Plant sciences

The study examines the phytochemical and phytohormone profiles of two ecotypes of Doliocarpus dentatus (Capadulla) from Guyana with an emphasis on their potential therapeutic applications, particularly in the context of erectile dysfunction. The research concentrates on the red and white ecotypes, which are visibly differentiated by the colouration of their inner and outer bark; moreover, the red ecotype is widely regarded by traditional knowledge to be more efficacious in treating erectile dysfunction. The study seeks to provide much-needed scientific evidence to support the traditional medicinal uses of D. dentatus and aims to accomplish this by providing semi-targeted and targeted analysis of its bioactive compounds.The thesis employs a multi-faceted approach, commencing with Chapter 2, an examination of the international and national frameworks that govern natural product research in Guyana, thereby ensuring ethical and sustainable practices. Chapter 3, Liquid chromatography-mass spectrometry-based metabolomics was utilized to analyze the phytochemical profiles of the red and white ecotypes of D. dentatus. Approximately 847 compounds were identified, with 138 tentatively classified as potentially therapeutic, particularly polyphenols exhibiting antioxidant properties. Chapter 4 focus on the phytohormone profiling revealed that the red ecotype generally exhibited higher levels of active cytokinins, particularly trans-zeatin (tZ) and cis-zeatin (cZ), compared to the white ecotype. A strong correlation was identified between specific cytokinins and secondary metabolites such as flavonoids and alkaloids, suggesting complex interactions that may enhance the therapeutic potential of D. dentatus. Chapter 5 further analysis of total phenolic content, flavonoid content, and their antioxidant capacities demonstrated differences between the red and white ecotypes, with the red ecotype generally showing higher levels of these compounds. Both ecotypes demonstrated significant antioxidant activity, with the red ecotype showing enhanced radical scavenging capacity. The study also revealed a diverse range of bioactive compounds, including flavonoids, terpenoids, and alkaloids, which may contribute to the plant's reported medicinal properties. This semi-targeted and targeted analysis provides novel insights into the phytochemical and phytohormone profiles of D. dentatus ecotypes, offering a scientific basis for their traditional medicinal uses and potential therapeutic applications. The findings suggest that D. dentatus, particularly the red ecotype, may have significant potential for developing new drugs, especially in the context of erectile dysfunction therapy.

Author Keywords: antioxidant, Bioactive compounds, liquid chromatography-electrospray ionization tandem mass spectrometry, metabolomics, secondary metabolites, therapeutic properties

Hybridisation is an important evolutionary mechanism with diverse outcomes, including the formation of new lineages, the exchange of alleles between species, or their extinction through genetic swamping. In some cases, hybrids exhibit higher fitness than their parental species, which can lead to hybrid invasions that threaten ecosystems. In North America, hybridisation between the native cattail Typha latifolia and the non-native T. angustifolia produces the hybrid T. × glauca, which is a highly impactful invader in wetlands across large areas of southern Canada and northern USA; contrastingly, in Europe and Asia, where its parental species also co-occur, T. × glauca is rare to non-existent. This thesis examines the evolutionary history of T. latifolia and T. angustifolia, which, in addition to being the parental species of T. × glauca, are two of the most globally widespread Typha species, a genus of plants foundational to wetlands. First, we developed genomic resources, including ~12M nuclear SNPs and plastome assemblies to facilitate genetic research on Typha; we also described a cost-effective library preparation and genotyping protocol that makes population genetic studies of freshwater plants accessible. Then, we applied those genomic resources to investigate the roles of drift, selection, and hybridisation in the divergence of T. angustifolia, T. latifolia, and their sister species, T. domingensis and T. shuttleworthii. We found that speciation in these taxa was driven by drift under allopatry, resulting from historical bottlenecks, and that natural selection has played a minimal role in the divergence of these species; additionally, we observed introgressive hybridisation from T. latifolia into T. angustifolia. Finally, we reconstructed the demographic histories of T. angustifolia and T. latifolia from North America and Europe. We observed that reproductive isolation is strong in Europe, where the two species have potentially been sympatric for ~800,000 years, and weak in North America, where they have been sympatric for only a few centuries. Our results exemplify how the divergence and demographic histories of species can correlate with their strength of reproductive isolation. We emphasise that preventing invasions by hybrids will require limiting the movement of Typha and other historically allopatric species, which likely lack reproductive barriers.

Author Keywords: biological invasions, cattails, demographic histories, evolutionary histories, population genomics, reproductive isolation

Ecological restoration of grassland ecosystems is increasing in scope in Ontario, as a result the demand for genetically appropriate, high-quality seeds of native plants is also increasing. This mixed methods study characterized fitness and seed quality traits using genetic, demographic and growth trial data for a keystone tallgrass prairie species Big bluestem (Andropogon gerardii). To estimate the ploidy levels in Big bluestem, our flow cytometric results indicated an average of 6.32 picograms of nuclear DNA within sixteen populations surveyed showing that hexaploid (6x) cytotypes are dominant in Southern and Central Ontario populations, aside from one 9x occurrence in Norfolk county, ON. Seed quality, measured through germination and viability testing did not change based on whether a population was remnant or restored. Concerningly, our study shows that remnant populations of Big bluestem are at risk of being lost as high quality seed sources likely because of the absence of stewardship and the resulting loss in population fitness. Workshops with prairie restoration practitioners suggest that there is significant vision behind the future of this work in the province, and that an ecosystem wide seed strategy for keystone tallgrass prairie species is a necessary next step to increase the sustainability of seed-based restoration strategies and preserve remnant site genetics.

Author Keywords: Big bluestem (Andropogon gerardii), Ecological Restoration, fitness, polyploidy, practitioner, seed quality

Macrophytes play an important role in aquatic ecosystems, and thus are integral to ecological risk assessments of environmental contaminants. In this dissertation, I address gaps in the assessments of contaminant fate and effects in macrophytes, with focus on glyphosate herbicide use for invasive plant control. First, I evaluated the suitability of Typha as future standard test species to represent emergent macrophytes in risk assessments. I concluded that Typha is ecologically relevant, straight-forward to grow, and its sensitivity can be assessed with various morphological and physiological endpoints. Second, I assessed effects from glyphosate (Roundup WeatherMAX® formulation) spray drift exposure on emergent non-target macrophytes. I performed toxicity tests with five taxa, Phragmites australis, Typha × glauca, Typha latifolia, Ammannia robusta, and Sida hermaphrodita, which in Canada collectively represent invasive, native, and endangered species. I found significant differences in glyphosate sensitivity among genera, and all species' growth was adversely affected at concentrations as low as 0.1% (0.54 g/L), much below the currently used rate (5%, 27 g/L). Third, I assessed the potential for glyphosate accumulation in and release from treated plant tissues. I found that P. australis and T. × glauca accumulate glyphosate following spray treatment, and that accumulated glyphosate can leach out of treated plant tissues upon their submergence in water. Finally, I assessed effects of released glyphosate on non-target macrophytes. I found that P. australis and T. × glauca leachate containing glyphosate residues can stimulate the germination and seedling growth of T. latifolia, but can exert an inhibiting effect on A. robusta, although leachate without glyphosate caused similar responses in both plants. Additionally, I found no negative effects in A. robusta when exposed to glyphosate residues in surface water, or when grown with rhizosphere contact to an invasive plant that was wicked (touched) with glyphosate. My results show that non-target macrophytes can be at risk from glyphosate spray for invasive plant control, but risks can be mitigated through informed ecosystem management activities, such as targeted wick-applications or removing plant litter. Integrating contaminant fate and effect assessments with emergent macrophytes into ecological risk assessments can support the protection of diverse macrophyte communities.

Author Keywords: Ecosystem management, Ecotoxicology, Glyphosate, Herbicide, Invasive plant, Species at risk

Glycine max (soybean) is an economically important plant species that registers a relatively low yield/seed weight compared to other food and oil seed crops due to higher rates of flower and pod abortion. Alleviation of this abortion rate can be achieved by altering the sink strength of the reproductive organs of soybeans. Cytokinin (CK) plays a fundamental role in promoting growth of sink organ (flowers and seeds) by increasing the assimilate demand. Cytokinin oxidase/dehydrogenase (CKX) is an enzyme that catalyses the irreversible breakdown of active CKs and hence reduce the cytokinin content. The current thesis uncovers the members of CKX gene family in soybeans and the natural variations among CKX genes within soybean varieties with different yield characteristics. The identification of null variants of OsCKX2 that resulted in large yield increases by Ashikari et al. (2005) provided a rationale for current thesis. The soybean CKX genes along with the ones from Arabidopsis, Rice and Maize were used to construct a phylogenetic tree. Using comparative phylogeny, protein properties and bioinformatic programs, the potential effect of the identified natural variations on soybean yield was predicted. Five genes among the seventeen soybean CKXs identified, showed polymorphisms. One of the natural variations, A159G, in the gene GmCKX16 occurred close to the active site of the protein and was predicted to affect the activity of enzyme leading to higher accumulation of CKs and hence increased seed weight. Use of such natural variations in marker assisted breeding could lead to the development of higher yielding soybean varieties.

Author Keywords: CKX, Cytokinins, Seed weight, Seed Yield, SNPs, Soybeans

Abstract

Early Responses of Understory Vegetation After One Year of Above Canopy Nitrogen Additions in a Jack Pine Stand in Northern Alberta

Nicole Melong

Nitrogen (N) emissions are expected to increase in western Canada due to oil and gas extraction operations. An increase in N exposure could potentially impact the surrounding boreal forest, which has adapted and thrived under traditionally low N deposition. The majority of N addition studies on forest ecosystems apply N to the forest floor and often exclude the important interaction of the tree canopy. This research consisted of aerial NH₄NO₃ spray applications (5, 10, 15, 20, 25 kg N ha-1yr-1) by helicopter to a jack pine (Pinus banksiana Lamb.) stand in the Athabasca Oil Sands Region (AOSR) in northern Alberta, Canada. The main objective was to assess the impacts of elevated N after one year of treatment on the chemistry of understory vegetation, which included vascular plants, terricolous lichens, epiphytic lichens and a terricolous moss species. Changes in vegetation chemistry are expected to be early signs of stress and possible N saturation. Increased N availability is also thought to decrease plant secondary compound production because of a tradeoff that exists between growth and plant defense compounds when resources become available. Approximately 60% of applied N reached the ground vegetation in throughfall (TF) and stemflow (SF). Nitrate was the dominant form of N in TF in all treated plots and organic N (ON) was the dominant form of N in SF in all plots. The terricolous non-vascular species were the only understory vegetation that responded to the N treatments as N concentration increased with increased treatment. Foliar chemistry of the measured epiphytic lichens, vascular species, and jack pine was unaffected by the N treatments. Based on biomass measurements and N concentration increases, the non-vascular terricolous species appear to be assimilating the majority of TF N after one year. Vegetation from the high treatment plot (25 kg N ha-1yr-1) was compared to a jack pine forest receiving ambient high levels of N (21 kg N ha-1yr-1) due to its proximity to Syncrude mining activities. Nitrogen concentrations in plant tissues did not differ between the two sites; however, other elements and compounds differed significantly (Ca, Mg, Al, Fe). After one year of experimental N application, there were no environmental impacts consistent with the original N saturation hypothesis.

Author Keywords: Athabasca Oil Sands Region, Canopy Interactions, Jack Pine, Nitrogen, Secondary Chemistry, Understory Vegetation

Cis-isomers of the cytokinin plant hormone family are thought to have low activity or impact on plant growth and development. Mutants with independent silencing of the pathway leading to cis-CK (cis-cytokinin) were investigated at the phenotype and metabolite levels. Phenotypic deviations were noted in trichome development, fresh weight, rosette diameter, number of non-rosette leaves, shoot height, delayed flowering, flower number, and carotenoids. Exploratory metabolomic analysis detected a number of metabolite features that have been associated with CK, auxin, and ABA (abscisic acid) activity. Evidence from both phenotype and metabolomic analysis support the hypothesis that cis-CK production is biologically important for plant growth and development.

Author Keywords: arabidopsis, cytokinin, IPT, metabolmics

In Canada, eastern flowering dogwood (Cornus florida L.) is an endangered tree that occurs only in the Carolinian forest of southern Ontario. Threats to this species include habitat fragmentation and the fungal pathogen dogwood anthracnose (Discula destructiva). I conducted a population genetic analysis using seven nuclear microsatellite markers to determine if fragmented populations are genetically isolated from one another and have low levels of genetic diversity. Genetic comparisons suggest on-going dispersal among sites and relatively high genetic diversity within most sites; however, smaller populations and younger trees were less genetically diverse. I also used linear mixed effects models to assess potential relationships between several ecological variables and the prevalence of dogwood anthracnose. Disease severity was higher in trees on shallow slopes and in larger trees; the latter also had higher likelihood of infection. Insights from this study will be important to incorporate into future management strategies.

Author Keywords: Cornus florida, Discula destructiva, dogwood anthracnose, Eastern flowering dogwood, endangered, population genetics

Wild lupine (Lupinus perennis) restoration efforts seek to increase and connect populations, using seeds, to facilitate the recovery of endangered butterflys in Ontario. This study observed plant growth and phytohormone levels of L. albus, L. polyphyllus, and L. perennis through stages of seed development, each with varying strategies in growth and reproductive investment. L. polyphyllus is similar to L. perennis in morphology, acting as similar comparable with L. albus, a well-studied annual, as an outgroup comparator.

Wild lupines showed a lack of sexual reproductive effort as they did not put as much effort into above ground growth, and few in the population reproduces. They also showed cis-zeatin, a weaker cytokinin, throughout development and had higher amounts of abscisic acid at the end of seed maturity, impacting their ability to develop and germinate. These factors contribute to why wild lupines are difficult to restore using seeds, limiting expansion and challenging restoration.

Author Keywords: L. albus, L. perennis, L. polyphyllus, plant physiology, seed development, Wild blue lupine

The activities of phytohormones during the reproductive phase have been partially clarified in seed physiology while the biological role of plant hormones in oil accumulation during seed development has been investigated in part only. In this research, fatty acid (FA) contents and hormone profiles, including abscisic acid (ABA) and cytokinins (CKs) of seed samples in four different stages and comparing six soybean varieties have been investigated in order to examine the hypothesis that the endogenous plant hormones play important roles in FA production in soybean seeds. The FA contents increased significantly during this period while the hormone concentrations gradually declined towards the seed physical maturation. However, the interactions between FA contents and hormone profiles were complex and went beyond linear correlations. Hormone metabolism in the earlier stages of seed maturation period demonstrated numerous robust relationships with FA accumulations, as derived from several simple and multiple regression models in the determination of different FA contents. Evaluation of the effects of exogenous ABA and trans-Zeatin (tZ) on FA biosynthesis has revealed that ABA appears to be involved in the accumulations of unsaturated FAs while tZ participated in the synthesis of saturated and unsaturated FAs. Notably, the alterations of FA synthesis differ according to what exogenous hormone concentrations could be used.

Author Keywords: Abscisic acid, Cytokinin, Fatty acid, Seed development, Soybean