Arsenic transfers and toxicology are important topics of research and a public health concern because arsenicosis affects millions of people worldwide every year. One of the most significant sources of arsenic in the environment is industrial wastes, such as by-products of mining operation. In Yellowknife, Northwest Territories, Canada, there were two large gold mines—Giant Mine and Con Mine, along with dozens of small-scale mines. The combined by-product of emission from these roasters might have contributed to high concentrations of arsenic found in the city. This thesis presents the results of two related studies to address the environmental health concern: (1) to investigate the arsenic transfers and arsenic species accumulation in freshwater food webs near large legacy mining operations in Yellowknife, and (2) to assess the long-term health risk of inorganic arsenic exposure from the consumption of fish in Yellowknife among the general residents and the Yellowknives Dene First Nation. We found that inorganic arsenic is biominified in food webs (i.e. inorganic arsenic concentrations diminish at higher trophic positions relative to lower trophic positions). Higher-trophic organisms have low inorganic arsenic concentrations in tissue due to biotransformation of inorganic arsenic to non-toxic organic arsenobetaine, and effective elimination of arsenic from their tissue. The trophic positions of freshwater organisms can be used to predict the range of arsenic concentrations and its species composition, accounting for more than 80% of variance. Dietary study results show that the Yellowknives Dene First Nation consumed significantly more fish in their diets (adults: 19 g/day, children: 9 g/day) compared to the general residents of Yellowknife (adults: 9 g/day, children: 5 g/day). Our probabilistic risk assessments showed no significant long-term non-carcinogenic and carcinogenic health risks of inorganic arsenic exposure from fish consumption for the majority of Yellowknife residents, but elevated cancer risks among the adult heavy fish consumers in Yellowknife. However, our data suggested that the residents of Yellowknife were not exposed to higher cancer risks from inorganic exposure compared to the general population in Canada. Therefore, due to fish health benefits and the values associated with its consumption, fish should continue to be a major source of sustenance in Yellowknife.

Microplastic particles (MPs) are widely distributed in aquatic environments and present a potential risk to marine life. This thesis considers several issues relating to methodologies for sampling and analyzing MPs and the sources and fate of these particles in the marine environment, wild and farmed shellfish, and food webs of southern coastal British Columbia, Canada. Chapter 1 introduces MPs as a contaminant, methodologies for studying them, and reviews what is known about their source, fates, and ecotoxicology in marine environments. Chapter 1 also outlines the goals of this thesis. Chapter 2 compares the potential MP concentration estimates provided by two different seawater sampling methods. Jar samples filtered to 8-µm yielded MP concentrations averaging approximately 8.5 times higher than bucket samples filtered to 63 µm, per L of water (at the site level), driven largely by differences in the number of microfibres. An analysis of MP concentrations and mesh sizes reported in the literature suggests that using a 300–350-µm mesh may underestimate total MP concentrations by one to four orders of magnitude compared with samples filtered through much smaller mesh sizes (e.g., less than 100 µm), and despite the effect of sample volume. Particles less than 300 µm in diameter make up a large component of MPs commonly found in fish and invertebrates. As such, common sampling practices fail to adequately measure a biologically relevant class of MPs, thereby undermining the ability to quantify ecological risk. Chapter 3 determines the influence of shellfish aquaculture activity, and its use of plastic equipment, on MPs in bivalves and their environment by comparing MP concentrations in Manila clams (Venerupis philippinarum) and Pacific oysters (Crassostrea gigas) grown on commercial shellfish beaches with those grown on non-aquaculture beaches from six areas. MP concentrations in water and sediment were also determined in four of the areas. MP concentrations did not differ between shellfish aquaculture and non-aquaculture sites for either bivalve species or for sediment and water samples. Beach sediment type had a minor effect, with more gravelly or sandier beaches associated with higher MP concentrations in oysters or clams, respectively. Oysters on sites using many synthetic anti-predator nets had more MPs than those on sites without any plastic, but analysis of particles using Fourier-transform infrared spectroscopy suggested a predominance of textile fibres including nylon and polyester, which are not typically used in shellfish aquaculture. Chapter 4 uses stable isotope food web analysis and hierarchical Bayesian generalized linear mixed models to explore whether bioaccumulation and biomagnification are occurring in coastal marine food webs at three locations. Bioaccumulation was higher for smaller-bodied suspension feeding animals such as bivalves. However, biomagnification was not occurring in animal digestive tracts, and trophic dilution was demonstrated in fish livers. Trophic transfer was shown to occur between prey and predator for rockfish, but higher concentrations in full stomachs compared with empty ones suggested rapid excretion of ingested MPs. Chapter 5 supplies some general conclusions on the status of MPs in the British Columbian environment, as well as risks to seafood consumers. It also explores future work that will be needed to understand the complex ecotoxicology of MPs. / Graduate / 2022-04-12

Arctic tundra wetlands support abundant waterbirds, but invertebrate prey communities may change with climate warming. Increased influx of nutrients and labile dissolved organic matter (DOM) from thawing permafrost may alter the relative importance of organic matter (OM) sources, with associated changes in relative biomass of taxa dependent on different sources. In six wetland types, we used stable isotopes (δ13C, δ15N) to compare contributions of four OM sources (periphytic microalgae, cyanobacteria, macrophytes, and peat) to the diets of nine macroinvertebrate taxa. Relative OM contributions within invertebrate taxa were similar among wetland types. Cyanobacteria comprised only 2–7% of OM sources for all taxa in shallow wetland types (<1 m), but up to 25% for oligochaetes and Physidae in deeper wetlands. Macrophytes were minor OM sources (<13%) in all wetland types except deep open lakes (21–26%). Peat typically comprised 20–40% of OM sources except for Physidae (mostly 50–80%). Microalgae were the dominant OM source for most taxa (47–78%, mean ⁓60%), although less for Oligochaeta and much less for Physidae (9–32%). High periphyton production with very depleted δ13C values likely results from continuous daylight illuminating shallow depths, high N and P levels, and very high pCO2 derived from bacterial respiration of DOM leached from thawing permafrost. Invertebrate consumption of microalgae and peat appears often to involve bacterial intermediates. Impacts of warming on invertebrate prey availability will likely depend not on shifts in OM sources, but on changes in overall area or number of shallow ponds.

The cold water of Antarctica has a unique endemic fauna, where durophagous predators are rare or absent. Due to climate change the water is heating up and the predators have begun to return to the Southern Ocean, which could bring a lot of changes to the food web. There is a high risk it will lead to losses in the unique marine fauna of Antarctica. The aim of this study is therefore to examine the potential effect these invasive species has on the food web structure in the Weddell Sea. To study this, several general network metrics were used (connectance, number of interactions, vulnerability and generality, trait distributions), as well as a number of centrality metrics (betweenness, closeness, PageRank). The analyses showed that none of the invasive species become important in the Weddell Sea food web. Nor do they significantly change the food web structure in any way which impact the importance of the native species. Their great opportunism regarding their prey species, and thereby their connectedness and thus their position in the network, are probably the main reason why theses invasive species did not become important in this food web. The lack of changes in the food web structure due to the presence of these invasive species are probably also a result of not including factors such as abundances and network dynamics in the analyses, which seem to be the driving forces when it comes to changes in food web structure caused by invasion of species.

The Arctic is warming faster than any other region of the globe. To conserve and manage many thousands of lakes across arctic landscapes, scientists need to understand historic and present conditions within these lakes to predict how the lakes, and the organisms that inhabit them, may respond to a changing climate. The goal of my research was to improve our understanding of what physical, chemical, and biological factors contribute to: 1) how lake food webs are assembled; and, 2) how these food webs may change in the future. First, I used long-term observations and lab experiments to determine how fish food, including zooplankton and snails, may respond to a warming climate. I then used field measurements of arctic char (Salvelinus alpinus) body characteristics, genetic samples, and fish diets to investigate if, and potentially why, populations of arctic char across a series of lakes achieve different maximum body sizes. Finally, as a method of monitoring population-level changes of fish abundance, I collected samples of arctic char DNA in lake water to test if estimated arctic char population abundances within a given lake correspond to the amount of DNA collected. Fish will require more food to eat as their metabolism increases with warming lake temperatures. Based on a thirty-year period of record, I determined zooplankton abundance increases in warmer years, indicating there is likely to be enough food for fishes in the future. Accordingly, zooplankton and snail abundance and development was also faster in warmer treatments of my lab experiments. My field observations indicated these are important prey items for arctic char. Small arctic char eat more zooplankton and large arctic char eat more snails, and these observations were consistent whether or not other predators are found in the particular lake. Similarly, my analyses did not indicate morphological or genetic differences between small and large arctic char within the same lake, suggesting arctic char size structure is determine by biological characteristics, including primary productivity and arctic char density. Indeed, estimates of arctic char population abundances across a series of lakes followed a gradient of arctic char densities, and my DNA sampling corresponded with this gradient. As there are thousands of lakes across the Arctic, my research demonstrates lake food webs, and the fishes within them, are likely to adapt to a warming climate. However, biological, chemical, and physical properties of these lakes can vary widely such that management and conservation plans may need to be developed at relatively small spatial scales across a large landscape.

Spider silk is one of the most versatile material.s in nature with great mechanical properties, exceeding some of the best man made materials. Native and synthetically produced silk has been used in a wide array of applications throughout the history of mankind including nets, bandages and cloths. It is recognized that spider silk can be a suitable replacement material for many existing materials such as ropes, body armor, parachutes and biodegradable bottles - all of which could show cost and environmental 4 benefits relative to other currently used man made materials. An added advantage to these types of applications is the potential for the products to have intrinsic antimicrobial activity. Studies have demonstrated a level of antimicrobial activity in native silk, a property that may have evolved in order to resist microbial decomposition, to protect developing eggs, and to resist decomposition or destruction by predators, parasites, or fluctuations in the environment. In this study, the novel aqueous glue coating peptides found on the silk fiber of the black widow spider, spider coating peptide 1 and 2, were investigated. Using circular dichroism, it was determined that SCP-1 and SCP-2 display predominantly alpha-helical secondary structures. In temperature gradient studies, SCP-1 is structurally stable at high temperatures while SCP-2 unfolded and lost its alpha-helical structure. The two peptides remained structurally stable both in an acidic and basic environment. This study was the first to characterize the secondary structure of the peptides found coating various silk fibers in Latrodectus hesperus, the black widow spider. The function of the SCPs is unknown but has-been hypothesized to potentially have antimicrobial properties. We investigated this role and found no significant antibacterial activity of the peptides against Escherichia coli and Bacillus subtitlis in growth studies. This study is the first to investigate the functional role of SCPs.

Spider silk from the female black widow spider, Latrodectus hesperus, is made of extraordinary biomolecules of nature. Efforts of the scientific community to commercially synthesize silks have become a collaborating, yet competitive race, to characterize the proteins that contribute to its intriguing biomechanics. Little has been reported on aqueous silk molecules in black widow spider silk, which are quite different from the large water insoluble core fibroins. In this study, a novel, aqueous aggregate gland derived silk factor (AgSF 1) was investigated using proteomics and immunological approaches. Western blot analyses of whole tissue lysates and solublized silk fibers revealed high levels of AgSFl in the aggregate gland, in the web scaffolding junctions, and in wrapping silk. MS/MS analyses of tryptic digest products from solubilized wrapping silk and aggregate gland whole cell lysate also confirmed the presence of AgSFl in these samples. Possible post-translational modifications were also analyzed by two-dimensional gel electrophoresis (2DE) and MS/MS analysis. AgSFl was localized in the web scaffolding junctions and our data supports a role as an adhesive silk protein that serves as a center for connecting scaffolding fibers that functions to reduce the tensile strength of scaffolding fibers, which facilitates capture of aerial insects.

Little is actually known about the transcriptional regulation of spider silk as most studies have focused on the material properties of silks. We isolated and mapped the TuSp1 core promoter from the black widow spider, Latrodectus hesperus. Using a genomic DNA walking strategy, we have isolated an upstream segment (581 bp) of genomic DNA containing the promoter as well as the first exon of the TuSp1 gene. This upstream regulatory element was able to initiate transcription in insect cells when placed upstream the promoterless firefly luciferase reporter gene. Initiation of transcription was orientation dependent, as insertion of this upstream regulatory module in the reverse orientation led to inefficient transcriptional initiation. Only 170 bp of upstream sequence was required for strong transcriptional initiation, showing that core promoter resides within the first 170 bp of upstream 5' -flanking DNA. We also demonstrate the bHLH factor SGSF1 can repress gene transcription of the TuSp1 core promoter, implying SGSF I might participate in the transcriptional regulation of the TuSp1 gene in vivo.

Belonging to the diverse order Araneae, the black widow spider Latrodectus 4 hesperus produces high-performance silks with a broad range ofbiological functions and mechanical properties. The cob weaver spider spins different fibers by using seven specialized glands located in its abdomen. Egg case silk originates from the tubuliforrn gland and to date, no proteins that participate in the assembly process of egg case silk proteins have been identified. The goal of this project was the expression, purification, and characterization of such protein products. De novo sequencing of peptides from in-solution tryptic digestion of black widow spider dragline silk, the most studied type of silk, identified a novel cysteine-rich nonfibroin- like peptide that we named cysteine-rich component or CRC- 1. Further analysis of a large pool of nucleic acid sequences deposited in our custom eDNA database revealed 4 additional sequences with similarities to each other at the amino acid level called CRC-2, CRC-3, CRC-4, and CRC-5, suggesting a new family of proteins. Specifically, Q-PCR analysis revealed that the CRC-5 mRNAs were predominantly expressed in the tubuliform and aggregate glands. Since the aggregate gland manufactures a more complex aqueous solution compared to the tubuliforrn gland, we focused these studies on the tubuliform gland and resultant egg case fibers. Westem blot analysis using a cross-reactive polyclonal anti-CRC-1 antiserum conoborated the presence of CRC-5 in the tubulifmm gland and egg case silk, supporting the colocalization ofTuSpl, a tubuliform gland-specific protein, and CRC-5. Thus, we have demonstrated that these two proteins are present within tubuliform silks. In vitro studies suggested that recombinant CRC-5 displayed enzymatic activity similar to a sulfhydryl oxidase. Collectively, our findings provide new insights into novel proteins that have a potential role in the silk assembly and extrusion pathway of egg case silk fibers.

Spider silk is a natural high-performance biopolymer with superior mechanical propetiies. Although these fibers out perfmm several man-made and natural biomaterials, there are cha llenges to be circumvented before commercialization. One of the silkproducing glands warranting further study is the pyrifonn gland, which produces gluelike threads functioning to cement dragline silk to substrates. We focused on the molecular properties of PySp 1, the major component of pyrifonn silk from Latrodectus hersperus, and its putative Oiiholog, PySp2, from Nephi/a clavipes. To date, there are no reports describing the secondary structure of PySp internal block repeats. Moreover, because the PySp C-terminus amino acid residues are distinct from MaSp C-terminus and the morphology of these glands is different, we hypothesized that PySp C-terminal domains form distinct secondary structures. The MaSp C-terminus has been shown to regulate the silk assembly process and whether the PySp C-terminus performs a similar function is unknown. In order to test this supposition, we used the following experimental approaches: I) we developed a series of PySp prokaryotic expression constructs carrying various block repeat modules representative of the internal iterations found within the protein chain; 2) we constructed prokaryotic expression vectors coding for the PySp C-terminal domains; 3) we expressed and purified the PySp C-terminal domains from bacteria; 4) we performed structural analyses of the purified PySp C-terminal domains using cd spectroscopy and atomic force microscopy. After expression and purification of the PySp C-tennini proteins, our studies support that this domain displays a predominantly ~-sheet structure, distinctive from the NMR-determined ahelical nature of MaSp C-tennini. The difference in secondary structure implies the MA and pyriform glands use different biochemical mechanisms during fiber extrusion to control protein folding and assembly. By investigating protein folding and fiber formation for different spider silk types, its characteristics can be customized for spinning different materials for industrial applications.