Battling pollution in coastal areas / Lutter contre la pollution dans les zones côtières

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    IDRC Research Chair in Environmental Health and Management : final technical report
    (2014-12) Bianchi, Adalto; Wood, Christopher
    The project developed alternative management strategies to guide industrial regulation, human settlement and urban policy in coastal areas, using ecotoxicology modeling (Biotic Ligand Model - BLM) for metal release regulatory purposes, and biomonitoring of water quality. Environmental pollution originating from human activities contaminates estuarine and other coastal areas due to net drainage from the watershed. The report provides a review of project fieldwork in Brazil and Canada, methodology, participants, outcomes, outputs, recommendations and a bibliography of sources. The tools developed in the project were established as reliable and valuable for evaluating water quality and ecosystem health in coastal areas.
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    Biochemical composition and performance of Atlantic cod (Gadus morhua L.) eggs and larvae obtained from farmed and wild broodstocks
    (Elsevier, 2012) Ceccon Lanes, Carlos Frederico; Tilahun Bizuayehu, Teshome; Bolla, Sylvie; Martins, Camila; de Oliveira Fernandes, Jorge Manuel
    Farming of Atlantic cod, Gadus morhua L., has been facing several challenges lately. Biological issues such as differences in egg quality and lack of high quality larvae are considered as prominent limiting factors. The aim of this study was to compare fertilization and hatching rates, incidence of malformed larvae at hatching, larval viability (high-salinity stress and starvation tolerance tests) and the biochemical profile (proteins, amino acids, lipids, fatty acids and minerals) between eggs produced from wild and farmed broodstocks. Twenty-five batches of eggs from farmed broodstock (FB) and 27 egg batches from wild broodstock (WB) were collected during the reproductive season. Fertilization and hatching rates were significantly higher in WB group (91±3% and 86±3%, respectively) than in FB (75±3% and 66±3%, respectively). In highsalinity stress test, WB larvae survived for a significantly longer time than FB larvae. No differences between egg batches from the two broodstocks were observed in starvation tolerance tests, incidence of malformed larvae at hatching, as well as in the total content of proteins, lipids, fatty acids, zinc, and calcium concentrations. Regarding fatty acids, FB eggs had significantly higher levels of C16:0, C18:0, C18:2n−6, C18:3n−3, C18:3n−6, C20:4n−6, C20n:5n−3, C22:5n−3 and C22:5n−6, whereas WB eggs had significantly higher levels of C14:0, C16:1n−7, C18:4n−3, C20:4n−3, C22:6n−3 and C24:1n−9. As for the amino acid profile, WB eggs had significantly higher levels of aspartic acid, serine, glycine, threonine, taurine and leucine, while FB eggs had significantly higher levels of proline, tryptophan, phenylalanine and arginine. The concentrations of minerals such as copper and phosphorus were significantly higher in WB eggs, whereas in FB eggs iron concentration was significantly higher. Overall, the biological parameters indicated that egg and larvae originating from WB were of superior quality than those from FB. The information on the biochemical profile may be useful in improving the quality of broodstock feeds for Atlantic cod.
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    Mortality, bioaccumulation and physiological responses in juvenile freshwater mussels (Lampsilis siliquoidea) chronically exposed to copper
    (Elsevier, 2013) Jorge, Marianna B.; Loro, Vania L.; Bianchini, Adalto; Wood, Chris M.; Gillis, Patricia L.
    As little is known about the toxic mode of action and sub-lethal effects of copper exposure in freshwater mussels, the study examined physiological effects of long-term copper exposure (survival, growth, copper bioaccumulation, whole-body ion content, oxygen consumption, filtration rate, ATPase activities, and biomarkers of oxidative stress) in juvenile (6 month old) mussels (Lampsilis siliquoidea). Results indicate that ionoregulatory disruption in freshwater mussels chronically exposed to copper is the main mechanism of toxicity and that redox parameters do not appear to be useful as indicators of sub-lethal copper toxicity.
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    Estimation of zooplankton secondary production in estuarine waters : Comparison between the enzymatic (chitobiase) method and mathematical models using crustaceans
    (Elsevier, 2012) Ramos Avila, Tatiana; Abel de Souza Machado, Anderson; Bianchini, Adalto
    Sampling was seasonally performed in the Patos Lagoon estuary (Rio Grande, RS, Southern Brazil) to estimate zooplankton biomass and production comparing values obtained using an enzymatic (chitobiase) method and the traditional mathematical models based on growth. Comparison of data obtained from zooplankton samples collected with 90 and 200-␣m mesh nets showed that net selectivity in␣uences the estimation of zooplankton biomass and production. Furthermore, it showed differential results for dominance of taxa and proportions of developmental stages in samples. Differences among samples collected at the different sites in the same season were observed using either the mathematical models or the enzymatic method. The two different approaches were also able to detect the seasonal variation in production. In a broad view, data obtained using the chitobiase method showed a similar pattern of zooplankton production compared to those obtained with the traditional mathematical models based on growth. However, values estimated using the enzymatic method were systematically higher than those obtained with the mathematical models. Maxi- mum total production values were estimated as 12.5, 9.2 and 7.9 mg C m␣ 3 day␣ 1 for the “chitobiase method”, “Huntley model”, and “Hirst model”, respectively. Considering all sampling sites and seasons, the magnitude of this difference corresponded to 1.95 and 2.49 mg C m␣ 3 day␣ 1 for the “Huntley model” and the “Hirst model”, respectively. These ␣ndings indicate the reliability of the enzymatic method in estimating crustacean production also in estuarine environments of changing salinity, as previously demonstrated for marine waters. In addition, the use of this method is more practical and comparatively less time-consuming and cheaper than the use of the mathematical models based on growth.
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    Copper effects on key metabolic enzymes and mitochondrial membrane potential in gills of the estuarine crab Neohelice granulata at different salinities
    (Elsevier, 2012) Machado Lauer, Mariana; Bento de Oliveira, Camila; Inocencio Yano, Natalia Lie; Bianchini, Adalto
    The estuarine crab Neohelice granulata was exposed (96 h) to a sublethal copper concentration under two different physiological conditions (hyperosmoregulating crabs: 2 ppt salinity, 1 mg Cu/L; isosmotic crabs: 30 ppt salinity, 5 mg Cu/L). After exposure, gills (anterior and posterior) were dissected and activities of enzymes involved in glycolysis (hexokinase, phosphofructokinase, pyruvate kinase, lactate dehydrogenase), Krebs cycle (citrate synthase), and mitochondrial electron transport chain (cytochrome c oxidase) were analyzed. Membrane potential of mitochondria isolated from anterior and posterior gill cells was also evaluated. In anterior gills of crabs acclimated to 2 ppt salinity, copper exposure inhibited hexokinase, phosphofructokinase, pyruvate kinase, and citrate synthase activity, increased lactate dehydrogenase activity, and reduced the mitochondrial membrane potential. In posterior gills, copper inhibited hexokinase and pyruvate kinase activity, and increased citrate synthase activity. In anterior gills of crabs acclimated to 30 ppt salinity, copper exposure inhibited phosphofructokinase and citrate synthase activity, and increased hexokinase activity. In posterior gills, copper inhibited phosphofructokinase and pyruvate kinase activity, and increased hexokinase and lactate dehydrogenase activity. Copper did not affect cytochrome c oxidase activity in either anterior or posterior gills of crabs acclimated to 2 and 30 ppt salinity. These findings indicate that exposure to a sublethal copper concentration affects the activity of enzymes involved in glycolysis and Krebs cycle, especially in anterior (respiratory) gills of hyperosmoregulating crabs. Changes observed indicate a switch from aerobic to anaerobic metabolism, characterizing a situation of functional hypoxia. In this case, reduced mitochondrial membrane potential would suggest a decrease in ATP production. Although gills of isosmotic crabs were also affected by copper exposure, changes observed suggest no impact in the overall tissue ATP production. Also, findings suggest that copper exposure would stimulate the pentose phosphate pathway to support the antioxidant system requirements. Although N. granulata is very tolerant to copper, acute exposure to this metal can disrupt the energy balance by affecting biochemical systems involved in carbohydrate metabolism.
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    Assessment of water quality in coastal waters of Fernando de Noronha, Brazil : biomarker analyses in Amphistegina lessonii
    (GeoScienceWorld, 2012) de Freitas Prazeres, Martina; Eslava Martins, Samantha; Bianchini, Adalto
    Symbiont-bearing foraminiferal assemblages from reef environments have proven useful as ecological indicators for water quality. Studies with other organisms have utilized cellular biomarkers to elucidate mechanisms of stress response. Our goal was to combine these approaches, focusing on populations of the symbiont-bearing foraminifer Amphistegina lessonii from Fernando de Noronha Archipelago (northeastern Brazil). Reef-rubble samples containing A. lessonii were collected at five sampling sites on the leeward side of the main island. Three sites were located inside and two outside the Fernando de Noronha National Marine Park (FNNMP) area. Foraminifers were evaluated regarding their density, degree of bleaching, antioxidant capacity against peroxyl radicals (ACAP), lipid peroxidation (LPO), protein carbonylation (PC), and concentrations of metallothioneinlike proteins (MTs). Densities of A. lessonii were lower at sites located outside the FNNMP, compared with those inside the FNNMP, where the maximum average was 159±29 individuals/100 cm2. In individuals from sites located outside FNNMP, bleaching frequency reached 25%, though ,2%; at FNNMP sites (,1.8%;). ACAP was higher (two-fold) in individuals collected at the FNNMP sites, which showed lower LPO and PC levels. MTs concentration did not differ significantly among sites. Zinc, copper, and lead concentrations were very high at stations located outside FNNMP. Dissolved organic carbon concentration was high at all sampling sites. Redundancy analysis revealed a strong correlation between biomarkers and water quality among sampling sites. These results show that the biomarker approach would be effective in identifying impacted areas and underlying causes, providing a reliable diagnostic for environment health.
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    Growth hormone transgenesis affects osmoregulation and energy metabolism in zebrafish (Danio rerio)
    (Springer, 2013) Volcan Almeida, Daniela; de Martinez Gaspar Martins, Camila; de Azevedo Figueiredo, Márcio; Ceccon Lanes, Carlos Frederico; Bianchini, Adalto; Marins, Luis Fernando
    Growth hormone (GH) transgenic fish are at a critical step for possible approval for commercialization. Since this hormone is related to salinity tolerance in fish, our main goal was to verify whether the osmoregulatory capacity of the stenohaline zebrafish (Danio rerio) would be modified by GH-transgenesis. For this, we transferred GH-transgenic zebrafish (T) from freshwater to 11 ppt salinity and analyzed survival as well as relative changes in gene expression. Results show an increased mortality in T versus non-transgenic (NT) fish, suggesting an impaired mechanism of osmotic acclimation in T. The salinity effect on expression of genes related to osmoregulation, the somatotropic axis and energy metabolism was evaluated in gills and liver of T and NT. Genes coding for Na(+), K(+)-ATPase, H(+)-ATPase, plasma carbonic anhydrase and cytosolic carbonic anhydrase were up-regulated in gills of transgenics in freshwater. The growth hormone receptor gene was down-regulated in gills and liver of both NT and T exposed to 11 ppt salinity, while insulin-like growth factor-1 was down-regulated in liver of NT and in gills of T exposed to 11 ppt salinity. In transgenics, all osmoregulation-related genes and the citrate synthase gene were down-regulated in gills of fish exposed to 11 ppt salinity, while lactate dehydrogenase expression was up-regulated in liver. Na(+), K(+)-ATPase activity was higher in gills of T exposed to 11 ppt salinity as well as the whole body content of Na(+). Increased ATP content was observed in gills of both NT and T exposed to 11 ppt salinity, being statistically higher in T than NT. Taking altogether, these findings support the hypothesis that GH-transgenesis increases Na(+) import capacity and energetic demand, promoting an unfavorable osmotic and energetic physiological status and making this transgenic fish intolerant of hyperosmotic environments.
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    Cobia Rachycentron canadum L. reared in low-salinity water : does dietary sodium chloride affect growth and osmoregulation?
    (Blackwell Publishing, 2012) Santos, Renato A.; Bianchini, Adalto; Jorge, Marianna B.; Romano, Luis A.; Sampaio, Luís A.; Tesser, Marcelo B.
    The effects of NaCl supplementation (0.0%, 2.5%, 5.0%, 7.5% and 10.0% dry weight of a basal diet) on growth, gill histological alterations and osmoregulation of juvenile cobia reared in low-salinity water (5 g L−1) were assessed. At the end of the experiment, gills were sampled for Na+, K+-ATPase activity determination and histological evaluation. In all treatments, no mortality was observed. Results showed that dietary NaCl supplementation did not alter growth. At the highest supplementations (7.5% and 10.0%), juvenile cobia showed higher feed intake and feed conversion ratio. Na+, K+-ATPase activity was higher in fish fed the diet without salt supplementation than in those fed with NaCl-supplemented diets. The number of chloride cells significantly increased with increasing dietary salt level, being 2.5-fold higher in fish fed with 10.0% NaCl supplementation (41 cells mm−2) than in those from the non-supplemented fed group (16 cells mm−2). These findings indicate that dietary salt supplementation stimulated chloride cell proliferation paralleled with a reduction in the gill Na+, K+-ATPase activity, suggesting a possible decrease in energy consumption associated with osmoregulation. However, the suggested energy sparing did not have a significant impact on juvenile cobia growth.
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    Acute waterborne copper toxicity to the euryhaline copepod Acartia tonsa at different salinities : influence of natural freshwater and marine dissolved organic matter
    (SETAC Press, 2013) Carvalho Rodrigues Monteiro, Sandra; Lopes Leães Pinho, Grasiela; Hoffmann, Karine; Barcarolli, Indianara Fernanda; Bianchini, Adalto
    The influence of natural dissolved organic matter (DOM) on acute waterborne Cu toxicity was evaluated in the euryhaline copepod Acartia tonsa at 3 different water salinities. Three sources of freshwater DOM (extracted by reverse osmosis) and 2 sources of marine DOM (extracted using a solid-phase technique) were used. Artificial salt water was used to prepare the experimental media. Different combinations of Cu concentrations and DOM sources and concentrations were tested at salinities of 5, 15, and 30 ppt. Toxicity data (48-h median lethal concentration [LC50] values) were calculated based on dissolved Cu concentrations. In a broad view, data showed that increasing salinity was protective against the acute waterborne Cu toxicity. In general, Cu toxicity was also lower in the presence than in the absence of DOM. Toxicity (48-h LC50) values from all treatments at the same salinity showed a positive linear relationship with the dissolved organic carbon (DOC). Thus, the protective effect of DOM against the acute Cu toxicity seems to be dependent mainly on the DOM concentration. However, it seems also to be dependent to some extent on the source of DOM used. In summary, findings reported in the present study clearly indicate that both salinity and DOM (source and concentration) should be taken into account in the development of an estuarine version of the biotic ligand model.
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    Cadmium and calcium transport along the gastro-intestinal tract of rainbow trout : more than “gut feelings” on mechanisms of uptake
    (Biology Department, McMaster University, Hamilton, ON, CA, 2011-10) Klinck, Joel S.; Biology Department, McMaster University
    Teleosts take up metals by two major pathways: gills and/or gut. Past research is heavily focused on branchial uptake despite evidence that the gastro-intestinal tract (GIT) is the dominant route in some natural environments. To address this information gap, my thesis characterizes uptake mechanisms of Cd and Ca along the GIT of Oncorhynchus mykiss. Toxic effects of Cd, protective effects of Ca against Cd uptake, and Cd distribution within fish after ingestion are also explored. Four-week dietary Cd exposure affected growth and Ca regulation, while causing toxicity at the subcellular level. Elevated Ca in diets protected against accumulation and altered subcellular handling of Cd. Pre-exposure to different diets changed unidirectional uptake and binding rates of Cd and Ca, although they remained highly correlated. Binding rates of Cd to mucus can predict absorption rates of Cd. A variety of in vitro and in vivo experiments on four distinct GIT segments (stomach, anterior-, mid-, and posterior- intestine) were undertaken to determine specific mechanisms of Cd and Ca uptake in freshwater trout. Cd transport was unaffected by solvent drag, but was stretch- and temperature- sensitive. Strong evidence for a common pathway for Ca and Cd transport was obtained. Cd also appeared to be taken up in part by zinc (ZIP-like) transporters and the divalent metal transporter DMT1. Ca uptake along the GIT appeared to be carrier-mediated, time- and concentration- dependent, but was not affected by solvent drag, or by Na concentration. Mucosal Cd did not inhibit Ca uptake. Ca, but not Cd, uptake rates were much lower in sea water-acclimated trout. Seawater and freshwater fish accumulated similar whole body Cd concentrations when fed contaminated diets for three weeks, but the majority of Cd in seawater trout remained in the posterior intestine tissue, while freshwater trout had higher internal burdens. This thesis advances the knowledge of metal uptake along the GIT of trout.
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    Waterborne copper exposure inhibits ammonia excretion and branchial carbonic anhydrase activity in euryhaline guppies acclimated to both fresh water and sea water
    (Elsevier, 2012) Zimmer, Alex; Indianara Fernanda Barcarolli; Chris M. Wood; Adalto Bianchini
    Inhibition of ammonia excretion (J(amm)) is a common response to Cu exposure in freshwater (FW) and seawater (SW) organisms. To determine the mechanism of this response, a euryhaline species of guppy (Poecilia vivipara) was exposed to 20 μg Cu/l in FW (0 ppt) and SW (25 ppt) for 96 h. In both salinities, Cu transiently inhibited ammonia excretion (J(amm)) followed by a full recovery by the end of the 96 h exposure. The activities of Na(+)/K(+)-ATPase, H(+)-ATPase, and carbonic anhydrase (CA) were examined in the gills at 12 and 96 h of Cu exposure. In both salinity acclimations, CA activity was significantly inhibited following 12h of Cu exposure in P. vivipara, marking the first in vivo evidence of Cu-induced inhibition of CA in fish. Moreover, the inhibition and recovery of this enzyme were correlated with the inhibition and recovery of J(amm) in both salinity acclimations. The blockade of CA potentially acts as a common mechanism of J(amm) inhibition in FW and SW. There were no significant effects on Na(+)/K(+)-ATPase or H(+)-ATPase activity at either time point or salinity. However, H(+)-ATPase activity was upregulated at 96 h relative to the 12h time point, potentially involving this enzyme in re-establishing J(amm).
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    Transepithelial potential in the Magadi tilapia, a fish living in extreme alkalinity
    (Springer, 2012) Wood, Chris M.; Bergman, Harold L.; Bianchini, Adalto; Laurent, Pierre; Maina, John
    We investigated the transepithelial potential (TEP) and its responses to changes in the external medium in Alcolapia grahami, a small cichlid fish living in Lake Magadi, Kenya. Magadi water is extremely alkaline (pH = 9.92) and otherwise unusual: titratable alkalinity (290 mequiv L(-1), i.e. HCO(3) (-) and CO(3) (2-)) rather than Cl(-) (112 mmol L(-1)) represents the major anion matching Na(+) = 356 mmol L(-1), with very low concentrations of Ca(2+) and Mg(2+) (<1 mmol L(-1)). Immediately after fish capture, TEP was +4 mV (inside positive), but stabilized at +7 mV at 10-30 h post-capture when experiments were performed in Magadi water. Transfer to 250% Magadi water increased the TEP to +9.5 mV, and transfer to fresh water and deionized water decreased the TEP to -13 and -28 mV, respectively, effects which were not due to changes in pH or osmolality. The very negative TEP in deionized water was attenuated in a linear fashion by log elevations in [Ca(2+)]. Extreme cold (1 vs. 28°C) reduced the positive TEP in Magadi water by 60%, suggesting blockade of an electrogenic component, but did not alter the negative TEP in dilute solution. When fish were transferred to 350 mmol L(-1) solutions of NaHCO(3), NaCl, NaNO(3), or choline Cl, only the 350 mmol L(-1) NaHCO(3) solution sustained the TEP unchanged at +7 mV; in all others, the TEP fell. Furthermore, after transfer to 50, 10, and 2% dilutions of 350 mmol L(-1) NaHCO(3), the TEPs remained identical to those in comparable dilutions of Magadi water, whereas this did not occur with comparable dilutions of 350 mmol L(-1) NaCl-i.e. the fish behaves electrically as if living in an NaHCO(3) solution equimolar to Magadi water. We conclude that the TEP is largely a Na(+) diffusion potential attenuated by some permeability to anions. In Magadi water, the net electrochemical forces driving Na(+) inwards (+9.9 mV) and Cl(-) outwards (+3.4 mV) are small relative to the strong gradient driving HCO(3) (-) inwards (-82.7 mV). Estimated permeability ratios are P (Cl)/P (Na) = 0.51-0.68 and [Formula: see text] = 0.10-0.33. The low permeability to HCO(3) (-) is unusual, and reflects a unique adaptation to life in extreme alkalinity. Cl(-) is distributed close to Nernst equilibrium in Magadi water, so there is no need for lower P (Cl). The higher P (Na) likely facilitates Na(+) efflux through the paracellular pathway. The positive electrogenic component is probably due to active HCO(3) (-) excretion.
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    Oxidative stress parameters and antioxidant response to sublethal waterborne zinc in a euryhaline teleost Fundulus heteroclitus : protective effects of salinity
    (Elsevier, 2012) Loro, Vania Lucia; Jorge, Mariana Basso; Rios da Silva, Kassio; Wood, Chris M.
    Zinc is an essential trace metal, but many aspects of its toxicity remain unclear. In this study, we investigated zinc effects on oxidative stress parameters and antioxidant profile in four tissues (gill, liver, kidney, and white muscle) of Fundulus heteroclitus. Possible interactive effects of salinity were also studied. Killifish were exposed to sublethal level (500 μgL(-1)) of waterborne zinc for 96 h in 0% (fresh water), 10% (3.5 ppt), 30% (10.5 ppt) and 100% sea water (35 ppt). Salinity per se had no effect on any parameter in the control groups. Zinc exposure clearly induced oxidative stress, and responses were qualitatively similar amongst different tissues. Salinity acted as a strong protective factor, with the highest levels of reactive oxygen species (ROS) and greatest damage (protein carbonyls, lipid peroxidation as indicated by thiobarbituric acid reactive substances (TBARS) in 0 ppt, the least in 100% sea water (35 ppt), and gradations in between in many of the observed responses. Increases in total oxidative scavenging capacity (TOSC) occurred at higher salinities, correlated with increases in the activities of superoxide dismutase (SOD) and glutathione-S-tranferase (GST), as well as in tissue glutathione (GSH) concentrations. However, TOSC was depleted in zinc-exposed fish at 0 ppt, accompanied by decreases in SOD, GST, GSH, and also catalase (CAT) activity. Our results confirm that sublethal waterborne zinc is an oxidative stressor in fish, and highlight the important protective role of higher salinities in ameliorating the oxidative stress associated with zinc toxicity in this model estuarine teleost.
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    Quantifying the direct and indirect effects of Dissolved Organic Matter (DOM) on aquatic organisms : interaction with pH and quality measures
    (McMaster University, 2012-09) Al-Reasi, Hassan Ali
    This thesis has advanced understanding of the relationships between dissolved organic matter (DOM) quality and its indirect and direct effects on aquatic organisms, and points to new directions for future work. DOM in natural waters is a heterogeneous mixture of organic molecules with direct and indirect influences on aquatic organisms. Aquatic natural organic matter (NOM) is composed of extremely heterogeneous mixtures of organic compounds. The major constituents of NOM are humic substances generated from dead organic biomass through humification, a process which is poorly understood yet one of the most important ecosystem functions on the earth.
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    Investigations into the formation of rainbow trout (Oncorhynchus mykiss) social hierarchies and possible hierarchical disruption by an environmental perturbation
    (Department of Biology, McMaster University, 2011-12) Grobler, Josias; Department of Biology, McMaster University
    In environments with limiting resources, such as space or food, Salmonids such as rainbow trout form social hierarchies, consisting of dominant and subordinate individuals. There were two main objectives for this thesis: to investigate the physiological consequences of being in a social hierarchy, and secondly, to investigate the behavioural and physiological changes of groups of rainbow trout exposed to ammonia concentrations which are above chronic protected guidelines.
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    Effect of Pb, Zn, Cu and Ni on the embryonic and larval stages of S. purpuratus
    (Department of Biology, McMaster University, 2012) Tellis, Margaret Silvia; Department of Biology, McMaster University
    In light of the paucity of information on the toxicity of lead (Pb), zinc (Zn), copper (Cu) and nickel (Ni) in the marine environment, the aim of this thesis was to generate data on the mechanisms of toxicity of Pb, Zn, Cu and Ni in the early life stages of a very sensitive marine organism, the purple sea urchin (Strongylocentrotus purpuratus). Previous studies in other systems indicated that these metals can impact ionoregulation, especially calcium homeostasis, so an initial study focused on ionoregulatory changes during the first 96h of development under control conditions in embryonic and larval stages of S. purpuratus. Control tests showed that patterns of Ca, K, Na and Mg had an interesting pattern of accumulation over 96 h development and that the most pronounced changes were observed during the gastrulation stage. A variety of biological endpoints were subsequently utilized to examine potential mechanisms of toxicity. Toxicity tests were performed to determine median toxicity threshold values (i.e. EC50: median effective concentration and LA50: median lethal accumulation) for Zn and Pb. Growth, unidirectional Ca uptake rates, whole body ion concentrations (Na, K, Ca, Mg), and Ca ATPase activity were also monitored every 12 h over the first 84 or 96 h of early development to investigate the mechanisms of toxicity during acute and chronic exposures to lethal and sublethal concentrations of Pb, Zn, Cu and Ni in 100% sea water. Sea urchin embryos were very sensitive to Zn with an EC50 of 2.3 µmol/L (95% C.I. = 1.97–2.71 μmol/L) and LA50 of 4.8 (2.16-11.33) μmol/kg. Embryos displayed even higher sensitivity to Pb with an EC50 of 0.36 (0.25-0.49) µmol/L and LA50 of 1.92 (1.67-2.78) μmol/kg. The toxic effects of these metals were increased when tested in combination with extracts of dissolved organic carbon (DOC) from marine and freshwater sources. From studying larvae chronically exposed to Pb (60 µg/L), Zn (139 µg/L), Cu (6 µg/L) and Ni (47 µg/L) it was apparent that these metals rendered their toxic effects, at least in part, through disruption of Ca homeostasis. Unidirectional Ca uptake rates as well as Ca ATPase activity were significantly inhibited at various time points over development, in larvae in these metal exposures. This resulted in significantly lower levels of Ca accumulated in the larvae. Interestingly, larvae showed some capacity for recovery as Ca uptake rates and internal Ca levels returned to control values periodically over development. Also metal initially accumulated in the larvae often returned to control levels at 72 h of development with the exception of Pb, which was the only non essential metal out of the four tested. Metal effects on the whole body levels of three other ions (Na, K, Mg) as well as larval weight were modest. Surprisingly, acute exposure to much higher levels of these same metals at various stages during development had negligible inhibitory effects on unidirectional Ca uptake rate, suggesting a mechanism other than direct competition for the Ca uptake sites. We propose studying the toxicity of contaminants periodically over development as an effective way to detect sub-lethal effects, which may not be displayed at the traditional endpoint of 72 h.
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    Efeito da temperatura na sobrevivência, consumo alimentar e crescimento de pós-larvas do camarão-rosa Farfantepenaeus paulensis
    (Open Journal Systems (OJS), 2012-02) Soares, Roberta; Peixoto, Sílvio; Bianchini, Adalto; Cavalli, Ronaldo; Wasielesky, Wilson
    Uma sequência de três experimentos foi realizada para avaliar o efeito da temperatura na sobrevivência, consumo alimentar e crescimento de pós-larvas de Farfantepenaeus paulensis. Foram avaliadas as faixas de resistência das pós-larvas (PL20) em altas e baixas temperaturas. Grupos de pós-larvas foram submetidos a elevação ou redução gradual (1°C por dia) da temperatura da água até a mortalidade total ser observada. A temperatura mínima para sobrevivência de 100% dos animais foi estimada em 13,0°C e a mortalidade total em 8,3°C. Já a temperatura máxima para sobrevivência de 100% dos animais foi estimada em 33,2°C e a mortalidade total em 40,4°C. Para determinação da temperatura letal média (LT50) pós-larvas foram submetidas a seis temperaturas, variando de 7°C a 14°C (baixas temperaturas) e de 30°C a 40°C (altas temperaturas) por 96 horas. O LT50 inferior foi estimado em 9.3°C, enquanto o LT50 superior em 35.5°C. O crescimento e o consumo de Artemia pelas pós-larvas foram medidos nas temperaturas 18; 20; 23; 26; 29 e 32°C durante 30 dias. Foi observado um incremento significativo (p<0,05) no crescimento dos animais e no consumo de alimento com o aumento da temperatura. Os resultados sugerem que as pós-larvas sejam cultivadas em tanques de berçário com temperaturas entre 29-32°C. Já para berçários realizados diretamente no ambiente estuarino da Lagoa dos Patos recomenda-se que sejam limitados aos meses de primavera e verão.
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    New insights into gill epithelial transport: Linking ammonia excretion and sodium uptake
    (2011) Wright, P A; Wood, C M; Cooper, C A
    Models of gill epithelial transport are rapidly changing as researchers discover more complex and specialized cell types and use genomic data to uncover completely new protein transporters. Maetz and Garcia Romeu (1964) first suggested a linkage between ammonia excretion and active Na+ uptake at the gills of freshwater animals. Although many studies have provided supporting evidence for such a linkage, only in the last 4 years with the discovery of ammonia-transporting Rhesus (Rh) glycoproteins in gill tissue have we been able to more fully understand the model of gill ammonia transport in freshwater animals. Recent evidence supports an apical “Na+/NH4 + exchange complex” consisting of several membrane transporters (Rhcg, V type H+ATPase, Na+/H+ exchanger (NHE 2/3, Na+ channel) working together as a metabolon providing an acid-trapping mechanism for apical excretion in freshwater fish. The model for gill ammonia transport in seawater is more controversial, although Rhcg also appears to be associated with Na+ transport.
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    Mechanisms of sodium extrusion in isolated mitochondria-rich cells of the freshwater mussel Lasmigona costata after copper exposure
    (2011) Nogueira, L S; Wood, C M; Bianchini, A; Gillis, P L
    Gill tissues in aquatic animals are composed of various cell types, each with specific functions such as respiration, excretion, acid-base balance and ionic regulation. In the bivalves, the gill is the main route of contaminant uptake and metals can be absorbed through specific transporters in cell membranes. Among the gill cells, mitochondria-rich cells (MRCs) are considered to be the primary sites responsible for iron transport. In order to determine the mechanisms of sodium extrusion in MRCs after copper exposure, gill cells from Lasmigona costata were isolated and layered on a discontinuous Percoll gradient. The pavement cells (PCVs) and MRCs populations were confirmed by a specific mitochondrial dye (DASPEI) and Na, K-ATPase activity. It is the first time at literature that isolated fill cells from freshwater mussels are identified in two different populations and used in toxicological experiments. MRCs were then exposed to a range of copper concentrations environmentally relevant (5, 10 and 20 ug/L) for 3 hours. Copper accumulation increased in all copper exposures. In contrast, the sodium content did not change in low exposures and decreased significantly when cells were exposed to high copper concentrations. To further explore this result, both sodium and copper were measured after specific pharmacological blockade of Na, K-ATPase and carbonic anhydrase enzymes. Carbonic anhydrase inhibition suggest that sodium and copper uptake mechanisms are not dependent on a supply of protons and intracellular increase of both ions might be associated to blocked of Na extrusion via Na/HCO3-. The inhibition of Na, K-ATPase by ouabain also resulted in an increase in sodium and copper content in the MRCs. This enzyme is responsible for sodium extrusion from the cell, and the results suggest that it is also a route for copper extrusion.
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    Avaliação da lipoperoxidação em peixe como potencial biomarcador de contaminação aquática
    (2011) Gomes, E G; Machado, A A S; Hoff, M L M; Bianchini, A
    A contaminação aquática por pesticidas, metais e hidrocarbonetos pode ser monitorada usando-se biomarcadores. A lipoperoxidação (LPO) é um biomarcador que pode ser induzido pela exposição a contaminantes como atrazina (pesticida), cobre (metal) e fenantreno (hidrocarboneto). Avaliar LPO em músculo, brânquia e fígado do peixe Poecilia viviparaapós exposição a cobre, atrazina e fenantreno. Machos de P. vivípara foram coletados no Arroio do Gelo (Rio Grande, RS), aclimatados à salinidade 24, fotoperíodo 12:12 h C:E e 20°C, e expostos (96 h) aatrazina(controle, controle metanol, 2, 10, 100µg/L), Cu (controle, 5, 9 e 20µg Cu/L) e fenantreno (controle, controle DMSO, 10, 20, 200µg/L) em água do mar filtrada (0,5 µm). Após expostos, os peixes foram anestesiados e retirados brânquias, fígado e músculo. A LPO foi avaliada nestes tecidos através do método TBARS. ANOVA, Kruskal Wallis e coeficiente de correlação linear de Spearman foram utilizados. Os peixes expostos ao cobre apresentaram um aumento da LPO branquial e hepática em função da concentração do metal (brânquia r = 0,83; fígado r = 0,90). A exposição a 100 µg/L de atrazina induziu aumento da LPO branquial. Por sua vez, a exposição a 10 µg/L causou uma diminuição da LPO muscular. No fígado, observou-se uma tendência de diminuição da LPO com o aumento da concentração de atrazina (r = -0,88). A exposição a 10 µg/L de fenantreno causou um aumento da LPO branquial. No músculo, este contaminante causou um aumento da LPO em função da concentração (r = 0,94). Por sua vez, não foi observada variação significativa na LPO hepática. Os resultados obtidos indicam que o cobre, a atrazina e o fenantreno afetam a LPO em tecidos de P. vivipara, porém o efeito é dependente do tecido analisado e da concentração testada. Assim, um aumento da LPO muscular remete à exposição ao fenantreno, enquanto uma diminuição da LPO hepática em paralelo a um aumento da LPO branquial remete à exposição à atrazina e um aumento da LPO hepática conjuntamente a um aumento da LPO branquial remete à exposição ao cobre. Estes resultados indicam que a LPO se constitui em um potencial biomarcador para identificar a exposição de peixes à contaminação por fenantreno, atrazina e cobre em ambientes estuarinos.