Charlotte Ngo MSc Student, Yakimowski Lab Investigating the evolutionary origin(s) of EPSPS copy number variation in Amaranthus palmeri Agricultural use of herbicides has led to the repeated evolution of herbicide resistance in numerous agricultural weeds, posing significant challenges for food production and agriculture sustainability worldwide. The development of herbicide resistance over contemporary timescales has provided us with a unique opportunity to investigate the origins of this adaptation - has herbicide resistance arisen from de novo mutations or from standing genetic variation? Amaranthus palmeri (A. palmeri), a major agricultural weed originating from southwestern United States and Mexico, has been expanding northward, recently was detected in Canada. Many A. palmeri populations exhibit resistance to glyphosate, a common herbicide, primarily through EPSPS gene copy number variation. Yet it remains unclear whether such variation predated the commercialization of glyphosate in 1974. This project aims to answer an important evolutionary question concerning glyphosate resistance in A. palmeri - did the copy number variation of EPSPS exist as standing genetic variation, or did it evolve rapidly in response to glyphosate selection pressure? To investigate the origins of glyphosate resistance, I collected historic samples from the United States and Mexico from herbaria across the United States, spanning three different time periods (pre-1974, between 1974 and 1995, and post-1995), representing a broad geographic distribution. I extracted leaf tissue samples using a modified CTAB protocol, followed by quantification of EPSPS gene copy number using digital drop PCR (ddPCR). The project unveiled intriguing evolutionary patterns that can be explored further through genomic analyses of whole genomes. Given the expanding range of A. palmeri across North America, understanding the evolutionary mechanisms of herbicide resistance is imperative for unraveling the spread of not only A. palmeri but also other weed species. Moreover, the utilization of historical samples in this research serves as an exemplar for studying rapid adaptation to human-induced environmental changes.
Mia Akbar MSc Student, Colautti Lab Examining phenological trade-offs under biotic and abiotic selection in Lythrum salicaria Studies measuring phenotypic selection on day of first flower or “flowering time” in natural populations commonly observe directional selection for early flowering. Paradoxically, phenological responses to climate change are highly variable and do not demonstrate a unilateral shift to earlier flowering time. The timing of flowering is only one part of the overall “flowering schedule” and may trade-off with other adaptive aspects such as duration and peak flowering time, offering one potential resolution to this apparent contradiction. Furthermore, variation in growing conditions across years may maintain variation in the flowering time phenotype and offer an additional explanation for the lack of evolution in response to selection for earlier flowering. Focusing on the North American invasive plant Lythrum salicaria, I use common garden experiments to investigate whether flowering time correlates with other characteristics of the flowering schedule, how variation in annual growing conditions and insect herbivory have the potential to alter the strength of selection on flowering time and whether these relationships differ by latitude of origin. I discovered that detailed metrics of the flowering schedule vary along a latitudinal gradient and that responses to insect herbivory were highly variable across years of the experiment, demonstrating two potential mechanisms that contribute to the “paradox of evolutionary stasis” in the flowering time trait.
Alex Row MSc Student, Tufts Lab Spatial Ecology of Smallmouth Bass (Micropterus dolomieu) in the St. Lawrence River Smallmouth Bass (Micropterus dolomieu) are an important top predator in many freshwater ecosystems in North America. They are also one of the most popular species for recreational anglers. Although there is a lot of research describing the ecology of Smallmouth Bass, most studies have been carried out on populations in relatively small inland lakes and rivers. At present, much less is known about the ecology of this species in aquatic ecosystems at the scale of the Great Lakes and St Lawrence River, which have very different physical attributes and food webs. In recent years, Smallmouth Bass populations in the Great Lakes and St Lawrence River have also been experiencing intense pressure from competitive fishing events (bass tournaments), as well as invasive nest predators (Round Gobies). Since management and conservation efforts are most effective when there is a strong foundation of biological information, there is an urgent need for more information about the ecology of Smallmouth Bass in these unique aquatic ecosytems. On this background, this study uses acoustic telemetry to describe the spatial ecology of Smallmouth Bass in an area where Lake Ontario flows into the St Lawrence River. The annual movements and home ranges reported in this study may be the largest described for this species. The results of this thesis also provide important considerations for assessing, managing, and conserving this population of Smallmouth Bass.
Jamie Would MSc Student, Smol Lab Tracking long-term changes in climate and salinity using cladoceran and other markers from saline lakes in Yukon In the Canadian sub-Arctic, inland saline (athalassic) lakes are rare, as they are typically only found in temperate or equatorial regions with arid climates. These lakes, characterized by unusually high salinities and shallow, closed-basin morphometries, are especially sensitive to climate change as salinity increases markedly with increased evaporation. However, due to their remote location, little is known about the ecology of northern athalassic lake systems and their responses to climate warming. Here, we used paleolimnological approaches to reconstruct multi-trophic level responses to climate warming using biological, chemical and physical information preserved in dated lake sediments. Siliceous algae indicated increasing salinity since the early 1900s, closely tied to rising air temperatures. Cladoceran (commonly known as water fleas) microfossils, representing intermediate-trophic levels, tracked declines in species diversity with increasing salinity. Interestingly, a major lake drawdown (for road maintenance) between 1948 and the mid-1970s did not affect cladoceran assemblage composition (but did result in a change in diatom composition), allowing us to independently separate the effects of lake salinization and water level changes. This research sheds light on the impact of climate-induced salinization on northern lake ecosystems and also demonstrates the value of cladoceran microfossils as paleolimnological indicators of salinity.
Austin Macklem MSc Student, Aristizabal Lab Uncovering the functional consequences of cancer associated histone H2B mutations In eukaryotes DNA is wrapped into chromatin, a nucleoprotein structure that controls DNA-templated processes like DNA replication, transcription and DNA repair. Nucleosomes are the basic unit of chromatin and consist of 146 base pairs of DNA wrapped around two copies of histone H2A, H2B, H3, H4. The increased availability of cancer genomics datasets has revealed a large repertoire of mutations on histone genes and shown that these occur frequently in cancer patients. Although, the significance of the large histone gene mutational landscape remains understudied, some of these mutations (H3K27M, H3K36M and H3G34R) have been shown to drive oncogenesis and are now used as biomarkers of specific tumor types. My project aims to advance our understanding of the functional consequences of cancer-associated histone mutations using Saccharomyces cerevisiae as a model system. To this end, we have mined The Cancer Genome Atlas (TCGA) and identified cancer-associated mutations on histone H2B, a histone that remains poorly characterized with regards to cancer-associated histone mutations. Focusing on mutations that fall on residues conserved between yeast and humans, I have generated a panel of strains carrying only mutant or mutant and wild type version of H2B. These strains will be examined for effects on growth, chromatin structure and function in vivo.
Graydon Gillies MSc Student, Eckert Lab Metapopulation structure and dynamics may maintain a species’ range edge Species range limits may be enforced by various evolutionary and ecological processes.
It is thought that many range limits are imposed by low fitness beyond a species’ range or the inability to disperse to suitable habitat beyond the range. However, transplant experiments and geographic surveys often find that these hypotheses fail to adequately explain species ranges, requiring the use of a more comprehensive framework. The metapopulation-hypothesis outlines how changes in habitat patch colonization rates, extinction rates, and habitat availability may cause metapopulation collapse and the generation of an abrupt range limit. Using coastal dune plant Camissoniopsis cheiranthifolia, I evaluated the metapopulation-hypothesis by conducting a multi-year survey across the species’ northern range. I found that patch structure, including the frequency, size, isolation, and quality of patches, changes towards the northern range edge. Furthermore, I found that colonization rates and habitat availability decline towards the range edge, while extinction rates increase non-significantly. With this survey, I provide the first empirical estimates of metapopulation parameters towards a species’ range edge and demonstrate that metapopulation dynamics may contribute to the maintenance of species’ range limits. Faiqa Amin MSc Student, Monaghan Lab Phosphorylation of the Arabidopsis thaliana E3 ubiquitin ligase ATL6 by the Ca2+-dependent protein kinase CPK4 Post-translational modifications (PTMs) such as reversible phosphorylation integrate signalling and gene expression with cellular metabolic networks and represent some of the earliest responses of plant cells to (a)biotic stress that rapidly controls the functions of many proteins. Ca2+ -dependent protein kinases (CPKs) transduce Ca2+ signals via the catalytic activity of their kinase domain to phosphorylate specific residues of target proteins. Our research discovered that the Arabidopsis CPK isozyme CPK4 phosphorylates the E3 ubiquitin ligase ARABIDOPSIS TÓXICOS EN LEVADURA 6 (ATL6) at multiple residues including Ser278. ATL6 is a positive regulator of immune signalling as it polyubiquitinates CPK28, leading to its proteasomal degradation and subsequent activation of pathogen defense responses. Rabbit polyclonal antibodies were raised against purified, heterologously expressed ATL6 (anti-ATL6), whereas anti-(phosphoSer278-specific) antiserum (anti-pSer278) was raised against a synthetic ATL6 phosphopeptide. These antibodies will augment various biochemical and genetic tools that are being integrated to investigate the functional relationship of CPK4 and ATL6 during Arabidopsis stress signaling. For example, immunoblot time-course assays using anti-pSer278 have established the ATP- & Ca2+ -dependent phosphorylation of ATL6 at Ser278 by CPK4. In vitro E3 ubiquitin ligase assays are being developed to test the hypothesis that CPK4-mediated phosphorylation enhances ATL6’s ability to polyubiquitinate CPK28. Overall, This work will provide an insightful understanding of the role of AtATL6 in biotic or (a)biotic stress responses.
Jules Petrenko MSc Student, Bonier Lab Do urban environments ameliorate range-limiting challenges for urban-tolerant species? Species’ distributions are constrained by challenges which usually increase in severity approaching the range edge. While urban environments might generally be expected to exacerbate such range-limiting challenges, some urban-tolerant species take advantage of the opportunities provided by cities, and persist in urban areas. For these urban-tolerant species, cities might ameliorate range-limiting challenges; however, no direct tests of this hypothesis exist. To test how urban environments affect urban-tolerant species’ distributions, I will compare the relative abundance of North American, urban-tolerant bird species across their distributions in both urban and nonurban habitats. I will use a large community science dataset (eBird Status & Trends Products) to quantify the relative abundance of each species in approaching climatic and geographic range edges of its distribution, in urban compared to nonurban habitats. If cities do ameliorate range-limiting challenges, then species abundance will decline less approaching the range edge in urban habitat, relative to nonurban habitat. Alternatively, if cities do not affect or exacerbate range-limiting challenges, then species abundance will decline at a similar rate or even more steeply approaching the range edge in urban habitat, relative to nonurban habitat. By examining whether relative abundance differs in urban compared to nonurban habitats at the range edge, I will identify whether urban environments provide enough advantages for urban-tolerant species to support persistence among range-limiting challenges. This insight into the effects of urban environments on range limits can increase our understanding of how anthropogenic environments influence species’ distributions.
Harshavardhan Thyagarajan PhD Candidate, Chippindale Lab Intralocus sexual conflict explored through male-limited selection in Drosophila melanogaster Intralocus sexual conflict (IaSC) arises from the opposing forces of sex-specific selection pressures and positive genetic trait correlations between sexes, impacting the evolutionary genetics of sexually reproducing populations. This conflict is implicated in maintaining genetic variance for fitness, shaping trait dimorphism and influencing sex-biased gene expression. However, direct tests of its contributions remain limited and challenging. Here, I present findings from a sex-limited selection experiment using Drosophila melanogaster haplotypes, artificially ‘resolving’ IaSC under male-limited (ML) selection without opposing female fitness selection. I estimate heritable variance for fitness in selected and matched control (MC) lines. In alignment with theory, I observed increased estimates of male fitness and a sharp decline in heritable variance for the same in selected lines. Surprisingly however, I found no significant improvement in selected male fitness compared to controls under generic test conditions, challenging both the findings about genetic variance, and assumptions about ubiquity of IaSC in D melanogaster lab populations. Female fitness of selected haplotypes declined as predicted. Upon further investigation, this system reveals substantial impact of selection from unforeseen sources, likely obscuring our ability to identify adaptive responses to a release from IaSC. I demonstrate evidence of local adaptation to a novel environment of sexual selection and compensatory adaptations in a new genetic background, in the form of nuclear recovery from mother’s curse and adaptations to unique Y chromosomes. These results highlight the complexity of selection experiment design, raising concerns about the generality and accuracy of previous findings.
Nick Smith MSc Student, diCenzo Lab Characterizing bifunctional sugar-processing enzymes from thermophilic bacteria Nucleotide sugars are the donor substrates for glycosyltransferase enzymes, which are widely studied throughout biochemistry, medicine, and glycobiology. However, nucleotide sugars are prohibitively expensive, prompting the development of in-lab chemoenzymatic synthesis protocols to circumvent this cost. One such example is the usage of the bifunctional Fucokinase/GDP-fucose pyrophosphorylase enzyme from Bacteroides fragilis (BfFKP) to catalyze the production of the costly compound GDP-Fucose. The BfFKP enzyme contains a C-terminal Fucokinase domain and an N-terminal GDP-Fucose pyrophosphorylase domain that catalyze sequential steps in the GDP-Fucose synthesis pathway. As a result, this bifunctional protein is used in “one-pot” syntheses, resulting in appreciable amounts of GDP-Fucose for a fraction of the commercial cost. Since the discovery of the BfFKP enzyme, studies have mainly focused on using this enzyme rather than sourcing novel FKP proteins, despite an enormous sequence space of putative FKP orthologs. To address this lack of knowledge, a sequence similarity network (SSN) was used to identify putative bifunctional FKP enzymes. To test the hypothesis that FKP enzymes from thermophilic organisms would exhibit altered properties and/or activities, an uncharacterized FKP sequence from the thermophile Thermophagus xiamenesis (i.e., TxFKP) was selected. I recombinantly overexpressed both the TxFKP and BfFKP enzymes in Escherichia coli and purified the proteins using Immobilized Metal Ion Affinity Chromatography. To compare thermostability between the two enzymes, I used Thermal Shift Assays to approximate their melting temperatures (Tm). Unexpectedly, both proteins demonstrated biphasic melt curves, suggesting that the enzymatic domains are modular enough to unfold at separate temperatures in vitro. In addition, I performed kinetic analyses of the Fucokinase domains of both enzymes, revealing key differences and similarities between the two. These results will shed light on the variance of properties within a fascinating group of biocatalytic tools.
MSc Student, Aristizabal Lab Characterizing the effects of human cancer-associated histone H2AZ mutations using S. cerevisiae Nucleosomes, the building blocks of chromatin, are composed of 146 base pairs of DNA wrapped around an octamer of histone proteins (typically two copies of H2A, H2B, H3, and H4). Chromatin, the packing structure by which DNA is condensed into chromosomes, is dynamic and can be altered in several ways, including: (1) the sliding, eviction, and deposition of nucleosomes, (2) the addition of posttranslational modifications (PTMs) to histone proteins and (3) the incorporation of histone variants in place of canonical counterparts. A wide range of mutations on histone-encoding genes have been identified among publicly available whole-exome sequencing data from human cancer patients. Some of these mutations have been demonstrated to disrupt the histone octamer, alter higher order chromatin structure, and affect histone tail dynamics, DNA accessibility and transcription factor binding. Although most of these mutations remain to be studied, some, termed “oncohistones,” have been shown to drive cancer development and are correlated with poor disease prognosis. My project seeks to advance our understanding of the effect(s) of these cancer-associated missense mutations to the histone H2A variant, H2A.Z, using budding yeast (Saccharomyces cerevisiae) as a model system. H2A.Z is implicated in a wide range of molecular processes, including transcriptional regulation, DNA replication, cell cycle progression, and DNA damage repair. It is also highly conserved from yeast to humans. Focusing on cancer-associated mutations that map to identical residues between yeast and humans, we have generated a library of 21 mutations which will be screened for effects on cellular growth, chromatin dynamics, and gene expression. This work will improve our understanding of the various mechanisms by which cancer-associated mutations affect genome function, providing some clues as to how they may contribute to cancer development and progression.
MSc Student, Snedden Lab Arabidopsis CML13 and CML14 Interact with Myosins and Function as Plant-Specific Myosin Light Chains Calcium ions (Ca2+) are widely present as secondary messengers in eukaryotes. Ca2+-signals are interpreted by Ca2+-binding proteins called sensors, which then regulate various responses. Apart from the highly conserved calmodulin (CaM), plants possess a distinct family of CaM-like proteins (CMLs) that function as Ca2+-sensors. CMLs primarily consist of Ca2+-binding EF-hands and lack any other functional domains. They are believed to act as sensor-relays by undergoing conformational changes induced by Ca2+ and interacting with target proteins. Among the 50 CMLs found in Arabidopsis, AtCML13/14 are particularly intriguing due to their unique biochemical properties and high expression levels in vivo. To investigate the function of CML13/14, we screened a yeast two-hybrid library to identify potential interacting proteins. Our screen led to the discovery of three unrelated families of putative targets: IQ67 domain proteins (IQDs; microtubule scaffolds), CAMTAs (transcription factors), and myosins (motor proteins). These proteins possess a structural characteristic known as tandem IQ-motifs, which are a special type of CaM-binding domain. Through in vitro and in vivo protein-interaction assays, we found evidence suggesting that CML13, CML14, and CaM are the primary interactors of these targets via their IQ domains within the CML family. Focusing on myosins as representative targets of CML13/14, we utilized confocal microscopy, in vitro kinetic assays, and in vitro binding tests to demonstrate that these CMLs act as novel myosin light chains. To gain further insights into their functions in vivo, we employed an inducible RNAi system to specifically silence either CML13 or CML14 in Arabidopsis. The resulting phenotypes were pleiotropic, indicating that these CMLs play crucial roles in development by regulating cytoskeletal function through their interactions with myosins and IQDs. In summary, our data suggests that CML13/14 are important regulators in various biological processes. They modulate cytoskeletal activity via their association with myosins and IQDs, while potentially influencing gene expression through interactions with CAMTAs.
PhD Student, Regan Lab Surveying the Hormonome of Hazelnut Flower Dormancy Hazelnut is an emerging crop in Ontario but elite, European cultivars must adapt to Ontario winters to support local demand. Hazelnut is a winter flowering tree, meaning flowers begin their development in the summer, go dormant over the winter, and bloom in the spring. Brief, early, warm spells are typical of Ontario winters and can signal hazelnut’s male flowers (catkins) to bloom prematurely. Premature bloom makes catkins especially vulnerable to incoming cold weather, damaging pollen and reducing nut yield. To help establish hazelnut as a reliable crop in Ontario, premature catkin bloom must be prevented. Plant hormones are known to regulate flower dormancy and hormones have been manipulated to delay bloom in select crops. This study has surveyed the endogenous levels of canonical plant hormones and their metabolites (the hormonome) of catkins from early, mid, and late-season blooming hazelnut cultivars across a full season of dormancy. This hormonome is the first of its kind within deciduous woody perennials and provides a valuable resource for those studying flower dormancy in fruit crops. Preliminary analysis shows increased ethylene precursor levels in the early blooming cultivar, revealing a potential target pathway for the manipulation of catkin bloom.
Julia Paton, MSc Candidate, Smol Lab Long-term changes in lake ecosystems linked to smelter emissions on the Atikameksheng Anishnawbek First Nations Reserve Mining operations in Sudbury, Ontario, may have caused acidification, metal contamination, and other disturbances on lakes on the Atikameksheng Anishinabek First Nation (AAFN) Reserve. Despite the societal importance of the Reserve’s many lakes, little direct long-term limnological data are available. Here we use paleolimnology and a multi-proxy approach to reconstruct the potential long-term effects of mining operations on water quality and aquatic biota.
Whitefish Lake is a relatively shallow lake that is situated adjacent to the Sacred Lands of the AAFN community and located approximately 15 km southwest of the Vale Copper Cliff Complex. A sediment core was retrieved in September 2022, radioactively dated, and analyzed for geochemistry. Sedimentary geochemical data reconstructions show increased metal inputs linked to mining operations, with arsenic and copper reaching probable effects levels during peak mining emissions, circa 1960s. Further, the sediment chlorophyll a concentration profile records changes similar to mining-impacted lakes of the Sudbury region. Meanwhile, diatom assemblages show only subtle changes in response to mining, indicating that there was no biological evidence of acidification. Round Lake is a relatively deep lake, located approximately 20 km southwest of the Vale Copper Cliff Complex and less than 5 km west of the abandoned Long Lake Gold Mine, which contains over 163,000 m3 of mining tailings. A sediment core was retrieved in September 2022, radioactively dated, and analyzed for geochemistry. Geochemical reconstructions show increased metal inputs linked to mining operations, with arsenic, cadmium, copper, lead, and zinc reaching probable effects levels during peak mining emissions, circa 1960s. Interestingly, and in contrast to Whitefish Lake, Round Lake’s sedimentary chlorophyll a concentration profile showed no change with the onset of mining. The largest shift in biological data shows a striking change in diatom assemblages post-mining; however, these changes were not linked to acidification. This research offers a unique opportunity to collaborate with Indigenous communities and apply western scientific approaches to provide data critical to proper lake management and ultimately protect the societal value of aquatic ecosystems within the Atikameksheng Anishinabek First Nation community. Evan Jones, MSc Candidate, Smol Lab Tracking the long-term limnological impacts of silver mining near Keno City on the traditional territory of the First Nation of Na-Cho Nyäk Dun (Yukon, subarctic Canada) Mining in Northern Canada has caused many major environmental problems; however, historical data are often non-existent. Here, a multi-proxy (metals, bioindicators, pigments) paleolimnological approach is used to reconstruct the historical impacts of mining activity near Keno City, on the traditional land of the First Nation of Na-Cho Nyäk Dun in central Yukon (Canada). Silver was discovered in the Keno region in the early 1900s and intensive mining has taken place ever since.
Christal Lake, a shallow water body, lies near many historical and current mines, and was once the site of a processing mill. A sediment core was retrieved from Christal Lake in September 2022. Geochemical data from the dated sediment core were used to reconstruct metal inputs linked to the mining activity. The largest shift in biological indicators was a striking decline in sedimentary chlorophyll-a concentrations, indicating declining algal populations. Meanwhile, subfossil diatom assemblages only changed subtly in response to mining. There was no biological evidence of acidification, likely due to the neutralizing effect of the carbonate-rich catchment. The Hanson lakes are situated ~10 km from Keno City and are outside the Christal Lake watershed. These lakes are being studied to determine the potential extent of aerial deposition of mining contaminants. A sediment core was retrieved from a basin in the Hanson lakes system in October 2023. Preliminary results indicate that, despite proximity to mines, it is climate that is the major driver of ecological change in this system. Collectively, the data from these two sites help document the long-term impacts of silver mining in this subarctic environment. Yael Lewis, MSc Candidate, Orihel Lab The effects of microplastics on zooplankton and emerging insect communities in a littoral limnocorral experiment Although microplastics research has rapidly accelerated in the last few years, our understanding of their effects on freshwater invertebrates, particularly under environmentally relevant conditions, remains limited. Using in-situ limnocorrals at the International Institute for Sustainable Development – Experimental Lakes Area (IISD-ELA), we investigated the effects of a microplastic mixture on zooplankton and emerging insect communities. We installed 12 open-bottom limnocorrals in the littoral zone of a boreal lake in June of 2022 and added an environmentally relevant range of microplastic concentrations in a regression-based design. Our microplastic mixture consisted of distinctively coloured polystyrene, polyethylene, and polyethylene terephthalate fragments in equal parts by count, manufactured with common plastic additives. I sampled the zooplankton and emerging insect communities pre-addition and every week thereafter for the 8-week duration of the experiment. To evaluate possible longer-term effects, I sampled the emerging insect community again in May of 2023 over the partially enclosed bases of the limnocorrals. I assessed the relationship between nominal microplastic concentration and the abundance and community composition of zooplankton and insects. Microplastic concentration was negatively related to total zooplankton abundance at weeks 1 and 5 post-addition and drove changes in community composition at week one. Evidence for microplastic effects on the emerging insect community was more limited – microplastic concentration was negatively related to insect emergence only at week 2, and the total seasonal emergence exhibited only a weak relationship with microplastic concentration. There were no effects of microplastic on insect community composition. I anticipate my results will inform policy regarding ecological risk thresholds for microplastics in freshwater ecosystems.
The Departmental I-EDIAA committee will be screening the 30-minute film ‘Signal Fire’. This film progressed out of the working group composed of Canadian scientists and Indigenous elders and scholars that published “Towards Reconciliation: 10 Calls to Action to Natural Scientists Working in Canada” in Facets (Wong et al., 2020).
More information about the film, and its trailer, can be found at: https://www.signalfirefilm.ca/watch Following the screening, everyone is welcome to participate in a discussion (~20 minutes). Those interested will break into small groups (using several rooms on the 3rd floor) and will be provided a few questions to guide reflection and discussion. We look forward to seeing everyone at this screening and discussion. Zoe Kane, PhD Candidate, Smol Lab Using changes in Cladocera assemblages to determine how ornithogenic inputs structure freshwater ecosystems Seabirds can be considered biovectors, transporting large concentrations of nutrients (e.g., nitrogen (N), phosphorous (P)) and metals (e.g., Cd, Zn, Hg) from their marine feeding grounds to their terrestrial breeding grounds. Seabird fertilizes their nesting sites by depositing feces, feathers, carcasses, and eggshells, which can be tracked directly (e.g., sterol/stanol to characterize guano deposits) or from their influence on nearby waterbodies (e.g., nutrient enrichment and/or pH changes traced using subfossil algal assemblages over time). Recent paleolimnological studies from PEARL reconstructed and hindcasted the population dynamics of Baccalieu Islands vulnerable Leach's Storm petrel population (Hydrobates leucorhous, hereafter LESP) to assess the effects of natural and anthropogenic stressors.
Adding to the several paleolimnological proxies used to track and reconstruct historical LESP inputs on ponds on Baccalieu island (Lunin, Brister, Gull and Mainland Reference ponds), I have implemented Cladocerans, or water fleas, which are well-known paleolimnological indicators (but as yet unexplored in these novel ornithological-limnological studies) as their species-specific exoskeletal remains are well preserved in sediment. They have a pivotal role in aquatic food webs, occupying an intermediate trophic position between top-down regulators and bottom-up factors, providing a critical link between the eutrophication process and the implications of elevated nutrient conditions on higher trophic levels. My results build on previous work that examined how seabird-derived nutrients influence primary production and algal assemblages in highly impacted ponds. I demonstrate that shifts from littoral/benthic to pelagic cladoceran taxa coincide with peaks in the inferred LESP population, highlighting the importance of seabird inputs to limnological conditions and eutrophication in several ponds on Baccalieu Island. Christina M. Tschritter, PhD Candidate, Lougheed Lab Mapping pathogen distributions and population connectivity of a sentinel Arctic species, the polar bear (Ursus maritimus) across a changing North American Arctic Large-scale environmental shifts are expanding pathogen distributions making many northern species more vulnerable to disease. To understand such rapidly changing host-pathogen dynamics and potentially mitigate impacts of novel pathogens on northern peoples and ecosystems, I quantify population connectivity and pathogen presence in a sentinel Arctic species, the polar bear (Ursus maritimus). Polar bears are apex predators that can provide insight into pathogen distribution and prevalence across marine and terrestrial ecosystems. The focus of my dissertation was to develop molecular tools to monitor polar bear populations and to promote co-management through non-invasive and harvest-based sampling. More specifically I aimed to: (i) Delineate polar bear population structure using genome-wide panels of Single Nucleotide Polymorphic markers (SNPs) to interpret population connectivity that might impact pathogen spread; (ii) Develop and validate a sensitive multiplexed, magnetic-capture, and digital PCR tool for surveillance of five zoonotic pathogens (three bacteria Erysipelothrix rhusiopathiae, Francisella tularensis, and Mycobacterium tuberculosis complex (MTBC), and two parasites T. gondii and Trichinella spp.) relevant to wildlife and human health; and (iii) Quantify the spatial distributions of focal pathogens in polar bear tissues and observe associations between pathogen detections and predictors. Despite the mobility of polar bears and their large home ranges I found three population clusters that coincide with Arctic ice ecoregions. I made novel pathogen detections (first detection of E. rhusiopathiae in a polar bear, first molecular detection of F. tularensis in the tundra, and the first detection of a MTBC member in Arctic wildlife) and provide insights on how populations might respond to future exposure to novel pathogens. Overall, we found that harvest season and human settlements were important predictors of presence for some pathogens. I envision the establishment of a long-term harvest-based monitoring program that incorporates the powerful molecular tools that we have created, enabling territorial governments to monitor changes in prevalence and/or the geographic advance of select pathogens. The continuation and expansion of this work into a monitoring program would present an unprecedented opportunity to provide critical, real-time, and community-based disease surveillance across the Arctic and ultimately improve opportunities for the co-management of the polar bear species.
Rui Huang, PhD Candidate, diCenzo Lab and Snedden Lab Characterizing the functional impact of Sinorhizobium meliloti bacA allelic variation on symbiosis with legume hosts The symbiosis between legume plants and rhizobial bacteria are highly specific and require both partners to be compatible with each other during the process of symbiotic development. A range of host and symbiont properties regulate the specificity of these relationships. In certain clades of legume plants, the rhizobial BacA transporter and legume-expressed NCR peptides are thought to play a role in defining host/symbiont compatibility. The inverted repeat-lacking clade (IRLC) of legumes expresses NCR peptides to initiate and maintain terminal differentiation of rhizobia, a developmental process that leads to mature, nitrogen-fixation bacteroids. The BacA transporter is essential in transporting NCR peptides and protecting the rhizobia from the antimicrobial activity of these peptides. A previous study found that replacing the bacA allele of Sinorhizobium meliloti with the ortholog from Rhizobiumleguminosarum allowed symbiosis with the host Melilotus officinalis but not with Medicago sativa. In this thesis, I investigated the mechanisms underlying the symbiotic specificity of bacA alleles. On the host legume side, the nodule transcriptomes of M. sativa and M. officinalis were assembled, and their NCR peptide transcripts were identified. The transcriptomic data revealed an approximately 2-fold greater abundance of transcripts encoding highly cationic NCR peptides (isoelectric point > 9.5), which are known to have antimicrobial properties, in M. sativa vs M. officinalis. I hypothesize that the difference in the abundance of highly cationic NCR peptide contributes to why the R. leguminosarum BacA can support symbiosis with M. officinalis but not M. sativa. In addition, I hypothesize that unlike the S. meliloti BacA, the R. leguminosarum BacA is unable to efficiently transport all of the highly cationic NCR peptides of M. truncatula, resulting in a build-up in the bacterial periplasm, leading to cell death. On the rhizobia side, I combined genetic analysis and computational modelling to better understand the BacA transporter. Site-directed mutagenesis analyses showed that S. meliloti
Q193G and N312G BacA mutants recreated the symbiotic phenotypes of S. meliloti carrying the R. leguminosarum bacA allele when paired with hosts M. sativa and M. officinalis. In addition, I generated mutant libraries of S. meliloti and R. leguminosarum bacA through error-prone PCR that identified amino acid residues important for BacA function. Together with BacA structural models that I generated, these data provide novel insight and hypotheses into the functional differences between S. meliloti and R.leguminosarum BacA and their ability to transport NCR peptides. Collectively, the data presented in this thesis provides a deeper understanding of how BacA and NCR peptides may contribute to host/symbiont compatibility in rhizobium – legume symbioses. AAKANX PANCHAL - MSc Candidate Tackling the rhizobium competition problem by engineering exclusive compatibility between legumes and inoculant rhizobia Legume plants can benefit tremendously from their relationship with nitrogen-fixing soil bacteria called rhizobia. Within structures rhizobia induce along legume roots called nodules, rhizobia fix atmospheric dinitrogen into ammonia. High nitrogen-fixing rhizobium inoculants can provide legume crops with enough bioavailable nitrogen under ideal conditions to limit nitrogen fertilizer use and its negative environmental impacts. However, low nitrogen-fixing rhizobia native to the soil can outcompete high nitrogen-fixing rhizobia for nodule occupancy, reducing the benefit to the legumes and ultimately perpetuating dependence on nitrogen fertilizer. A potential solution to this "rhizobium competition problem" is engineering exclusive compatibility at the level of nodule occupancy between high nitrogen-fixing rhizobium inoculants and their associated legume crops. Nodule occupancy begins with rhizobia producing Nod factors specific to legume Nod factor receptors, where Nod factors and Nod factor receptors are like keys and locks. It is the ability of low nitrogen-fixing rhizobia native to the soil to produce the same Nod factors as high nitrogen-fixing rhizobium inoculants that allows low nitrogen-fixing rhizobia to nodulate the legume. Exclusive compatibility between the rhizobium inoculant and legume crop would involve inoculant rhizobia producing Nod factors that are unique in the target environment and legumes producing matching Nod factor receptors, such that low nitrogen-fixing rhizobia producing the original Nod factors should no longer be able to nodulate the legume. To test the feasibility of using this strategy, we have been working towards genetically engineering a model rhizobium-legume pair, Sinorhizobium meliloti and Medicago truncatula, to use Nod factors and Nod factor receptors from a different rhizobium-legume pair, Mesorhizobium japonicum and Lotus japonicus. In my seminar, I will present our progress in bringing the Lotus japonicus Nod factor receptor genes together into a single construct for their eventual expression in Medicago truncatula.
MSc Candidate, Aristizabal Lab Understanding the regulation and function of Cdk8 Cdk8 is a conserved protein kinase and a member of the Mediator transcription co-activator complex. Accumulating evidence highlights Cdk8 as a crucial oncogene in colorectal cancer, emphasizing the need to understand its function and regulation. Notably, CDK8 is amplified or overexpressed in approximately 60% of colon cancer tumors, and elevated CDK8 levels correlate with tumor stage and reduced patient survival. Supporting its oncogenic role, knockdown of CDK8 diminishes the proliferation of colon cancer cell lines, while overexpression induces tumor formation in immunodeficient mice, effects that depend upon Cdk8's kinase activity. Beyond colorectal cancer, growing research also implicates CDK8 in melanoma, leukemia, as well as breast, pancreatic, and prostate cancer, evidence that has stimulated the development of Cdk8 inhibitors for cancer therapy. Despite a growing interest to target Cdk8 for anti-cancer treatment we have limited information about Cdk8 regulation, a knowledge gap that may complicate efforts to block its activity. For my exit seminar, I will discuss my work using the budding yeast model system to understand the function and regulation of Cdk8 by leveraging disease-associated variability and recent insight from structural and biochemical analyses. Specifically, I have leveraged data from The Cancer Genome Atlas and ClinVar to identify CDK8 missense mutations that are likely to affect function. This work led me to find a cluster of mutations that localize to the kinase ATP binding pocket or sites of interaction with CycC and Med12, proteins known to regulate Cdk8 kinase activity. My findings so far suggest that most of the mutants disrupt Cdk8 function, resulting in a feedback mechanism that leads to an increase in Cdk8 protein and mRNA levels. In addition, my work examining the role of Med12 in regulating Cdk8’s activity in vivo showed that this interaction contributes to Cdk8 function in a condition specific manner. Collectively, this research sheds light on the function and regulation of Cdk8, work that will inform the development of targeted therapies and enhance our understanding of the mechanisms by which Cdk8 contributes to cancer development.
MSc Candidate, Lougheed Lab Mapping the Contact Zone of Two Chorus Frog Mitochondrial Lineages in Southern Ontario Contact zones facilitate the study of diverging evolutionary lineages in primary (evolved in situ) or secondary (evolved in allopatry) contact; outcomes of contact zone dynamics have implications for species range limits, can clarify questions regarding species delineation and speciation itself, and may be relevant for conservation. Chorus frog populations within Ontario and Quebec (currently collectively classified as Pseudacris triseriata) possess distinct mitochondrial haplotypes representing Boreal (P. maculata) and Western Chorus Frogs (P. triseriata). These diverging mitochondrial lineages are in secondary contact in Southern Ontario, yet the exact location of contact is uncertain, lying somewhere between Toronto and southern Georgian Bay (a linear span of ~150km). I used environmental DNA (eDNA) from water sampled at ~50 chorus frog breeding ponds to better delineate the chorus frog contact zone. Environmental DNA facilitates extensive geographic sampling, allows for the identification of single wetlands with co-occurring populations, and is less invasive than tissue sampling (e.g. toe clipping, buccal swabbing, lethal sampling). I used droplet digital PCR with custom primer-probe mitochondrial cytochrome b sets that target each lineage (one primer-probe set for each lineage). My data refine the location of the contact zone and identify at least three ponds with both lineages present. My findings set the stage for future work in hybridization and speciation, and have implications for conservation and designations by the Committee on the Status of Endangered Wildlife in Canada (COSEWIC). My research also showcases the effectiveness of eDNA in mapping the distributions of cryptic or secretive species or evolutionary lineages.
Troy Martin, MSc Candidate, Arnott Lab Effects of 'eco-friendly' road de-icer alternatives on freshwater ecosystems Increasing use of de-icing salt on roads and paved surfaces is contributing to rising salinity in freshwater, threatening aquatic ecosystems. In response, novel road de-icers advertised as “eco-friendly” have been developed and are widely used in large cities across North America. Despite this, research on road salt alternative toxicity rarely extends beyond individual species, and community and ecosystem-level testing remain limited. We used outdoor mesocosms to test how zooplankton communities, important primary consumers in aquatic systems, responded to three de-icers: rock salt (NaCl), an organic alternative (beet-brine), and an inorganic alternative (NaCl, CaCl2, MgCl2). We found that both alternatives were toxic to zooplankton and decreased total abundance and richness of communities. Cladocerans and copepods were more sensitive than rotifers for all de-icers, indicating differential sensitivities that would impact community composition, species interactions, and ecosystem function. We further saw impacts at the zooplankton species level and at the bacterial community level, indicating that de-icers might have impacts at various trophic levels. These results are worrisome because they suggest that some de-icer alternatives might not be as “eco-friendly” as advertised, despite their growing use across North America.
Emma Graves, PhD Candidate, Smol Lab Assessing long-term anthropogenic impacts on lakes within the Rideau Canal system: A paleolimnological assessment The Rideau Canal was constructed in the early-1830s, primarily as a means to transport military personnel, but is now exclusively recreational. The construction of the canal and associated flooding, as well as other land-use changes, likely impacted lakes within the system, but long-term monitoring data are not available. Furthermore, recent environmental changes, including accelerated climate warming, have likely affected lake ecosystems. Paleolimnology can be used to reconstruct these missing data sets. Earlier diatom-based paleolimnological studies were conducted on lakes within the Rideau Canal system ~25-30 years ago and more recently by K. Balasubramaniam (MSc, 2022). However, nothing is known concerning the long-term changes in primary consumer (i.e., Cladocera) assemblages linked to the various environmental disturbances. Using the biological information stored in dated sediment profiles from five lakes within the Rideau Canal system (Opinicon, Upper Rideau, Lower Rideau, Big Rideau, and Indian), my study examines how cladoceran assemblages have changed over the past ~250 years in response to these multiple environmental stressors. Changes linked to canal construction and subsequent flooding are recorded in the sediment profiles. However, the most striking shifts in both diatoms and Cladocera have occurred since the 1970s, with assemblage changes linked to accelerated climate warming. These changes will likely have cascading effects on other ecosystem services.
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