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Rhododendron is a monophyletic group that inhabits many different climates. One clearly defined diversification was from temperate ancestors into tropical habitats. The focus of this work was to explore leaf and stem anatomical traits in relation to habitat (temperate and tropical) and elevation of the native range. A closely-related group of Rhododendron was selected to reduce variation in genetic history and reveal environment–associated adaptive traits. Vessel anatomical traits of Rhododendron accessions were assayed for the trade of between safety (protection against catastrophic failure) and efficiency (high theoretical conductivity). Rhododendron wood and vessels were found to be relatively safe. The metrics of wood efficiency were higher for the tropical species. Thus, a trade-off between safety and efficiency was found although the wood of Rhododendron is characterized as highly safe. Leaf anatomical traits of Rhododendron were assayed for habitat and elevation. Leaves on tropical species were thicker and denser compared with temperate species. Idioblasts were always found in tropical leaves but not in temperate species. Leaves of tropical species were more xeromorphic (drought tolerant) than those of temperate species. Increasing elevation of the native range did not influence leaf anatomical traits. Idioblast abundance and leaf water relations traits were assayed for tropical Rhododendron species. Idioblast expression varied from 5% to 28% and stomatal pore index varied from 0.08 to 3.3. Idioblast expression was highly correlated with leaf succulence, and water deficit at the turgor loss point. Idioblast expression was positively associated with leaf capacitance for thin (< 0.5 mm) leaves. Thus, idioblasts can serve as a water buffer for relatively thin leaves. Synthesis–Wood traits of evergreen Rhododendron shrubs reflect adaptation for safety. Although tropical species have significantly higher efficiency, wood safety is still the dominant feature. The implication of high wood safety is constrained water flow and a potential for low water potential. Both leaf succulence and the presence of idioblasts in thin leaves enhances leaf capacitance and provides some buffering against short-term drought. These leaf adaptations in tropical Rhododendron shrubs likely reflect the abundance of epiphytes in this group. Ph. D.

The study focuses on the genus Bothriochloa (Andropogoneae, Poaceae) in Australia. Despite morphological features separating this genus from the closely related two genera Capillipedium and Dichanthium, (the three hereafter will be called BCD), De Wet and Harlan introduced the compilospecies complex to show the interbreeding phenomena that occurred among species of these genera. This study was carried out to assess species/genus relatedness of the BCD complex using different evidences from morphology, molecular information and genomic content. Nineteen morphological characters were observed, three regions (trnT-F, rps16 intron and 3'trnK) of chloroplast genome phylogenetic were used in phylogenetic reconstruction, and chromosome counting as well as flow cytometry for chromosome number and genome size were conducted during the study. Phylogenetic trees were constructed using MP with NJ for morphological data, and MP, RAxML, and BI for molecular data. Based on morphology, all three genera were separated as monophyletic units. Bothriochloa consisted of two clades. However, phylogenetic analyses based on chloroplast genomic regions reveal that Bothriochloa and Dichanthium are paraphyletic clades and only Capillipedium is resolved as a monophyletic clade. The concatenated data set has performed better than individual data sets in terms of resolution and support for clades. Flow-cytometry and chromosome counting only found diploid and tetraploid but not hexaplod species. TCS network reveals that tetraploidization followed different pathways from the ancestral diploid species. This study provided new insight onto the evolution of the chloroplast genome in the compilospecies and empirical evidence of species grouping of the compilospecies based on morphology. Master of Science

Canary grasses (Phalaris, Poaceae) include 21 species widely distributed throughout temperate and subtropical regions of the world with centers of diversity in the Mediterranean Basin and western North America. The genus contains annual/perennial, endemic/cosmopolitan, wild, and invasive species with basic numbers of x=6 (diploid) and x=7 (diploid/tetraploid/hexaploid). The latter display vastly greater speciation and geographic distribution. These attributes make Phalaris an ideal platform to study species diversification, dispersal, historic hybridization, polyploidy events, and chromosome evolution in the grasses. This body of research presents the first molecular phylogenetic and phylogeographic reconstruction of the genus based on the nuclear ITS and plastid trnT-F DNA regions allowing species relationships and the importance of polyploidy in speciation to be assessed. Divergence dates for the genus were determined using Bayesian methods (BEAST, version 1.6.2) and historic patterns of dispersal were analyzed with RASP (version 2.1b). Self-incompatibility and the feasibility of hybridization between major groups within the genus were studied with a series of greenhouse experiments. Acetocarmine and fluorescent staining techniques were used to study the morphology of the chromosomes in a phylogenetic context and the nuclear DNA content (C values) was quantified using flow cytometry. Four major clades were revealed in the genus with cytological and geographic affinities leading to the establishment of two subgenera and four sections in the first comprehensive infrageneric treatment of Phalaris. Divergence dating revealed a Miocene emergence (20.6-8.4 MYA) for the genus which is concurrent with studies of other genera in the Aveneae tribe. The hypothesis stating that Phalaris originated in the Mediterranean Basin and dispersed to the New World via a western route leading to a secondary center of diversification in western North America was supported by phylogeographic and cytological analyses. An empirical study comparing the weight, length, and width of the florets by morphological type and cytotype revealed significant differences that support a dispersal advantage among the New World and Arundinacea species. The x=6 species displayed greater intraspecific C value variation, higher DNA content per haploid chromosome set, and a distinct karyotype compared with the x=7 species indicating a complex history of chromosome evolution. Ph. D.

Plants have evolved a variety of constitutive and induced chemical defense mechanisms against biotic stress. Emission of volatile compounds from plants facilitates interactions with both beneficial and pathogenic organisms. However, knowledge of the chemical defense in roots is still limited. In this study, we have examined the root-specific biosynthesis and function of volatile terpenes in the model plant Arabidopsis. When infected with the root rot pathogen Pythium irregulare, Arabidopsis roots release the acyclic C11-homoterpene (E)-4,8-dimethylnona-1,3,7-triene (DMNT), which is a common constituent of volatile blends emitted from insect-damaged foliage. We have identified a single cytochrome P450 monooxygenase of the CYP705 family that catalyzes a root-specific oxidative degradation of the C30-triterpene precursor arabidiol thereby causing the release of DMNT and a C19-degradation product named arabidonol. We found that DMNT shows inhibitory effects on P. irregulare mycelium growth and oospore germination in vitro, and that DMNT biosynthetic mutant plants were more susceptible to P. irregulare infection. We provide evidence based on genome synteny and phylogenetic analysis that the arabidiol biosynthetic gene cluster containing the arabidiol synthase (ABDS) and CYP705A1 genes possibly emerged via local gene duplication followed by de novo neofunctionalization. Together, our studies demonstrate differences and plasticity in the metabolic organization and function of terpenes in roots in comparison to aboveground plant tissues. Additionally, we demonstrated that the arabidiol cleavage product, arabidonol, is further modified by yet unknown enzymatic reactions into three products, which are found in root exudates. We suggested a pathway for their biosynthesis based on precursor feeding experiments and NMR analysis. Although DMNT biosynthetic genes are clustered on chromosome 4 along with several potential modification genes, we did not find a possible role of these genes in the derivatization of arabidonol. Preliminary experimental results using genetic and biochemical approaches for identifying genes involved in the modification steps are also presented. In summary, this study demonstrates an alternative route for volatile terpene formation belowground different from aboveground plant tissues via triterpene degradation and provides evidence for an unexplored triterpene catabolism pathway in Arabidopsis. Ph. D.

The recent call to reconstruct a detailed picture of the tree of life for all organisms has forever changed the field of molecular phylogenetics. Sequencing technology has improved to the point that scientics can now routinely sequence complete plastid/mitochondrial genomes and thus, vast amounts of data can be used to reconstruct phylogenies. These data are accumulating in DNA sequence repositories, such as GenBank, where everyone can benefit from the vast growth of information. The trend of generating genomic-region rich datasets has far outpaced the expasion of datasets by sampling a broader array of taxa. We show here that expanding a dataset both by increasing genomic regions and species sampled using GenBank data, despite the inherent missing DNA that comes with GenBank data, can provide a robust phylogeny for the plant order Caryophyllales (angiosperms). We also investigate the utility of trnK intron in phylogeny reconstruction at relativley deep evolutionary history (the caryophyllid order) by comparing it with rapidly evolving matK. We show that trnK intron is comparable to matK in terms of the proportion of variable sites, parsimony informative sites, the distribution of those sites among rate classes, and phylogenetic informativness across the history of the order. This is especailly useful since trnK intron is often sequenced concurrently with matK which saves on time and resources by increasing the phylogenetic utility of a single genomic region (rapidly evolving matK/trnK). Finally, we show that the inclusion of RNA edited sites in datasets for phylogeny reconstruction did not appear to impact resolution or support in the Gnetales indicating that edited sites in such low proportions do not need to be a consideration when building datasets. We also propose an alternate start codon for matK in Ephedra based on the presense of a 38 base pair indel in several species that otherwise result in pre-mature stop codons, and present 20 RNA edited sites in two Zamiaceae and three Pinaceae species. Ph. D.

Peanuts (A. hypogaea) are an economically important crop, a source of food allergies and a member of the South American genus Arachis. The eighty species of genus Arachis have been divided into nine sections. The largest, section Arachis, has been further subdivided into three genome groups. The current intuitive understanding of the evolutionary relationships among Arachis is based on morphological, geographic and cytogenetic data, but a comprehensive phylogenetic study for the genus is lacking. A total of 48 species representing all nine sections were used to reconstruct a phylogeny based on sequence information from plastid trnT-trnF and nuclear ITS genomic regions. Phylogenetic analysis resolved section Extranervosae at the base, followed by sections Triseminatae and Caulorrhizae. Two major terminal lineages were recovered. One is comprised of sections Erectoides, Heteranthae, Procumbentes, Rhizomatosae, and Trierectoides, referred to here as group erectoides. The other is comprised of two major clades, arachis I (B genome, D genome, and aneuploid species) and arachis II (A genome species). The phylogenetic trees show that sequence data partially agrees with the relationships described in the monograph; however, some further investigation is necessary to clarify relationships within and among species of the two terminal lineages. In addition, the major allergen Ara h 2 from 12 wild species from across the genus was analyzed for mutations that could potentially produce a hypoallergenic ortholog. It was found that the evolution of the allergen mostly reflected the species phylogenies based on ITS and combined. The majority of substitutions and length variations were concentrated in the loop connecting helices H2 and H3. Section Arachis species tended to have larger H2-H3 loops, while those from other sections had shorter loops. The immunodominant epitopes #6 and #7, located within this loop, tended to contain mutations or were truncated among species outside of section Arachis. Dot immunoblots showed reduced IgE-binding to peptides representing portions of the H2-H3 loop from A. guarantica and A. triseminata. Orthologs from wild species have demonstrated that they could potentially contain variations of the allergen Ara h 2 that could be utilized to develop a safer peanut cultivar. Ph. D.

Plants as sessile organisms are susceptible to many environmental stresses such as drought, and salinity. They have therefore evolved mechanisms to acclimate and tolerate environmental stresses. Knowledge of the molecular aspects of abiotic stress gleaned from extensive studies in Arabidopsis has provided much information on the complex processes underlying plant response to abiotic stresses. Nevertheless, there is a need for integration of the knowledge gained and a systematic molecular genetic dissection of the complex responses to abiotic stress. In this study in Arabidopsis, comparative expression profiling analysis of progressive (pDr) and moderate (mDr) drought treatments revealed common drought responses, as well as treatment specific signatures responses to drought stress. Under prolonged moderate drought plants develop different mechanisms for acclimation: induction of cell wall loosening at early stage, and a change in hormonal balance (ABA: JA) at late stage of moderate drought. Taking a reverse genetics approach, a MYB transcription factor (MYB109) has been identified as a regulator of growth under drought and salt stress. Global expression profiling showed possible mechanisms of how MYB109 modulates growth under drought conditions: as a regulator of RNA processing and splicing and as a negative regulator of jasmonic acid biosynthesis and signaling. A forward genetics screen for drought and salt tolerance of transposon activation tag (ATag) lines led to the discovery of novel genes, which shed light on unexplored areas of abiotic stress biology. Utilizing this strategy, a potential role for cell wall modification and MATE transporters in response to drought and salt stress has been discovered, which needs further analysis to integrate this information on the role of these biological processes in plant stress biology. Ph. D.

Aim: Fatigue during team sports requiring multiple sprints can result from the combined effects of metabolic, mechanical, neurological, and immune factors. The purpose of this study was to investigate the influence of capsaicin on performance of and the physiological response to an exercise test simulating the fitness demands of team sport game conditions. Methods: This study was a placebo-controlled, crossover design. Nineteen healthy male experienced athletes age 18-30 yr consumed either 3 g/d cayenne (25.8 mg/d capsaicin) or placebo for 1 wk. Directly following the supplementation period, they completed a repeated sprint test consisting of 15 30 m maximal effort sprints on 35 s intervals. Sprint times were recorded via electronic dual-beam timing system. Fasted blood draws for interleukin-6 (IL-6) were taken at baseline prior to supplementation, 45-min pretest, and immediately post test. Heart rate (HR), blood pressure (BP), rate of perceived exertion (RPE), muscle soreness (MS), and gastrointestinal distress (GD) were measured 1-min pretest, during, posttest, and 1-min posttest. MS was also measured for 3 d posttest. Results: Relative to the placebo, capsaicin significantly reduced maximum HR by 9.3%, total average HR by 8.5%, and sprinting average HR by 6.0% (P

The chloroplast matK gene has been identified as a rapidly evolving gene at nucleotide and corresponding amino acid levels. The high number of nucleotide substitutions and length mutations in matK has provided a strong phylogenetic signal for resolving plant phylogenies at various taxonomic levels. However, these same features have raised questions as to whether matK produces a functional protein product. matK is the only proposed chloroplast-encoded group II intron maturase. There are 15 genes in the chloroplast that would require a maturase for RNA splicing. Six of these genes have introns that are not excised by a nuclear imported maturase, leaving MatK as the only candidate for processing introns in these genes. Very little research has been conducted concerning the expression and function of this important gene and its protein product. It has become crucial to understand matK expression in light of its significance in RNA processing and plant systematics. In this study, we examined the expression, function and evolution of MatK using a combination of molecular and genetic methods. Our findings indicate that matK RNA and protein is expressed in a variety of plant species, and expression of MatK protein is regulated by development. In addition, matK RNA levels are affected by light. Furthermore, genetic analysis has revealed that although MatK has a high rate of amino acid substitution, these substitutions are not random but are constrained to maintain overall chemical structure and stability in this protein. We have also identified an alternate start codon for matK in some plant species that buffers indels (insertions and deletions) in the open reading frame (ORF) that are not in multiples of three in the gene sequence. Usually, indels not in multiples of three result in frame shifts that destroy the reading frame. Our results indicate that an out-of-frame matK start codon in some orchids compensates for these otherwise deleterious indels. This research represents the first in-depth analysis of matK gene expression and contributes to several fields of biology including plant systematics, genetics and gene expression. Ph. D.

Comparative analysis of multiple genomes presents many challenges ranging from management of information about thousands of local similarities to definition of features by combination of evidence from multiple analyses and experiments. This research represents the development stage of a database-backed pipeline for comparative analysis of multiple genomes. The genomes of three recently sequenced species of Brucella were compared and a superset of known and hypothetical coding sequences was identified to be used in design of a discriminatory genomic cDNA array for comparative functional genomics experiments. Comparisons were made of coding regions from the public, annotated sequence of B. melitensis (GenBank) to the annotated sequence of B. suis (TIGR) and to the newly-sequenced B. abortus (personal communication, S. Halling, National Animal Disease Center, USDA). A systematic approach to analysis of multiple genome sequences is described including a data model for storage of defined features is presented along with necessary descriptive information such as input parameters and scores from the methods used to define features. A collection of adjacency relationships between features is also stored, creating a unified database that can be mined for patterns of features which repeat among or within genomes. The biological utility of the data model was demonstrated by a detailed analysis of the multiple genome comparison used to create the sample data set. This examination of genetic differences between three Brucella species with different virulence patterns and host preferences enabled investigation of the genomic basis of virulence. In the B. suis genome, seventy-one differentiating genes were found, including a contiguous 17.6 kb region unique to the species. Although only one unique species-specific gene was identified in the B. melitensis genome and none in the B. abortus genome, seventy-nine differentiating genes were found to be present in only two of the three Brucella species. These differentiating features may be significant in explaining differences in virulence or host specificity. RT-PCR analysis was performed to determine whether these genes are transcribed in vitro. Detailed comparisons were performed on a putative B. suis pathogenicity island (PAI). An overview of these genomic differences and discussion of their significance in the context of host preference and virulence is presented. Master of Science

Chapter One: Raven's 1985 phylogenetic analysis of the Mygalomorphae placed a number of previously unrelated genera into the rastelloid family Cyrtaucheniidae. Although Goloboff's 1993 reanalysis of mygalomorph relationships retained the familial composition of the Rastelloidina it di not support cyrtaucheniid monophyly. This study resolves the issue of cyrtaucheniid monophyly within the context of the Rastelloidina. Using 71 morphological characters scored for 29 mygalomorph taxa we find that the Cyrtaucheniidae is polyphyletic and propose the following families in its place: Cyrtaucheniidae, Kiamidae (new family), Aporoptychidae (new rank), Ancylotrypidae (new family) and Euctenizidae (new rank). We also propose two new euctenizid genera, Apachella and Sinepedica, revise the taxonomy of the euctenizids of the Southwestern United States, and present a key for these six genera. In addition to the morphologically based phylogeny we test and refine the euctenizid intergeneric phylogeny using molecular data (mitochondrial 16S rRNA and COI genes and 28S rRNA nuclear genes). The results of the combined morphological and molecular analysis are used to construct a composite rastelloid phylogeny that is used to investigate biogeographical relationships, burrow entrance evolution, and homoplasy. Chapter Two: This systematic study of the predominately Californian trapdoor spider genus Aptostichus Simon, 1890 describes 28 species, 25 of which are newly described: A. atomus, A. improbulus, A. insulanus, A. icenoglei, A. ebriosus, A. muiri, A. cahuillus, A. luiseni, A. serranos, A. calientus, A. chemehuevi, A. shoshonei, A. pauitei, A. tipai, A. cochesensis, A. indegina, A. gertschi, A. kristenae, A. fornax, A. spinaserratus, A. brevifolius, A. brevispinus, A. agracilapandus, A. tenuis, and A. gracilapandus. Aptostichus stanfordianus Smith, 1908 is considered to be a junior synonym of A. atomarius Simon 1890. Using 72 quantitative and qualitative morphological characters we propose a preliminary phylogeny for this group. Based on the results of this phylogenetic analysis, we recognize the Atomarius, Simus, Hesperus and Pandus species groups. Additionally, our phylogenetic analysis indicates that adaptations favoring the invasion of the very arid desert habitats of southern California have evolved multiple times in the Aptostichus clade. The existence of both desert and non - desert species in three of the four species groups makes this genus an ideal candidate for the study of the evolutionary ecology of desert arthropods. Chapter Three: Aptostichus simus is a trapdoor spider that is endemic to the coastal dunes of southern California and is recognized as a single species on morphological grounds. Mitochondrial DNA 16S rRNA sequences demonstrate that populations from San Diego County, Los Angeles County, Santa Rosa Island, and Monterey County are extremely divergent (6 - 12%). These results are comparable to, or higher than recent reports of species - level differences in other invertebrate taxa. A molecular clock hypothesis shows that these four populations have been separated for 2 - 6 million years. A statistical cluster analysis of morphological features demonstrates that this genetic divergence is not reflected in anatomical features that might signify ecological differentiation among these lineages. The species status of these divergent populations of A. simus depends upon the species concept utilized. The time - limited genealogical perspective that is employed separates A. simus into two genetically distinct species. This study suggests that a species concept based on morphological distinctiveness in spider groups with limited dispersal capabilities probably underestimate taxonomic diversity. Ph. D.

DNA similarity studies were used to determine the species of thirty-one strains of bacteria isolated from the hemolymph of infected larvae from Mexico and throughout Central and South America. Twenty-one of the strains were determined to be Paenibacillus popilliae and ten were found to be more closely related to Paenibacillus lentimorbus. Only one of the P. popilliae strains, an isolate from Mexico, was resistant to the antibiotic vancomycin, a trait characteristic of P. popilliae strains from other geographic areas. As expected, all P. lentimorbus strains were sensitive to vancomycin. The polymerase chain reaction (PCR) was used to amplify a portion of a ligase gene necessary for vancomycin resistance in the Mexican strain. Sequencing of the amplicon revealed a sequence identical to that obtained from a North American strain of P. popilliae previously described. The ability of P. popilliae and the inability of P. lentimorbus to grow on medium supplemented with 2% sodium chloride has been used as a phenotypic trait for differentiating between the two species. Approximately 86% of the P. popilliae strains were capable of growth on medium supplemented with 2% sodium chloride and 60% of the P. lentimorbus strains were not capable of growth on this medium. Microscopic examination revealed that all of the Mexican and Central and South American strains of P. popilliae and P. lentimorbus produced a parasporal body. PCR was used to amplify two different regions of the cry18Aa1 gene encoding the paraspore in all of the isolates. One primer pair, CryBP2, detected the cry18Aa1 gene in 17 of the 21 P. popilliae strains and in all ten of the P. lentimorbus strains. The second primer pair, CryBP4, detected the parasporal gene in 20 of the 21 P. popilliae strains and in all ten of the P. lentimorbus strains. Thirty of the thirty-one P. popilliae and P. lentimorbus strains produced amplicons of approximately 616 bp with the CryBP4 primers. The CryBP4 primers did not detect the paraspore gene in one of the P. popilliae strains. The CryBP2 primer pair produced amplicons of three different sizes, indicating possible variability in the parasporal proteins of P. popilliae and P. lentimorbus. Eleven of the P. popilliae strains produced CryBP2 amplicons approximately 660 bp in size and six of the P. popilliae strains produced CryBP2 amplicons approximately 1100 bp in size. The cry gene was not detected in four of the P. popilliae strains with the CryBP2 primers. The P. lentimorbus strains produced CryBP2 amplicons approximately 750 bp in size. Three PCR products representing the variable CryBP2 amplicon sizes were sequenced and compared to the published cry18Aa1 gene sequence. Sequencing data revealed that the Central and South American CryBP2 amplicons are similar to the published cry18Aa1 sequence, however, the 1100 bp amplicon has a 453 bp insert that is not found in the published cry18Aa1 gene sequence. Paraspores are produced by P. popilliae and P. lentimorbus and is not a reliable phenotypic trait for differentiation between the two species. The ability of Mexican and Central and South American strains of P. lentimorbus to produce paraspores supports the previous findings of a North American group of P. lentimorbus strains that were also capable of producing paraspores. Except for one Mexican strain of P. popilliae, the Central and South American strains of P. popilliae are sensitive to vancomycin. This was unexpected since all North American strains of P. popilliae are vancomycin resistant. Vancomycin resistance could be useful in identifying strains of P. popilliae from North America but not for identifying strains of P. popilliae from Central and South America. So far, no vancomycin resistant strains of P. lentimorbus have been identified. There was variability in the ability of these organisms to grow on medium supplemented with 2% sodium chloride so the usefulness of this trait is debatable. However, the majority of P. popilliae strains from Mexico and Central and South America will grow on medium supplemented with 2% sodium chloride and the majority of the P. lentimorbus strains from these same areas will not grow on this medium. North American strains of P. popilliae and P. lentimorbus also showed variability of growth on medium supplemented with 2% sodium chloride. Master of Science

Montagnea arenaria [Hymenogastrales, Basidiomycota] , adapted to desert and xeric habitats, is morphologically and phenotypically variable. Species have been described on the basis of macromorphology and spore shape and size. This study was initiated to investigate populations of M. arenaria from Namibia in Africa and the Southwestern United States. It was hypothesized that biological species would exist in the widely separated populations. Spores from single sporocarps were germinated, single spore isolates were obtained and selfed to obtain mating types. On transfer, clamp connections were not maintained and mating patterns could not be achieved. Nuclear staining revealed multinuclei in the hyphae of both single spore isolates and compatible crosses. Spores were stained and found to have either 1 or 2 nuclei, but only four sterigmate basidia were observed. Limited partial compatibility was achieved and in some cases clamp connections formed within and between crosses from the two continents. Genomic DNA was extracted from old herbarium specimens. The ITS1, 5.8S, and ITS4 regions of nuclear ribosomal DNA were amplified and sequenced directly. Phylogenetic analysis using PAUP was performed. The hypothesis that Montagnea would form different biological species based on continental separation was rejected. In fact, the complex of isolates from widely varying locations not only had partial compatibility, but the variation in ITS sequences among widely distributed collections was relatively low. Lastly, no correlation between sporocarp size and gene flow among specimens from a wide variety of habitats was found. It appears that M. arenaria is a single, highly variable, widely distributed species. Master of Science

Nymphaea petersiana Klotzsch (Nyika) is an important wild tuber eaten in some districts of Malawi. The tubers were processed by boiling/freeze-drying(BFD) and sun-drying(USD). The tuber's nutrient and antinutrient composition was determined to produce a preliminary nutrient data base for use in sub-Saharan Africa. There was no significant difference (P > 0.05) in protein content of BFDand USD samples. Sun-dried samples were significantly (P < .05) higher in ash than boiled samples while boiled samples were significantly higher (P < .05) in crude fat and total carbohydrate. The protein content of the tubers (8.0 and 8.1 %) was higher than that of the staple maize (7.9%), African millets (unspecified) (7.5%), and polished rice (7.0%), but lower than sorghum (10.7%). Protein content was higher than tubers like cassava (1.3%), potato (2%), sweet potato (1.6%), yams (1.5%) and N. lotus (5.2). Nyika tubers have a well balanced amino acid content, limiting only in lysine. There were no significant (P > 0.05) differences in the mineral content of BFD and USD samples, except for iron, which was lower in the boiled samples. Nyika tubers have a higher calcium (1376 and 946 ug/g) and phosphorus (2250 and 2883 ug/g ) content than wild and domesticated cassava, potatoes, sweet potatoes and wild and domesticated yams. Sun-dried tubers have a higher iron content (88ug/g) than maize (20ug/g). The zinc content of tuber was higher (21and 25ug/g) than that of boiled maize flour, boiled sorghum flour, rice, cassava, and sweet potato. The predominant fatty acids in the tubers were oleic (47%), linoleic (32%), palmitic (21%) and linolenic (7%) acids. Ascorbic acid content was very low, only 0.1 and 0.003 mg/100g. Tannin content was lower (1 and 1.7 %) in the tubers than in Vulgare Pers. sorghum, DeKalb sorghum from U.S. and Kabale sorghum from Uganda. There was a significantly (P < 0.05) lower content of phytate in boiled (3.9ug/g) than in sun-dried tubers (6.0 ug/g). Phytate content of the tubers was lower than that of cooked maize flour, unrefined maize flour, cassava and sweet potato. Trypsin inhibitor activity in the tubers was reduced from 463 to 55 TIU/g tuber and chymotrypsin inhibitor activity was reduced to 50 from 267 CIU/g tuber by cooking. Nyika is a good source of iron and quality protein limiting only in lysine. Protein is comparable to staple maize and higher than root crops consumed in Malawi. It is not a good source of fat and ascorbic acid. Tannin, phytate, trypsin, and chymotrypsin inhibitor content lower than most food crops consumed in Malawi. Ph. D.

The Crepidotaceae (Imai) Singer (Basidiomycetes: Agaricales) represents a proposed family of saprophytic fungi containing five agaricoid (Crepidotus, Tubaria, Melanomphalia, Simocybe, Pleurotellus) and four cyphelloid (Episphaeria, Phaeosolenia, Pellidiscus, Chromocyphella) genera. Several contemporary classification systems exist that delegate some or all of these genera to other agaric families. Phylogenetic relationships for the most prevalent genera in the Crepidotaceae were investigated using nuclear large subunit ribosomal DNA (LSU rDNA) sequences. Parsimony analysis of the molecular data supports the Singer classification of Crepidotus, Melanomphalia, and Simocybe as a single monophyletic unit within the Agaricales. The affinities of the genus Tubaria remain uncertain. Crepidotus (Fr.) Staude is the largest and most phenotypically variable genus in the Crepidotaceae. Sequencing of the LSU rDNA region for a cross-section of morphologically diverse species suggests that Crepidotus is not a monophyletic genus. Analysis of morphological characters for 23 Crepidotus taxa shows that characters traditionally applied for infrageneric classification of Crepidotus are homoplasic in origin, but that less commonly emphasized characters such as spore shape and ultrastructure of spore wall ornamentation may be indicative of monophyletic clades for this complex. A unique pattern of basidiospore dormancy and germination, unknown in any other species of agaric, is reported for 11 species of Crepidotus. Similar patterns were also encountered in species of Simocybe and Melanomphalia. In these species an endogenous period of spore dormancy of four to six months is followed by an activation period where the factors necessary for subsequent germination appear to involve a minimal nutritional component, water, and exposure to light. Master of Science

Thickets of Rhododendron maximum (Rm) in the southern Appalachians impose severe limitations on the regeneration of hardwood and coniferous seedlings. Interactions between Rm thickets and ectomycorrhizal colonization were examined to explain seedling inhibition. Experimental blocks were established in and out of Rm thickets in a mature, mixed hardwood/conifer forest in Macon County, North Carolina. Planted seedlings of red oak (Quercus rubra) and hemlock (Tsuga canadensis) were harvested at the end of the first and second growing seasons. Litter manipulation had no effect on total mycorrhizal colonization. Mycorrhizal colonization and ramification index (# mycorrhizae cm-1) were depressed and colonization by Cenococcum geophilum increased in blocks with versus without Rm. After the first year, percent colonization of T. canadensis not in Rm thickets (62 %) was three times higher than in Rm thickets (19%), and the ramification index was increased by more than a factor of four (2.83 versus 0.61). Mycorrhizal colonization levels were correlated with root weight and shoot weight in both hemlock and oak seedlings, but did not explain most of the variation observed. Sporocarps of 69 putatively ectomycorrhizal species were collected on the blocks. Species diversity and overall community structure was similar in and out of Rm thickets. Individual species, i.e. Lactarius speciosus and Russula krombholzii, were significant indicators of forest without Rm thickets. Rhododendron maximum thickets probably affect the process of mycorrhization. The reduced level of mycorrhizal capacity under Rm thickets could be a factor in the increased seedling failure in Rm thickets. Master of Science

Broomrapes are achlorophyllous holoparasites of many economically important dicotyledonous crops. As weeds, they cause reductions in crop yield, adversely affect crop quality, and result in loss of cultivated land due to reduced crop alternatives. Few effective control measures exist for broomrapes. One of the most promising approaches is the use of low rates of glyphosate in hosts with tolerance to the herbicide. Recently, availability of glyphosate-resistant crops has provided an alternative in broomrape infested areas. Knowledge about the nitrogen status of broomrapes is essential for developing new control strategies. Broomrapes have two potential sources of amino acids. First, the haustorium aids in the translocation of amino acids from the host plant to the parasites. Second, broomrapes may be able to synthesize some amino acids themselves and obtain the rest from the host. However, the relative importance of these two modes of acquiring amino acids by broomrapes is not clear. Osmotic stress has been implicated as a possible reason for inhibition of broomrape germination by nitrogen. To date, there has been no attempt to correlate osmotic potential with nitrogen induced inhibition of broomrape germination. Optimum temperatures for conditioning and germination are different among broomrape species. Although temperature is known to influence germination in broomrape, its effect on subsequent development of the parasitic seedling has not been studied. Studies were conducted to determine the use of glyphosate in controlling broomrape in common vetch that is tolerant to low rates of glyphosate, and to compare this response with broomrape control in oilseed rape that has been genetically engineered for glyphosate resistance. Glyphosate dose response studies using a commercial formulation and patterns of absorption, translocation, and metabolism, using ¹⁴C-glyphosate, were determined for both host crops. Glyphosate significantly reduced the growth of broomrape at 0.18 and 0.36 kg ae ha⁻¹> in common vetch and 0.25 to 0.75 kg ha⁻¹ in oilseed rape. More than 25% of translocated ¹⁴C-glyphosate in both host crops accumulated in broomrape tubercles. Broomrape parasitism caused a redistribution of translocated ¹⁴C-glyphosate in the roots of both host crops. Glyphosate was metabolized up to 25% in common vetch, but remained intact in oilseed rape. Studies were conducted to analyze amino acid composition of both nonparasitized and broomrape-parasitized hosts and associated broomrape after hydrolysis and phenylisothiocyanate derivatization of amino acids. Results indicated that amino acid concentrations of leaves of parasitized carrot plants were lower than those of the leaves of nonparasitized carrot plants. Broomrape tubercles had equal or higher amino acid concentrations compared to those of the leaves of nonparasitized carrot plants. Levels of free alanine and arginine concentrations of broomrape callus were higher than those of any other tissue of either carrot or broomrape. The effect of glyphosate on the host-broomrape interaction regarding amino acid metabolism was examined. Glyphosate generally increased the amino acid concentrations in common vetch and oilseed rape plants, and broomrape attachments. The aromatic amino acids, phenylalanine and tyrosine, did not differ from this pattern. Concentrations of certain amino acids in broomrape were similar to those of parasitized common vetch and parasitized oilseed rape, whereas levels of several others, were higher in broomrape attachments compared to the host plants. In vitro studies were conducted to determine the influence of osmotic potential and temperature on broomrape germination. Osmotic potential significantly affected germination and radicle elongation of broomrapes. No correlation was found between osmotic potential and ammonium-induced inhibition of germination of broomrapes. Temperature significantly influenced germination and radicle elongation of all broomrape species tested. Ph. D.

Cereals represent one of the most important food crops in the world. Rice (Oryza sativa) is one of the most commonly consumed cereal grains, and as a result, has considerable economic and agricultural importance, despite their potential as a source of genetic material. The phylogenetic relationships among rice and its wild species are not well understood. The objectives of this study are to evaluate the rate and type of molecular variation present in the gene that encodes the 10 kDa prolamin polypeptide in Oryza (Poaceae), and use this information in understanding the evolution of the Oryza genus. Our principle hypothesis is that genetic diversity exists at the molecular level in wild species of Oryza, and that this diversity can provide useful information regarding the phylogenetic relationships among rice species and permit a more theoretical examination of the evolutionary processes, such as concerted evolution, within Oryza and its nine genomes. A phylogeny of Oryza is presented, and modes of evolution are discussed. Master of Science

Comparative DNA sequencing of matK, a maturase-encoding gene located within the intron of the chloroplast trnK gene, was evaluated for phylogenetic utility above the family level and within the grass family (Poaceae). There are three major objectives in the research. The first one is to study the utility of the matK gene in plant evolution. The second objective is to characterize the matK gene in the grass family. The last major goal is to address the phylogenetic questions in the Poaceae using the matK sequences from representatives of different grass groups. In order to study the potential application of matK to plant systematics above the family level, eleven complete sequences from GenBank representing seed plants and liverworts and nine partial sequences generated for genera representing the monocot families Poaceae, Joinvilleaceae, Cyperaceae, and Smilacaceae were analyzed. The study underscored the following useful properties of the matK gene for phylogenetic reconstruction: reasonable size (1500 bp), high rate of substitution, large proportion of variation at the first and the second codon positions, low transition-transversion ratio, and the presence of mutationally-conserved sectors. The use of different sectors of the gene and the cumulative inclusion of informative sites showed that the 3' region was the most useful in resolving phylogeny, and that the topology and robustness of the tree reached a plateau after the inclusion of 100 informative sites. The presence of a relatively conserved 3' region and the less conserved 5' region provides two sets of characters that can be used at different taxonomic levels from the tribal to the division levels. It also has demonstrated the potential of partial sequencing in resolving systematic relationships from the tribe to the division level. The matK gene in the Poaceae was characterized with complete sequences from 11 grass genera, representing 7 subfamilies and 11 tribes, and one outgroup (Joinvillea plicata, Joinvillaceae). The alignment of 1632 base pairs from 14 species yielded a data set of 601 (36.8)% variable sites and 246 (15.1%) informative sites. The variations at nucleic and amino acid levels evenly distributed throughout the entire gene, and the 5' region appears to have more variation than the 3' region. The changes at the third codon position are very low as compared to the total of the first and second positions. This has led to a similar variation pattern at nucleic and at amino acid levels. The average tr/tv ratio generated from 14 entire matK sequences is 1.29. It is intriguing to find that the tr/tv ratios were regionally related. RASA analysis of the alignment data indicated a relatively high phylogenetic signal in the data set of 14 taxa. In the two half analyses, while the tRASA of the 5' half of the matK gene (0.43) is not significant, the 3' of the matK gene showed a significant phylogenetic signal. Among the 5 sections of the 14 entire matK sequences, only the fourth sector contains a statistically significant phylogenetic signal. These results indicate that matK is a phylogenetically valuable gene and that the 3' region of the matK gene contains strong phylogenetic information. A single most parsimonious tree was obtained from the 246 informative sites of the 14 entire matK sequences. Seven major groups were well resolved on the most parsimonious tree, corresponding to the seven commonly recognized subfamilies: Aruninoideae, Bambusoideae, Centothecoideae, Chloridoideae, Panicoideae, Pooideae and Oryzoideae. Approximately 960 base pairs of the matK gene were sequenced from grass species representing 48 genera, 21 tribes, and seven subfamilies to reconstruct a phylogeny for the Poaceae. Joinvillea plicata (Joinvilleaceae) was used as an outgroup species. The aligned sequences showed that 495 nucleotides (51%) were variable and 390 (36%) were phylogenetically informative. RASA indicated that very significant phylogenetic signals exist in this data set. The cumulative addition of informative sites starting at the internal end of the sequences revealed that at 300 sites, tree topology and bootstrap values matched those of the consensus tree based on the entire sequence. Parsimony analyses using PAUP resulted in six most parsimonious trees and a strict consensus tree showing major lineages supported by high bootstrap values. These lineages corresponded to six subfamilies: Bambusoideae, Oryzoideae, Pooideae, Chloridoideae, Panicoideae, and Arundinoideae. The Bambusoideae, including woody and herbaceous taxa, diverged as the most basal lineage, and the monophyletic oryzoid species formed a sister group. The Chloridoideae, Panicoideae, Arundinoideae, and the centothecoid Zeugitis (PACC group) emerged as a monophyletic assemblage with 95% bootstrap support. The Aristideae branched off as a monophyletic line basal to the chloridoid clade. Stipeae appeared as a sister taxon to the Pooideae. The matK-based phylogeny did not reveal a major dichotomy in the family. The matK gene has provided sequence information sufficient for good resolution of the major grass lineages. Ph. D.

The cosmopolitan Pteridium aquilinum (L.) Kuhn is widespread throughout eastern North American, where it is represented primarily by Tryon's (1941) var. latiusculum (Desv.) Underw. and var. pseudocaudatum (Clute) Heller. The taxonomy of Pteridium is controversial. Fourteen isozyme loci and 12 morphological characters were used to assess the taxonomic relationship of these two varieties. Isozyme data indicated a high mean genetic identity (I = 0.976) between eleven bracken populations. Strong patterns of geographic variation for isozyme allele frequencies were also observed. The isozyme results did not separate the two taxa. Numerical analysis of the morphology distinguished the two taxa when the qualitative characters were used alone or in conjunction with some of the quantitative traits. All qualitative characters differed significantly between the two taxa. No perceptible geographic pattern of variation was observed. Morphological distinctiveness was maintained even in those localities where both taxa were present, with few or no intermediates being found. Isozyme evidence suggestive of gene flow between the two varieties was found at Greensboro, NC, where the two morphotypes were easily recognizable. The isozyme evidence strongly indicates conspecificity, while the morphological evidence supports their status at the varietal level. Master of Science

Chitinase is an enzyme that catalyzes the hydrolysis of chitin, an essential component of the cell walls of many fungi. Plant chitinases have been implicated in plant defense against pathogens. In plants, chitinase often exists as isoforms and three classes of chitinases have been proposed. In this study, isolation, characterization and expression of chitinase genes in Kentucky bluegrass (Poa pratensis L.) were studied. With primers designed from conserved regions of chitinase genes from other plant species, a 710 bp fragment (CH710) containing a partial chitinase gene sequence was amplified from Kentucky bluegrass by PCR. Using cassette ligation mediated PCR, we amplified four 5’ and five 3’ unknown sequences flanking CH710. The sequence information of these flanking fragments led us to the amplification of three genomic sequences from Kentucky bluegrass by PCR, KBCH1, KBCH2 and KBCH3, which contain full coding regions of chitinase genes. The chitinase genes carrying KBCH1, KBCH2 and KBCH3 were designated as chi1, chi2 and chi3, respectively. Southern blot hybridization indicated the presence of more than seven chitinase genes in the Kentucky bluegrass genome. Chi1 and chi2 each contain an open reading frame with no introns, encoding polypeptides of 340 and 320 amino acids, respectively. Both CHI1 and CHI2, the predicted proteins encoded by chi7 and chi2, are class I chitinases and share 94% amino acid identity. CHI1 has a short C-terminal extension, implicating that this protein may be a vacuolar protein. Although chi3 has high sequence similarity to chi7 and chi2, the potential open reading frame of chi3 is interrupted by a translation termination codon at the 51st amino acid indicating that it does not encode a functional chitinase. In this study, the expression of chitinase genes in Kentucky bluegrass under various stress conditions was also investigated. RNA blot hybridization showed that stresses such as cold, heat, salicylic acid and ethephon all induced an increased accumulation of chitinase MRNA in Kentucky bluegrass leaves with ethephon leading to the highest induction. After ethephon treatment, the accumulation of chitinase transcripts at a high level was observed from 2 days to 5 days. The expression of two individual chitinase genes, chi1 and chi2, was shown to be stimulated, while being coordinately regulated, by ethephon. Ph. D.

This systematic study recognizes 15 species of the genus Mallos Pickard-Cambridge and three species of the genus Mexitilia Lehtinen. Three species of Mallos and one species of Mexitilia are newly described. Two species are placed in synonymy and two species are transferred to Mallos. The males of two Mallos species are described for the first time. A cladistic analysis based on 22 morphological produced a cladogram that supports the monophyly of Mallos and the validity of Mexitilia. For five species of Mallos and one species of Mexitilia mitochondrial and nuclear DNA sequences were analyzed by restriction digest. The seventeen resulting restriction sites produced a cladogram that agreed with the one based on the morphology of these six species. When morphological and molecular characters were combined they produced a single tree that was identical to that based on molecular data alone. These molecular and morphological characters present the same picture of Mallos and Mexitilia phylogeny. Master of Science

Shale barrens are unique habitats located throughout the southern Appalachians. They are characterized by a south or south west aspect, a steep slope, and an exposed rocky surface (Platt, 1951). They have a high total irradiance and can experience temperatures higher than the surrounding deciduous forest. A variety of plant species, several of which are rare or endangered, are endemic to the shale barren habitat. One reason proposed fc)r their endemism is that the plants are obligate heliophytes (Keener, 1983). The purpose of this dissertation is tel examine the acclimation ability of the shale barren endemic Eriogonum alIeni to shade and high temperature. Ph. D.

Thirty-two monoploids (2n = x = 12) derived by anther culture of ten diploid clones of Solanum phureja were used to generate doubled monoploids through in vitro shoot regeneration. Doubled monoploids were compared to the anther donor and progenitor monoploids for morphological characteristics, and were evaluated for fertility in the greenhouse and progeny performance under field conditions. Monoploids varied for frequency and earliness of shoot regeneration, number of shoots formed per explant and frequency of chromosome doubling among regenerated shoots. Regeneration was greater when stock plantlets were frequently subcultured (2- or 4-week intervals) and maintained under a 16 h photoperiod, and when explants were incubated at 20°C compared to 25°C. In addition, leaf explants regenerated at higher frequencies than stem explants. Significant high correlations between monoploids and their doubled monoploids were observed for 14 of 17 characters in the greenhouse. Doubled monoploids were significantly greater than monoploids for 15 characters, indicating a positive effect of increasing gene dosage from monoploid to diploid. The anther donor was not significantly greater than the mean of doubled monoploids for 10 characters; therefore, for specific characters, doubled monoploids without homozygote depression can be obtained. Doubled monoploids varied for nurnber of days to flower, duration of flowering, abundance of flowers, flower quality, fruit set and seed set; they had lower fruit and seed set than the anther donor. A few clones produced low levels of stainable pollen which had high 2n pollen frequency but did not germinate in vitro. Therefore, they were considered male-sterile for practical purposes. Used as female parents, doubled monoploids were able to transmit the 2n pollen trait to their progenies. Two of four doubled monoploids exhibited superior general combining ability over the anther donor under field conditions. This demonstrates the potential of passage of a heterozygous genotype through the monoploid sieve. The advantage of the monoploid sieve may be more or less evident depending on the combining ability of the crossing partner and variable performance can be expected among doubled monoploids from an unselected anther donor. The performance of unselected doubled monoploids demonstrates the potential for their utilization in breeding and warrants further research in the area. Ph. D.