Biology

Cytokinins (CKs) are adenine derivative molecules that are present in all kingdoms of life. CKs are known to have a role in cell growth and development in plants. However, the role of CKs in vertebrate systems is not well understood. Frog virus 3 (FV3) is a type species of the Iridioviridae family, genus Ranavirus. FV3 is a major contributor to the amphibian population decline in North America. In this study, we demonstrate that concurrent and pretreatment of 20 µM of either N6-isopentyldenine (iP), N6-ispoentyladenosine (iPR), N6-furfurladenine/kinetin, and N6-furfurladenosine/kinetin riboside (KR) decreased FV3 replication. To understand the mechanism of inhibition, we assessed morphological changes in host cell nuclei to assess the effect of CKs on infected nuclei. Our results show that infection with FV3 and 20µM treatment of iP or iPR reduced nuclei size. These results are the first to reveal insight into the potential mechanism in which FV3 replication is inhibited by iP and iPR.  

Author Keywords: cytokinins, frog virus 3

Northern flying squirrels (NFS) are mycophagous specialists (fungi-dominated diet) thatmay be displaced with southern flying squirrel (SFS) range expansion, thereby limiting fungal dispersal in forest communities. To understand the implications of squirrel species turnover on mycophagy, we investigated the home ranges of both flying squirrel species who are living in stable sympatry. We found no significant difference in home range sizes and identified spatial overlap between the two species. Through habitat selection ratios we found SFS were strongly selecting for deciduous-dominated habitats more than NFS. Lastly, we conducted microscopy on flying squirrel scat and found NFS were eating more fungi than SFS. We conclude that the squirrels are sharing the same habitat landscape but are finding ways to partition the habitat accordingly to allow for sympatry. SFS may contribute to the spore-dispersal cycle similarly to their northern counterpart through moderate fungus consumption and large home range sizes.

Author Keywords: diet, flying squirrels, Glaucomys, home range, mycophagy, sympatry

Climate change is a major conservation concern, especially for many cold-adapted species. The rate of warming due to climate change will likely outpace adaptive responses, and many populations will likely need to rely on phenotypic plasticity to cope with environmental warming. It is currently unclear whether plasticity in physiological responses to warming will be sufficient to offset the negative consequences of chronic environmental warming in ectotherms. I studied within-generation and transgenerational plasticity in two cold-adapted species of fishes, lake trout (Salvelinus namaycush) and brook trout (S. fontinalis), following temperature acclimation. Adults of both species were acclimated to either cold or warm temperatures and offspring were generated using a fully factorial breeding design, whereby the family thermal histories included crosses made within each temperature treatment and bidirectional crosses between temperatures. Offspring families were subdivided into two groups and acclimated to either warm or cold temperatures, so that offspring thermal experience matched or mismatched that of one or both parents. Offspring metabolic rate and critical thermal maximum during an acute thermal challenge were measured for both species. Limited transgenerational plasticity was detected in both species, but had a lesser effect than within-generationacclimation. In brook trout, the paternal contribution was greater than the maternal contribution. In lake trout, a mismatch in thermal acclimation, where the offspring were cold-acclimated but the parents warm-acclimated, resulted in elevated offspring metabolic rate without a corresponding increase in growth, suggesting that a mismatch in temperatures across generations could be detrimental to offspring. Using RNA-sequencing, transgenerational plasticity was linked to differential gene expression in the liver of lake trout offspring, in that genes were differentially expressed depending on the parental acclimation temperatures. Within-generation warm acclimation had the greatest effect on gene expression profile of offspring, with more genes differentially expressed under conditions of within-generation warm acclimation compared with transgenerational warm acclimation. Although it has been suggested that transgenerational plasticity may help to buffer the impact of warming due to climate change, my work implies that transgenerational plasticity, like within-generation plasticity, will be insufficient for these two species of cold-adapted salmonids to cope with climate change.

Author Keywords: Brook trout, Climate change, Lake trout, Phenotypic plasticity, Thermal tolerance, Transgenerational plasticity

Acoustic communication is vital for mediating interactions between individuals and their environment. From echolocating bats to isolated rodent pups, ultrasonic vocalizations are a useful mechanism for producing localized and cryptic acoustic signals. Flying squirrels have been demonstrated to produce almost exclusively ultrasonic calls which is rare in both squirrels and mammals, though the significance of this acoustic range is unknown. To investigate the context of ultrasonic calls in these gliding mammals, I created phylogenetic comparative analyses to compare ecological and morphological traits against call frequencies. I found that nocturnality, a trait entangled with gliding in squirrels, was related to higher-frequency calls. Furthermore, by comparing all gliding mammals and their non-gliding counterparts, I found that gliding mammals produce significantly higher frequencies than these relatives. This form of cryptic communication is likely used to avoid predation, which was further supported by behavioural experiments wherein flying squirrels used significantly higher frequencies when predation risk increased. While high frequency communication was used by most gliders, I found that ultraviolet-induced photoluminescence, another potential form of crypsis, was strongly associated with nocturnality, with half of the tested gliding mammals showing evidence of this unique colouration. While ultrasonic vocalizations are widespread in echolocating bats and echonavigating mammals, I did not find evidence to support echonavigation in flying squirrels. Instead, I found that ultrasonic vocalizations are used in a variety of social contexts and during solitary foraging and exploration. These combined results demonstrate the unique ecological and evolutionary pressures acting on small-bodied, nocturnal gliding taxa and the resulting cryptic behaviours and communication.

Author Keywords: communication, flying squirrels, gliding mammals, Sciuridae, ultrasound, ultraviolet-induced photoluminescence

Fungi are a diverse group of organisms that play crucial roles in various ecological processes and have immense economic importance. Understanding the intricate mechanisms underlying fungal growth and development is fundamental to harnessing their potential and exploring their applications in different fields. Signalling molecules, such as hormones, have been identified as key regulators in fungal physiology, orchestrating intricate processes and modulating biological responses to the environment. Phytohormones, commonly associated with plants, have been proposed as potential myco-hormones due to their production in a wide variety of fungi. Metabolomic analyses were performed via LC-MS/MS to investigate the role of phytohormones, specifically cytokinins (CKs) and indole-3-acetic acid (IAA), along with lipids and energetic metabolites such as organic acids during the growth and development of the model fungus Sordaria macrospora. The results revealed a clear switch between CK ribosides and CK free bases during the ascosporegenesis stage, with increased levels of cZ and iP and decreased levels of iPR and cZR. A similar pattern was observed in the mutant strain smgpi1 but demonstrating higher levels of CK free bases and increased fruiting body formation compared to the wild type. These findings provide insights into the regulation of phytohormones especially during fungal fruiting body development. In terms of IAA, the levels increased during the transition to sexual development in all strains, with the per5 mutant, unable to produce lipids via the cytosol, demonstrating a higher concentration than the wild type. The interplay between energetic metabolites and IAA suggests a potential role in the transition to sexual development. Additionally, the dose-dependent effects of exogenous CK application were investigated, showing the potential of low concentrations, from 1 to 10 μM, of CKs in promoting biomass accumulation or sexual development. Furthermore, gene editing in S. macrospora was proposed as a future direction to explore the functions of CKs and other metabolites during fungal development. Overall, this research contributes to our understanding of phytohormone-mediated processes in fungi and opens avenues for future investigations in fungal biology.

Author Keywords: fungi, LC-MS, metabolomics, myco-hormone, phytohormone, sexual development

Migration, space use, and phenology play key roles in shaping avian populations and are therefore critical for conservation. My thesis examines the migration strategies and non-breeding spatial requirements of Virginia's beach-nesting American Oystercatchers (Haematopus palliatus), as well as the nesting phenology of mid-Atlantic American Oystercatchers and Piping Plovers (Charadrius melodus). I assessed migratory decisions using field-readable bands and GPS transmitters, finding that ~90% of Virginia's beach-nesting American Oystercatchers migrate out-of-state during the non-breeding season, though tested hypotheses did not strongly predict migration patterns. I then estimated home range sizes of American Oystercatchers during the non-breeding season, revealing high individual variation (12.1-201.6 km²) and a potential trend toward larger home ranges in males. Finally, I analyzed clutch initiation timing of American Oystercatchers and Piping Plovers in response to climate change, finding that American Oystercatchers advanced timing of breeding by 10.7 days between 2005-2022, while Piping Plovers showed no significant shift.

Author Keywords: behavioural ecology, climate change, home-range, migration, movement ecology, phenology

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

I investigated the climate vulnerability of a cold-water salmonid by observing the upper thermal tolerance of brook trout (Salvelinus fontinalis) across multiple life stages. Using hatchery- and wild-ancestry fish, I assessed variation in thermal tolerances and carryover effects within and among brook trout life stages and populations that were reared under varying winter thermal regimes as embryos. Embryonic, fry, and gravid adult upper thermal tolerance were tested via CTmax. Warm acclimation was associated with an increase in embryonic upper thermal tolerance. CTmax variability was markedly higher in embryos than juvenile or adult salmonids. Effects of thermal incubation treatments varied by life stage, with incubation temperature and life stage both influencing body size and thermal tolerance. Collectively, these results suggest that brook trout only exhibit short-term carryover effects from thermal stressors during embryo incubation, with no lasting effects on phenotype beyond the first few months (10 weeks) after hatch.

Author Keywords: acclimation response, climate change, ontogeny, salmonid, stress, thermal plasticity

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

Cadmium is a toxic metal that has detrimental effects on blood vessel development and function. To examine the effect of varying concentrations of cadmium on angiogenesis, two in vitro assays were used. First, developing yolk sac blood vessels were studied in gestation day 8 mouse embryos exposed to medium alone, 1.25, or 1.75 μM cadmium chloride (CdCl2). Embryos exposed to 1.25 μM cadmium experienced a significant increase in the number of vessels formed; however, they were smaller in size. Vessel morphology and signalling pathways were also investigated using the mouse aortic ring assay, with exposures of 0.0, 0.5, 1.0, 5.0, or 10.0 μM CdCl2. Samples exposed to 10 μM experienced a significant increase in vessel length. However, no significant differences in phosphorylated PTEN and AKT were observed. The results of this study suggest that low levels of cadmium may disrupt angiogenesis, particularly the development of the embryonic vasculature in the yolk sac.

Author Keywords: Angiogenesis, Cadmium, Embryonic Development, Teratogenicity, Vascular Development, Vasculogenesis