Botany

Diversity of flowers in flowering plants is understood to facilitate mating success, and yet understanding the relationships between plant sexual diversity and mating patterns remains a challenge. In this thesis, I used Sagittaria latifolia, an aquatic plant with unique sexual systems, to investigate relationships between plant sexual diversity, mating patterns, and reproductive success and to understand their evolutionary consequences. First, I measured the magnitude of sexual selection in a dioecious and a monoecious population of S. latifolia, while accounting for size variation. I showed that sexual selection does operate in natural populations of plants. Estimates of sexual selection were similar in both populations, possibly due to the temporal separation of female and male flowering in hermaphroditic S. latifolia, enabling individuals to act temporarily as unisexuals. Second, I examined how seasonal differences in sex allocation and flowering time can affect male mating opportunities, and the effect on reproductive success in hermaphroditic S. latifolia. I found that shifting allocations to be more male-biased with earlier flowering increased male mating opportunities. However, greater mating opportunities did not equate to higher rates of siring, indicating that factors other than temporal overlap contribute to male reproductive success. Finally, I determined the factors affecting mating and reproductive success of males and male-function hermaphrodites, distinguishing between the predictable effects of plant-level traits and population-level features, and unpredictable effects of stochastic factors. Across both populations, we found that plant-level traits had a greater impact on mating and reproductive success than population-level features. Nevertheless, these effects were frequently masked by stochastic factors. Furthermore, unexplained variation in mating and reproductive success may stem from additional plant traits affecting post-pollination gametophyte-sporophyte interactions. My findings indicate that while sexual selection does operate in natural populations of S. latifolia as expected under Bateman's principles, temporal overlap in flowering, plant traits and population features did not explain patterns of male mating or reproductive success, suggesting that post-pollination factors may be influencing mating outcomes.

Author Keywords: Phenology, Plant mating, Plant reproduction, Sagittaria latifolia, Sex allocation, Sexual selection

Darwinian fitness is determined by the outcome of allocations of energy and nutrients to plant growth, survival, and reproduction. Allocations to reproduction differ from allocations to growth and survival because the products of reproduction (pollen, ovules, and fruits) are dispersed away from the plant and so the resources used to produce them cannot be re-allocated. As a result, and because the total resource pool available to a plant is limited, allocations to reproduction can reduce future growth, reproduction, and/or survival, yielding fitness costs of reproduction. This study provides a novel approach to detecting the costs of reproduction through multiple reproductive currencies, including both biomass and nitrogen allocations and their temporal effects on current and future plant performance. My results demonstrate the environmental dependence of the costs of reproduction and show that under certain conditions, males can bear greater costs than females, contrary to the prevailing view.

Author Keywords: Cost of reproduction, dioecy, flower removal, life-history trade-offs, photosynthetic machinery, sexual dimorphism

Diversified agroecosystems supporting greater genetic, structural, and functional diversity improve soil health and ecosystem function. However, there is limited understanding of how multiple forms of diversification, such as mixing cover crop species and adding arbuscular mycorrhizal fungi (AMF), alter belowground carbon supply to soil. In a controlled environment experiment using rhizoboxes, I investigated the belowgound response of cover crops – red clover (Trifolium pratense) and barley (Hordeum vulgare) – grown in monoculture or mixture, with and without AMF inoculation. Root morphological and mycorrhizal traits that characterize the hypothesized root economics spectrum (RES) were integrated with novel sampling of dissoved organic carbon fluxes and easily extractable glomlin in rhizosphere soil. Results revealed species-specific shifts on the RES suggesting that diversification through species mixing and AMF additions can alter belowground carbon allocation pathways, with potential implacations for plant performance and soil carbon stabilization in agroecosystems.

Author Keywords: Arbuscular mycorrhizal fungi, carbon sequestration, cover crops

Using Drosophila deficiency (Df) and Over Expression (OE) (GAL4/UAS>dCas9-VPR; sgRNA) gene systems, it was demonstrated that Dmel_CG31381 and Dmel_CG11089 are functional tRNA isopentenyltransferase (EC 2.5.1.8) genes (tRNA IPT1 and IPT2) critical to the first committed step in insect cytokinin biosynthesis. IPT Df mutants showed significant decreases in total CK levels and IPT1/IPT2 transcript levels compared to parent lines. IPT OE mutants showed significant increases in total CK levels and IPT1/IPT2 transcript levels compared to parent lines. Further, endogenous CK analyte levels and qPCR relative fold gene expression of Dmel_CG31381 and Dmel_CG11089 (tRNA IPT1 and IPT2) genes demonstrated expression patterns with functional confirmation corresponding to the predicted IPT mutant variants. The functional confirmation of tRNA IPT1 and IPT2 as the first committed step was further supported by the bioinformatic detection of putative gene homologs to corroborate seven remaining enzyme transcripts supporting the novel description of a CK biosynthesis pathway in insects.

Author Keywords: Cytokinin Biosynthesis, Drosophila, gene expression, Insect Gall, mass spectrometry, tRNA IPT

Heated leaf extracts of Euphorbia neriifolia (sweet aloes) are used traditionally in Guyana for asthma therapy, but the phytoconstituents have not been studied and phytohormones are generally unexplored in Euphorbia species. Phytoconstituents in asthma-effective traditional heated extracts were analyzed and compared with unheated leaves and latex extracts as screens to identify candidate compounds for asthma treatment. Analysis employed untargeted and targeted mass spectrometry-based metabolomics. The untargeted approach revealed thousands of features in samples with higher abundance in heated extracts and confirmation of 33 metabolite identities (confidence levels 2 and 3) which are known to have antioxidant and anti-inflammatory properties. Highly accumulated and unique features were detected in latex, and those characteristic compounds may reflect the expanded therapeutic uses of E. neriifolia globally. A targeted metabolomic analysis identified and quantified several cytokinins, including aromatic kinetin, and acidic phytohormones. Traditional heating improved the content of several cytokinins with known therapeutic potential, indicating they may be candidates for asthma management.

Author Keywords: asthma, cytokinins, Euphorbia neriifolia, metabolomics, sweet aloes, traditional remedies

Increased plant size is expected to have negative consequences for mating by increasing pollen transfer among the same plant. However, recent theoretical studies have demonstrated that this may not be true for clonal plants. Instead, clonal expansion could enhance outcrossing opportunities without increasing selfing by reducing distances to potential mates. I investigated how the spatial structure of clones influences patterns of pollen dispersal, selfing rates and siring success in a natural population of Sagittaria latifolia. I found that pollen dispersal distances typically exceeded the spatial extent of clones and there was a positive association between clone size and the likelihood that clones were intermingled. Together, this resulted in a weak positive association between clone size and selfing rates, and a strong positive association between clone size and outcross siring success. This is the first empirical support for the theoretical expectation that any negative effects of selfing in large clones might be offset by increased siring success.

Author Keywords: clonal growth, fitness gain curve, geitonogamy, plant mating, plant reproductive ecology, sex allocation theory

Heterosis, expressed as phenotypic superiority over parental species, typically peaks in first generation hybrids (F1s), while later generations (F2 +) exhibit lower fitness. The decrease in hybrid fitness is called hybrid breakdown. The overall incidence of hybrid breakdown in invasive hybrid zones remains poorly understood. The Laurentian Great Lakes (LGL) region contains a hybrid zone comprised of: native Typha latifolia, Typha angustifolia, and hybrid Typha × glauca. F1 T. × glauca display heterosis and are invasive, while later generation hybrids are relatively rare. To investigate possible hybrid breakdown, I compared seed germination and plant growth of backcrossed and advanced-generation (F2) hybrids to F1s and T. latifolia. I found evidence for hybrid breakdown in F2s and backcrossed hybrids, expressed as reduced growth and germination rates. Expression of hybrid breakdown in F2s and backcrosses may explain their relative rarity in the LGL hybrid zone.

Author Keywords: Advanced-generation hybrids, Backcrossed hybrids, Hybridization, introgression, Invasive species, plant competition

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

Interspecific hybridization rates can vary depending on genomic compatibilities between progenitors, while subsequent hybrid spread can vary depending on hybrid performance and habitat availability for hybrid establishment and persistence. As a result, hybridization rates and hybrid frequencies can vary across regions of parental sympatry. In areas around the Laurentian Great Lakes, Typha × glauca is an invasive plant hybrid of native Typha latifolia and introduced Typha angustifolia. In areas of parental sympatry in Atlantic Canada and outside of North America, T. × glauca has been reported as either rare or non-existent. I investigated whether the low frequencies of hybrids documented in Nova Scotia, Atlantic Canada, are influenced by reproductive barriers that prevent hybrid formation or environmental factors (salinity) that reduce hybrid performance. I identified an abundance of hybrids in the Annapolis Valley (inland) and a scarcity of hybrids in coastal wetlands through preliminary site surveys throughout Nova Scotia. In Annapolis Valley populations, flowering times of progenitor species overlapped, indicating that asynchronous flowering times do not limit hybrid formation in this region. Viable progeny were created from interspecific crosses of T. latifolia and T. angustifolia from Nova Scotia, indicating that there are no genomic barriers to fertilization and germination of hybrid seeds. Typha × glauca germination in high salinity was significantly lower than that of T. latifolia, but there was no difference at lower salinities. Therefore, while germination of hybrid seeds may be impeded in the coastal wetlands where salinity is high, inland sites have lower salinity and thus an environment conducive to hybrid germination. However, I found that once established as seedlings, hybrids appear to have greater performance over T. latifolia across all salinities through higher ramet production. Moreover, I found that T. latifolia sourced from Ontario had reduced germination and lower survivorship in high salinities compared to T. latifolia sourced from Nova Scotia, which could indicate local adaptation by T. latifolia to increased salinity. These findings underline that interactions between environment and local progenitor lineages can influence the viability and the consequent distribution and abundance of hybrids. This, in turn, can help explain why hybrids demonstrate invasiveness in some areas of parental sympatry but remain largely absent from other areas.

Author Keywords: flowering phenology, Hybridization, invasive species, physiology, pollen compatibility, salinity tolerance

According to the Invasional Meltdown Hypothesis (IMH), invasive species may interact in their introduced range and facilitate future invasions. This study investigated the possibility that Alliaria petiolata, an invasive allelopathic herbaceous plant in Ontario, is facilitating invasions by additional alien species. Two allelopathic focal species were chosen for this study: the native Solidago canadensis and the invasive A. petiolata. Field surveys in southern Ontario that quantified plant biodiversity in plots that included one or both focal species revealed no support for the IMH, although fewer species co-existed with A. petiolata than with S. canadensis. A year-long recruitment experiment in Peterborough, Ontario, also produced results inconsistent with the IMH, although did provide some evidence that A. petiolata limited recruitment of other species. These results collectively show negative impacts on regional biodiversity by A. petiolata, even in the absence of an invasional meltdown.

Author Keywords: allelopathy, Alliaria petiolata, co-occurrence surveys, invasional meltdown hypothesis, invasive species, Solidago canadensis