Influence of Ligand Complexation on Nickel Toxicity, Speciation and Bioavailability in Marine Waters

Currently there are no site-specific bioavailability-based prediction models for assessing the impacts of nickel (Ni) in marine environments although there are indications that these may be warranted. The aim of this research was to characterize the complexation of Ni in relation to toxicity and speciation. Various complexing ligands were used, and it was predicted that the binding affinity (logKf) of ligands would be inversely correlated to toxicity based on dissolved Ni concentrations ([NiD]) but that on a free ion concentration ([Ni2+]) basis, toxicity would not vary. A two-phased approach was used; the first was a proof of principle where synthetic ligands with known logKf values [EDTA, NTA, tryptophan (TRP), glutamic acid (GA), histidine (HD) and citric acid (CA)] were tested and the second, natural waters were characterized for binding capacity and Ni toxicity. Chronic Ni toxicity assays were performed using two marine species sensitive to Ni, the purple sea urchin (Strongylocentrotus purpuratus; where EC50 and EC20 values were determined) and the mysid (Americamysis bahia; LC50 and LC20). Embryological development and mortality (respectively) were used as the toxicity endpoints. Ni was measured by graphite furnace atomic absorption spectroscopy (GFAAS). The [NiD] E/LC50 values in unmodified artificial seawater (ASW) were 3.6 μM (95% CI 3.0-4.5 μM) for S. purpuratus and 2.6 μM (2.3-2.8 μM) for A. bahia. Tests with synthetic ligands provided significant protection based on [NiD], particularly for those with strong complexation such as EDTA and NTA. However, when considered on the basis of [Ni2+], E/LC50 values were either similar or less than those in ASW. In natural seawater the E/LC50 values ranged from 2.0 to 7.0 mM based on [NiD] and variability was reduced when expressed on a [Ni2+] basis. There were no significant differences in Ni toxicity between natural waters and ASW. Overall, this study supports the theory that free Ni concentration is the best predictor of toxicity and confirms the applicability of a marine BLM for Ni. It also provides insight into the understanding of relationships between aquatic geochemistry, Ni speciation, complexation and toxicity from a biological perspective

Influence of Ligand Complexation on Nickel Toxicity, Speciation and Bioavailability in Marine Waters

Currently there are no site-specific bioavailability-based prediction models for assessing the impacts of nickel (Ni) in marine environments although there are indications that these may be warranted. The aim of this research was to characterize the complexation of Ni in relation to toxicity and speciation. Various complexing ligands were used, and it was predicted that the binding affinity (logKf) of ligands would be inversely correlated to toxicity based on dissolved Ni concentrations ([NiD]) but that on a free ion concentration ([Ni2+]) basis, toxicity would not vary. A two-phased approach was used; the first was a proof of principle where synthetic ligands with known logKf values [EDTA, NTA, tryptophan (TRP), glutamic acid (GA), histidine (HD) and citric acid (CA)] were tested and the second, natural waters were characterized for binding capacity and Ni toxicity. Chronic Ni toxicity assays were performed using two marine species sensitive to Ni, the purple sea urchin (Strongylocentrotus purpuratus; where EC50 and EC20 values were determined) and the mysid (Americamysis bahia; LC50 and LC20). Embryological development and mortality (respectively) were used as the toxicity endpoints. Ni was measured by graphite furnace atomic absorption spectroscopy (GFAAS). The [NiD] E/LC50 values in unmodified artificial seawater (ASW) were 3.6 μM (95% CI 3.0-4.5 μM) for S. purpuratus and 2.6 μM (2.3-2.8 μM) for A. bahia. Tests with synthetic ligands provided significant protection based on [NiD], particularly for those with strong complexation such as EDTA and NTA. However, when considered on the basis of [Ni2+], E/LC50 values were either similar or less than those in ASW. In natural seawater the E/LC50 values ranged from 2.0 to 7.0 mM based on [NiD] and variability was reduced when expressed on a [Ni2+] basis. There were no significant differences in Ni toxicity between natural waters and ASW. Overall, this study supports the theory that free Ni concentration is the best predictor of toxicity and confirms the applicability of a marine BLM for Ni. It also provides insight into the understanding of relationships between aquatic geochemistry, Ni speciation, complexation and toxicity from a biological perspective

Ecology of rays on tropical coral reefs

Rays are a diverse group of elasmobranchs in both their morphology and ecology. They are among the most threatened elasmobranchs according to the International Union for the Conservation of Nature (IUCN), however, little is known about their life history, behaviour or population status. To be able to improve management of declining ray populations, their distribution and ecology must be better understood. Coastal species are most at risk from anthropogenic effects, however, the extent of impacts on rays have not been widely documented. Rays serve important ecosystems functions including stabilizing food webs and acting as ecosystem engineers through bioturbation. As mesopredators, rays prey on a variety of primary consumers, while also being prey for apex predators. Because of the many linkages in food webs among apex predators to producers, food web stabilization by mesopredators prevents trophic cascades. Some rays also serve their ecosystems through bioturbation, the biological reworking of sediments. During the act of feeding, some rays create feeding pits which oxygenate sediments, provides habitat for small teleosts and crustaceans, and facilitates meiofaunal movement. While rays serve important roles in their ecosystems, there is little species-specific information available. The lack of research is partially due to the cryptic nature of rays, making them difficult to study, and partially due to the charismatic nature of their relatives the sharks, which have received much more attention. Therefore, more research is needed to address the deficits in our knowledge of ray ecology and distribution. Baited remote underwater video systems (BRUVS) are increasingly used to study fish communities, biomass, and animal behaviour. BRUVS entail deploying baited video cameras in the absence of human presence in order to survey fish and invertebrate populations. This methodology reduces human influence on the study species and encourages more natural behaviours than with human presence. Due to the popularity of BRUVS approaches, there are many analysis methods. MaxN, which refers to the maximum number of individuals observed of a species in a single frame of a video, is the most commonly used metric of relative abundance when analysing BRUVS data. Chapter 3 presents a novel metric for BRUVS analysis that involves identifying and counting distinct individuals (MaxIND) to quantify the accuracy of MaxN. Individual oriental bluespotted maskray (Neotrygon orientalis) and the bluespotted fantail ray (Taeniura lymma) were identified on BRUVS by spot patterns, tail characteristics, and sex at three sites in Malaysian Borneo. We demonstrated that MaxIND gave abundances that were 2.4 and 1.1 times higher than MaxN for N. orientalis and T. lymma, respectively. These differences between methods were consistent for each species between sites regardless of the presence of marine reserves. However, differences in abundance estimates from MaxN to MaxIND were apparent between species, indicating that correction factors need to be developed on a species basis to better estimate true abundance. While identifying individuals is time consuming, it provides improved accuracy and information about populations. We therefore recommend the use of MaxIND when rare and threatened species are present, in high density populations, and for behavioural analyses, where distinguishing features are present. Ecological sampling must yield consistent results in order to reliably quantify predator populations. BRUVS are increasingly being used to evaluate and monitor predator communities in marine ecosystems. Many BRUVS studies compare multiple coral reef sites sampled at a single point in time. As coral reef monitoring using BRUVS grows in its capacity to provide data relevant to sustainable management, marine protected area efficacy, and overall reef health, understanding repeatability of sampling results is vital. Chapter 4 examined the repeatability of BRUVS results for the elasmobranch community both within and between seasons (dry and wet) and years, and explored environmental factors affecting abundances at two sites in Bau Bau, Indonesia. A total of 1139 elasmobranchs (69% rays, 31% sharks) were observed on 956 BRUVS across six sampling events. Consistent results were found both in species composition and abundances within a season and across multiple years using the same sampling protocol (number and location of BRUVS). However, abundances of all sharks and rays were significantly higher in the wet season in both years. The elasmobranch community was significantly different between the two sites sampled in a consistent manner. The results demonstrated that while BRUVS are a consistent, reliable and repeatable method for surveying elasmobranchs, care must be taken in timing of sampling various regions to ensure accuracy when comparing multiple locations as season was an important factor in the results. Coral reef ecosystems are highly dynamic environments with complex trophic interactions and environmental drivers. Rays are important members of these systems, however, in areas like Southeast Asia they are often heavily fished. Their conservation is difficult, as many countries in which they are fished do not have the capacity for effective fisheries management. For chapter 5, BRUVS were deployed at 70 reefs in 11 countries across the Coral Triangle and Australasian regions to determine ray abundances and assemblage. In 3426 BRUVS deployments, 1069 ray individuals were observed. The three most abundance species / genera were maskrays (Neotrygon spp.), fantail rays (Taeniura spp.), and eagle rays (Generas: Aetobatus, Aetomylaeus, Myliobatus). Ray assemblage was relatively consistant across the study area, however, ray abundances varied greatly with only a single individual in Vietnam to a very high abundance of rays in Indonesia. The differences in abundances are likely a reflection of fishing pressure and fisheries management. In countries with low fishing pressure, communities were species rich in both rays and sharks. Countries with moderate fishing pressure began to lose species richness, especially of sharks, although abundances of rays remained similar. With high fishing pressure, only small productive species of rays were present, and these were abundant due to the lack of top predators (i.e. sharks). Finally, in countries with extremely high fishing pressure, even productive species were absent. In order to conserve rays and their ecosystem services, fisheries management must be addressed. In some cases this requires fisheries management implementation and in some cases may involve increased management efficacy. Additionally, habitat quality and characteristics also affect the ray community at finer scales than fishing pressure. Benthic relief was most important to all rays with some species preferring low relief areas and some preferring high relief (coral dominated) areas. Thus, in addition to fisheries management, habitat quality and conservation is also important for ray species. Sharks are decreasing in abundance in many coral reef habitats, but the ecosystem effects of this loss are poorly understood. Rays are a prevalent mesopredator in tropical coral reef ecosystems experiencing low fishing pressure that are preyed upon by top predators like sharks. Across Southeast Asia and the Western Pacific there are varying abundances of coral reef predators that consume rays. Studies have suggested reduced predator abundances leads to increases in mesopredator abundance (mesopredator release) and potentially trophic cascades. In this study, we examined the relationship between top predator abundance (sharks) and the abundance and behaviour of two genera of small benthic rays using BRUVS at 19 sites across six countries. Where predators were more abundant, the bluespotted maskray complex (Neotrygon spp.) and two species of fantail rays (Taeniura lymma and T. lessoni) were sighted less often, possibly because of lower abundances. However, small ray behaviour was significantly affected by predator abundance. Individuals of focal ray species visited BRUVS significantly fewer times at sites with higher predator abundances. Where predators were less abundant, rays spent significantly more time in the video frame, visited BRUVS more often, and were more likely to feed from bait bags. In addition to predator abundance, small ray presence was significantly influenced by relief and depth. Neotrygon spp. were more abundant on deeper, lower relief habitats, while Taeniura spp. were more prevalent in reef-associated shallow, high relief habitats. Overall, this chapter found that predator abundance had a significant effect on small benthic ray abundance and behaviour in the presence of BRUVS. The results demonstrate that changes in behaviour associated with the loss of predators may make the interpretation of phenomena like mesopredator release more challenging unless behavioural effects are taken into account. This thesis demonstrates the many uses of BRUVS as a tool for surveying ray abundances, behaviours, and assemblage. A variety of analysis techniques were used for BRUVS data, with results proving the effectiveness of this survey method. Using the newly described metric MaxIND, more accurate abundance estimates and behavioural analyses are able to be performed in a natural setting. As there is limited data about rays on coral reefs globally, this thesis provides basic information about ray assemblages and abundances across the Coral Triangle and Australasian regions. Countries within these regions have extremely variable fishing pressure and management capacity leading towards changes in ray populations. In order to conserve ray species, improved fisheries management and habitat preservation are needed

Surviving a Batterer: An Ideal Policy Approach to Combating Intimate Partner Violence (IPV)

Gender violence has plagued developed and developing societies for centuries, embedded in culture, structures, and ways of life. Women have been seen as pieces of property with no autonomy or individualism, just as extensions of their husbands. My research centers around finding an ideal policy solution to diminish rates of intimate partner violence (IPV) in the case of California. Interviews and data collection with legislators concerning education, rehabilitation or batterer intervention programs (BIP), and care providers in emergency shelters regarding victims’ services provided insight on a three-pronged approach targeted at curbing rates of IPV in California. My findings yielded that although these variables are present in California, there must be an allocation of more resources and funding in order for ideal policy to be effective in California and across the nation

Pharmaceutical pollution in marine waters and benthic flora of the southern Australian coastline

Environmental context Most human pharmaceutical waste is discharged to the environment. While the presence of pharmaceuticals in freshwater systems is well documented globally, little is known of the impact on marine ecosystems. We measured pharmaceuticals in a marine environment in south-eastern Australia and found pharmaceutical concentrations around 24 000 times higher in benthic flora than in the marine surface waters. We discuss the potential use of seaweeds as biological indicators of pharmaceutical pollution. Rationale Pharmaceuticals are emerging pollutants of concern with a range of adverse consequences for organisms and ecosystems. Their presence in freshwater and estuarine systems has been well documented, but less is known about their prevalence in open ocean, or their uptake by benthic flora. This preliminary survey of the southern Australian coastline sought to measure the concentrations of key pharmaceuticals in both surface waters and benthic flora. Methodology This study used LC-MS/MS to measure the concentration carbamazepine, tramadol and venlafaxine in (1) samples from wastewater treatment plants, (2) ocean surface waters and (3) several species of benthic flora. Surface waters and benthic flora were sampled at two sites near waste water treatment plant (WWTP) discharges, and one site away from any discharge. Results All three pharmaceuticals were detected in surface water samples with their risk assessed (via risk quotient) as medium risk (carbamazepine) or low risk (venlafaxine, tramadol). All three pharmaceuticals were also detected in benthic flora, particularly in brown macroalgae Tramadol was measured at a maximum of 34.7 ng 

A microRNA signature in circulating exosomes is superior to exosomal glypican-1 levels for diagnosing pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is a deadly malignancy that often presents clinically at an advanced stage and that may be confused with chronic pancreatitis (CP). Conversely, CP may be misdiagnosed as PDAC leading to unwarranted pancreas resection. Therefore, early PDAC diagnosis and clear differentiation between PDAC and CP are crucial for improved care. Exosomes are circulating microvesicles whose components can serve as cancer biomarkers. We compared exosomal glypican-1 (GPC1) and microRNA levels in normal control subjects and in patients with PDAC and CP. We report that exosomal GPC1 is not diagnostic for PDAC, whereas high exosomal levels of microRNA-10b, (miR-10b), miR-21, miR-30c, and miR-181a and low miR-let7a readily differentiate PDAC from normal control and CP samples. By contrast with GPC1, elevated exosomal miR levels decreased to normal values within 24 h following PDAC resection. All 29 PDAC cases exhibited significantly elevated exosomal miR-10b and miR-30c levels, whereas 8 cases had normal or slightly increased CA 19-9 levels. Thus, our exosomal miR signature is superior to exosomal GPC1 or plasma CA 19-9 levels in establishing a diagnosis of PDAC and differentiating between PDAC and CP

Type II Kinase Inhibitors Show an Unexpected Inhibition Mode against Parkinson’s Disease-Linked LRRK2 Mutant G2019S

A number of well-known type II inhibitors (ATP-noncompetitive) that bind kinases in their DFG-out conformation were tested against wild-type LRRK2 and the most common Parkinson’s disease-linked mutation, G2019S. We found that traditional type II inhibitors exhibit surprising variability in their inhibition mechanism between the wild type (WT) and the G2019S mutant of LRRK2. The type II kinase inhibitors were found to work in an ATP-competitive fashion against the G2019S mutant, whereas they appear to follow the expected noncompetitive mechanism against WT. Because the G2019S mutation lies in the DXG motif (DYG in LRRK2 but DFG in most other kinases) of the activation loop, we explored the structural consequence of the mutation on loop dynamics using an enhanced sampling method called metadynamics. The simulations suggest that the G2019S mutation stabilizes the DYG-in state of LRRK2 through a series of hydrogen bonds, leading to an increase in the conformational barrier between the active and inactive forms of the enzyme and a relative stabilization of the active form. The conformational bias toward the active form of LRRK2 mutants has two primary consequences. (1) The mutant enzyme becomes hyperactive, a known contributor to the Parkinsonian phenotype, as a consequence of being “locked” into the activated state, and (2) the mutation creates an unusual allosteric pocket that can bind type II inhibitors but in an ATP-competitive fashion. Our results suggest that developing type II inhibitors, which are generally considered superior to type I inhibitors because of desirable selectivity profiles, might be especially challenging for the G2019S LRRK2 mutant