Indianara Fernanda Barcarolli

Waterborne copper exposure inhibits ammonia excretion and branchial carbonic anhydrase activity in euryhaline guppies acclimated to both fresh water and sea water

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).

The effects of salinity on acute and chronic nickel toxicity and bioaccumulation in two euryhaline crustaceans: Litopenaeus vannamei and Excirolana armata.

We investigated the influence of salinity (5 ppt versus 25 ppt) on acute (96-h LC(50)) and chronic toxicity (15-30 day LC(50)) of Ni in two euryhaline crustaceans, the shrimp (Litopenaeus vannamei) and the isopod (Excirolana armata). 96-h LC50 values were 41 μmolL(-1) and 362 μmolL(-1) for L. vannamei and 278 μmolL(-1) and >1000 μmolL(-1) for E. armata at 5 ppt and 25 ppt, respectively. Speciation analysis demonstrated that complexation with anions such as SO(4)(2-), HCO(3)(-) and Cl(-) at 25 ppt had a negligible effect on reducing the free Ni(2+) ion component in comparison to 5 ppt. The salinity-dependent differences in acute Ni toxicity could not be explained by differences in Ni bioaccumulation. Therefore, differences in physiology of the organisms at the two salinities may be the most likely factor contributing to differences in acute Ni toxicity. Chronic LC(50) values (2.7-23.2 μmolL(-1)) were similar in the two species, but salinity had no significant effect, indicating that water chemistry and osmoregulatory strategy do not influence chronic toxicity. However chronic (15-day) mortality in both species could be predicted by acute (96-h) Ni bioaccumulation patterns.

Effect of copper on ion content in isolated mantle cells of the marine clam Mesodesma mactroides

The effect of copper on ion content (Na(+), K(+), Ca(2+), and Cl(-)) was evaluated in isolated mantle gills of the marine clam Mesodesma mactroides. Clams were collected at the Mar Grosso Beach (São José do Norte, Rio Grande do Sul [RS], southern Brazil), cryoanesthetized, and had their mantles dissected. Mantle cells were isolated and incubated in a calcium-free phosphate solution without (control) or with Cu (CuCl(2)). Cells were exposed to Cu for 1 h (5 µM) or 3 h (2.5 and 5 µM). In cells incubated with 2.5 µM Cu, a significant decrease in intracellular Cl(-) content was observed. However, in cells incubated with 5.0 µM Cu, significant reductions in Na(+), K(+), and Cl(-) intracellular content were observed. Given the mechanisms involved in ion transport in mantle cells of the marine clam M. mactroides, the findings described here suggest that Cu exposure inhibits carbonic anhydrase and Na(+)/K(+) -ATPase activity. Also, it can be suggested that Cu is competing with Na(+) for the same mechanisms of ion transport in the cell membrane, such as the Na(+) channels and the Na(+)/K(+)/2Cl(-) cotransporter. Results from the present study also clearly indicate that processes involved in cellular anion regulation are more sensitive to Cu exposure than those associated with the cellular cation regulation. Characterization of sites for Cu accumulation and toxicity in aquatic animals is important for derivation of metal binding constants at the biotic ligand. Also, identification of the mechanism of metal toxicity is needed for modeling metal accumulation in the biotic ligand and its consequent toxicity. Therefore, the findings reported here are extremely valuable for the development of a biotic ligand model version for marine and estuarine waters.

Metal contamination as a possible etiology of fibropapillomatosis in juvenile female green sea turtles Chelonia mydas from the southern Atlantic Ocean

Environmental contaminants have been suggested as a possible cause of fibropapillomatosis (FP) in green sea turtles. In turn, a reduced concentration of serum cholesterol has been indicated as a reliable biomarker of malignancy in vertebrates, including marine turtles. In the present study, metal (Ag, Cd, Cu, Fe, Ni, Pb and Zn) concentrations, oxidative stress parameters [antioxidant capacity against peroxyl radicals (ACAP), protein carbonyls (PC), lipid peroxidation (LPO), frequency of micronucleated cells (FMC)], water content, cholesterol concentration and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) activity were analyzed in the blood/serum of juvenile (29.3-59.5cm) female green sea turtles (Chelonia mydas) with FP (n=14) and without FP (n=13) sampled at Ubatuba coast (São Paulo State, southeastern Brazil). Green sea turtles were grouped and analyzed according to the severity of tumors. Individuals heavily afflicted with FP showed significantly higher blood Cu, Pb and Fe concentrations, blood LPO levels, as well as significantly lower serum cholesterol concentrations and HMGR activity than turtles without FP. Significant and positive correlations were observed between HMGR activity and cholesterol concentrations, as well as LPO levels and Fe and Pb concentrations. In turn, Cu and Pb concentrations were significantly and negatively correlated with HMGR activity and cholesterol concentration. Furthermore, Cu, Fe and Pb were positively correlated with each other. Therefore, the reduced concentration of serum cholesterol observed in green sea turtles heavily afflicted with FP is related to a Cu- and Pb-induced inhibition of HMGR activity paralleled by a higher LPO rate induced by increased Fe and Pb concentrations. As oxidative stress is implicated in the pathogenesis of viral infections, our findings support the idea that metal contamination, especially by Cu, Fe and Pb, may be implicated in the etiology of FP in green sea turtles through oxidative stress generation.

Concentrations and distributions of metals in tissues of stranded green sea turtles (Chelonia mydas) from the southern Atlantic coast of Brazil.

Silver (Ag), cadmium (Cd), copper (Cu), lead (Pb) and zinc (Zn) concentrations were analyzed in tissues of juvenile green sea turtles (Chelonia mydas) found stranded along the southern Atlantic coast in Brazil. Green sea turtles were collected (n=29), measured (curved carapace length: CCL) and had their muscle, liver, and kidney dissected for metal concentration measurements. Sex was identified in 18 individuals (10 females and 8 males) through gonad histology. No gender differences in CCL and tissue metal concentrations were observed. In the muscle, there was a negative correlation between CCL and Cd and Cu concentrations. Metal concentrations were lower in the muscle than in the liver and kidney. Zn concentration in the muscle was the highest of all metals analyzed (16.6 mg/kg). The kidney showed the highest concentrations of Pb, Cd and Zn (5.4, 28.3 and 54.3 mg/kg, respectively), while the liver had the highest values of Ag and Cu (0.8 and 100.9 mg/kg, respectively). Tissue Ag, Zn and Cd concentrations were similar to those found in green sea turtles from other regions while Cu and Pb values were elevated, likely due to the metal-rich water and sediment reported in the collection area. In the liver and kidney, concentrations of non-essential (Ag, Cd and Pb) and essential (Cu or Zn) metals were positively correlated, likely due to an induced metallothionein synthesis to protect tissue against the toxic effect of metals. This is the first study to report and correlate the concentrations of essential and non-essential metals in tissues of green sea turtles in the Brazilian southern Atlantic coast, an important feeding and developing area for this turtle species.

Effect of salinity on survival, growth and biochemical parameters in juvenile Lebranch mullet Mugil liza (Perciformes: Mugilidae)

Teleost fish growth may be improved under isosmotic condition. Growth and metabolic performance of juvenile Mugil liza (isosmotic point: 12‰) were evaluated after 40 days in different salinities (0, 6, 12 and 24‰). Tests were performed in quadruplicate (30 fish/tank; 0.48 ± 0.1 g body weight; 3.27 ± 0.1 cm total length) under controlled water temperature (28.2 ± 0.1oC) and oxygen content (>90% saturation). Fish were fed on artificial diet (50% crude protein) four times a day until apparent satiation. Results showed that salinity influenced juvenile mullet growth. Fish reared at salinity 24‰ grew better than those maintained in freshwater (salinity 0‰). Gill Na+,K+-ATPase activity and whole body oxygen consumption showed an U-shape-type response over the range of salinities tested, with the lower values being observed at the intermediate salinities. Although no significant difference was observed in liver glycogen content at different salinities, it tended to augment with increasing salinity. These findings indicate that energy demand for osmorregulation in juvenile M. liza can be minimized under isosmotic condition. However, the amount of energy spared is not enough to improve fish growth. Results also suggest that M. liza is able to alternate between different energy-rich substrates during acclimation to environmental salinity.

Acute waterborne copper toxicity to the euryhaline copepod Acartia tonsa at different salinities : influence of natural freshwater and marine dissolved organic matter

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.

Acclimation of juvenile Mugil liza Valenciennes, 1836 (Mugiliformes: Mugilidae) to different environmental salinities

A sobrevivencia e parâmetros fisiologicos associados ao metabolismo e a osmorregulacao foram avaliados em juvenis da tainha Mugil liza aclimatada a diferentes salinidades (5, 10, 20, 30 e 40‰) por 15 dias. Foram fixadas a temperatura (25oC) e o fotoperiodo (12L:12D) da sala onde os experimentos foram realizados. Os peixes foram alimentados duas vezes ao dia com racao comercial (28% de proteina bruta) ate a saciedade. Apos aclimatacao, foi medido o consumo corporal de oxigenio e os peixes foram eutanasiados e foram coletadas amostras de sangue, brânquias e figado. O consumo corporal de oxigenio e a osmolalidade plasmatica nao variaram na faixa de salinidade testada. O ponto isosmotico foi estimado em 412,7 mOsmol kg-1 (13,5‰). A atividade da Na+,K+-ATPase branquial tendeu a ser menor em 20 e 30‰, enquanto o conteudo de glicogenio hepatico foi significativamente maior em 20‰ do que em 5 e 40‰. Estes resultados indicam que o juvenil de M. liza e capaz de se aclimatar a uma ampla faixa de salinidade (5-40‰) por um curto periodo de tempo (15 dias). Esta condicao e atingida atraves de ajustes na atividade da Na+,K+-ATPase branquial e no metabolismo de carboidratos para regular a osmolalidade plasmatica e o metabolismo aerobico/energetico. Portanto, nossos achados suportam a ideia de que e possivel capturar juvenis da tainha M. liza em agua do mar e cultiva-los em aguas estuarinas e marinhas.