (D1/D2 domain of the 28 rDNA gene, ITS region, β-Tubulin 2, GPDH gene, GS gene and HMG box at the MAT1-2 mating-type gene) and RAPDs showed this fungus is closely related to phytopathogenic strains of Colletotrichum acutatum and that it may represent a single population lineage of this species (i.e., Colletotrichum acutatum forma specialis fiorinia). Though a large body of information exists regarding the phytopathogenic genus Colletotrichum, ours is only the second reported entomopathogenic strain. It is not clear whether the colonization of an insect by this fungus is truly rare or a common but undetected event. Sexual recombination, observed in planta and in vitro, could be the means by which new genetic variants are generated leading to new biotypes with a selective advantage to colonize new hosts, which in this case is a novel host in a different kingdom.