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- DNA damage and lipid peroxidation vs. protection responses in the gill of Dicentrarchus labrax L. from a contaminated coastal lagoon (Ria de Aveiro, Portugal)Publication . Ahmad, I.; Maria, Vera L.; Oliveira, M.; Serafim, Angela; Bebianno, Maria João; Pacheco, M.; Santos, M.A.The present research work aimed to investigate the damage vs. protection responses in gill of European sea bass (Dicentrarchus labrax) captured at a polluted coastal lagoon, Ria de Aveiro (Portugal), as a tool to evaluate the human impacts on environmental health. Damage was assessed as DNA strand breakage and lipid peroxidation (LPO) whereas protection was evaluated by measuring catalase (CAT), glutathione peroxidase (GPX), glutathione reductase (GR), glutathione S-transferase (GST), total glutathione (GSHt), thiols and metallothioneins (MT). Fish were caught at five locations: Gafanha (GAF), Rio Novo do Príncipe (RIO), Laranjo (LAR) and Vagos (VAG) presenting each a different recognized source of contamination, and Torreira (TOR), assumed as reference site. Among the surveyed sites, gill damage was observed only at GAF, as measured by DNA integrity loss and LPO increase. An overall induction in enzymatic antioxidant protection was perceptible in fish from GAF and VAG, expressed as higher GPX, GR and GST activities. In addition, LAR fish showed elevated GST and CAT activities. Thiols content was higher in all study sites irrespective to the pollution spectrum, whereas GSHt increase was only observed at GAF and VAG. The highest MT level was detected in fish from VAG and the lowest level from RIO suggesting a low contamination degree on this particular site. Globally, the results expressed site-specific response patterns, signalling two critical areas - GAF and VAG. Additionally, a joint analysis (damage vs. protection) allowed the following ordering of surveyed sites according to the contamination degree and risk to fish health; GAF>VAG>LAR>RIO>TOR. D. labrax gill responses demonstrated their efficacy as early warning signals of the contaminants presence. Moreover, the adopted approach, considering simultaneously protection responses and damaging effects, also revealed its usefulness on the pollution extent assessment.
- Antioxidant and lipid peroxidation responses in Mytilus galloprovincialis exposed to mixtures of benzo(a)pyrene and copperPublication . Maria, Vera L.; Bebianno, Maria JoãoThis study aimed to assess the antioxidant system potential and lipid peroxidative effects, in the gill and digestive gland of Mytilus galloprovincialis exposed to individual and binary mixtures of benzo(a)pyrene (BaP) and Cu for 7 days. Data demonstrated that in mussels exposed to BaP antioxidant enzymes (catalase--CAT, total glutathione peroxidase--tGPx, glutathione S-transferase--GST and glutathione reductase--GR) and lipid peroxidation (LPO) increased in the gill. On the contrary, in the digestive gland inhibitory antioxidant effects (superoxide dismutase-SOD, GR, metallothioneins-MT) and no changes in LPO levels were detected. Cu was also a potent oxidant agent since MT and LPO levels increased in mussel gill, despite no LPO effect in the digestive gland. For both single contaminants the organ specificity and distinct physiologic/metabolism roles were evident in terms of antioxidant capacity. Gill SOD inhibition, MT and GST unchanged was a result of "simple independent action" of exposure to BaP and Cu. "Interactions" in the binary mixtures, led to absence of changes in LPO effects. In the digestive gland, BaP and Cu interactions were also responsible for the GST and LPO enhancement (antagonistic effects). The current findings demonstrate the differences in antioxidant responses where the organ dependency highlights each contaminant particular mode of action. Generally, in the gill "non-interactive" effects occurred with the lowest Cu concentration while "interactions" exist for the mixture with the highest Cu concentrations. In the digestive gland, "interactions" and "no interaction" effects occurred in all the binary mixtures. Complex contaminant mixtures interact differently based on target tissue which may lead to an imbalance in the mussels health status.
- Hepatic metallothionein concentrations in the golden grey mullet (Liza aurata): relationship with environmental metal concentrations in a metal-contaminated coastal system in PortugalPublication . Oliveira, M.; Ahmad, I.; Maria, Vera L.; Serafim, A.; Bebianno, Maria João; Pacheco, M.; Santos, M.A.This field survey was designed to assess the environmental metal contamination status of Ria de Aveiro (Portugal). To achieve that goal, the concentrations of Cd, Hg, Cu and Zn in the sediments and water were assessed and Liza aurata hepatic metallothionein (MT) determined. The relationships between MT and environmental metal concentrations and hydrological factors were examined. Results revealed a wide distribution of metals both in water and sediments throughout the lagoon, mainly at Rio Novo do Principe (RIO) and Laranjo (LAR), at concentrations that may affect biota. MT concentrations were higher at the sites with high metal content (RIO and LAR). A significant positive correlation was found between MT and Cd in the sediments as well as with MT and Hg and Cu in the water. Moreover, a negative correlation between MT and salinity was found. Thus, the current data support MT use as a biomarker of metal exposure emphasizing the importance of hydrological parameters in its concentrations. Results suggest the continued monitoring of this lagoon system.
- Comparison of metal accumulation between ‘Artificial Mussel’ and natural mussels (Mytilus galloprovincialis) in marine environmentsPublication . Gonzalez-Rey, Maria; Lau, T. C.; Gomes, Tânia; Maria, Vera L.; Bebianno, Maria João; Wu, R.The passive sampler called 'Artificial Mussel' (AM) developed by Wu et al. (2007) can provide a time-integrated estimate of metal concentrations in the marine environment, and offers a potential device to assess and compare metal concentration in different marine environments worldwide. The aim of this study was to compare metal accumulation on AM and natural mussel Mytilus galloprovincialis at three sites with different metal loads along the Portuguese coast for four months. M. galloprovincialis were placed in cages alongside AMs at each site. Samples were collected monthly and Cd, Cr, Cu, Pb, and Zn concentrations in whole soft tissues and AMs compared. For both Cu and Cd, the results were similar between AMs and natural mussels. Higher concentrations of Zn were observed in natural mussels, whereas the inverse was shown for Pb (about 10-fold higher). Our results showed that AMs are promising tools for assessing metal concentrations in marine environments.
- Contaminant effects in shore crabs (Carcinus maenas) from Ria Formosa LagoonPublication . Maria, Vera L.; Santos, M. A.; Bebianno, Maria JoãoDefence and damage biomarkers signals were studied in female and male shore crabs Carcinus maenas transplanted between two sites at Ria Formosa Lagoon (South of Portugal). The cross transplantation occurred during 6 days at a hypothetical reference site (Ramalhete), and a contaminated site (Olhão). DNA unwinding technique was used for DNA integrity measurement. General enzymatic antioxidant responses in gills and hepatopancreas (catalase and gills glutathione peroxidase activities increase) showed that cross transplanted crabs (female and male) are suffering from pro-oxidant challenges at the Olhão site. Gills and hepatopancreas glutathione-S-transferase were reduced in both gender crabs transplanted from Ramalhete to Olhão. Metallothioneins induction occurred in crabs transplanted from Ramalhete to Olhão (contaminated by metals and polycyclic aromatic hydrocarbons (PAHs)). However metallothionein differences were also observed towards gender and organ specificities. High gills lipid peroxidation exist in male crabs transplanted from Ramalhete to Olhão, while in females it was the opposite. In both gender crabs from the Olhão site, gills DNA integrity decreased compared to the Ramalhete feral crabs. Moreover, hepatopancreas DNA integrity decreased in male crabs transplanted from Olhão to Ramalhete site which may be related to the environmental conditions (lower contamination levels) revealing the difficulty of selection of reference sites in field studies. Data demonstrated that female and male C. maenas antioxidant defences and damage biomarkers were sensitive to the mixture of contaminants present in these sites as well as good indicators of general stress.
- Contamination assessment of a coastal lagoon (Ria de Aveiro, Portugal) using defence and damage biochemical indicators in gill of Liza aurata – An integrated biomarker approachPublication . Oliveira, M.; Maria, Vera L.; Ahmad, I.; Serafim, Angela; Bebianno, Maria João; Pacheco, M.; Santos, M. A.Fish gill importance in toxicants uptake, bioconcentration and excretion allied to meagre knowledge on branchial damage/protection responses substantiate this study. Five critical sites in Ria de Aveiro (Portugal) were assessed in comparison with a reference site (Torreira), focusing on Liza aurata gill antioxidant defences versus damage (oxidative and genetic). Only in Barra fish displayed damage (lipid peroxidation) though no differences were found in antioxidants. In all other sites, except Rio, antioxidant alterations were found. Thus, fish from Gafanha, Laranjo and Vagos showed higher total glutathione, glutathione peroxidase and catalase. Higher glutathione reductase and glutathione S-transferase activity was also found in the first and the last sites, respectively. In Laranjo, metallothionein levels were higher though lower in Gafanha and Vagos. In general, damage was not accompanied by defences weakening confirming that predicting damage based on antioxidants depletion is not straightforward. The integrated biomarker response index ranked sites as: Gafanha>Barra>Laranjo>Vagos>Rio>Torreira.
- Evaluation of oxidative DNA lesions in plasma and nuclear abnormalities in erythrocytes of wild fish (Liza aurata) as an integrated approach to genotoxicity assessmentPublication . Oliveira, M.; Ahmad, I.; Maria, Vera L.; Ferreira, C. S. S.; Serafim, A.; Bebianno, Maria João; Pacheco, M.; Santos, M. A.Genetic lesions (8-hydroxy-2'-deoxyguanosine (8-OHdG) and erythrocytic nuclear abnormalities (ENA) were seasonally quantified in the blood of Liza aurata caught at Ria de Aveiro (Portugal), a multi-contaminated aquatic system. Thus, five critical sites were assessed and compared with a reference site (Torreira). Oxidative DNA damage was found in Gafanha (harbour-water area), Laranjo (metal-contaminated) and Vagos (contaminated with PAHs) in the spring; Rio Novo do Principe (near a former paper-mill effluent) in the autumn; Rio Novo do Principe and Vagos in the winter. ENA were higher than Torreira at VAG (spring and winter). Torreira did not display seasonal variation neither in terms of 8-OHdG or total ENA. A positive correlation between 8-OHdG and ENA was found, suggesting oxidative stress as a mechanism involved in the formation of ENA. This study clearly demonstrates the presence of DNA-damaging substances in Ria de Aveiro and recommends the use of 8-OHdG and ENA as biomarkers of environmental contamination.
- Detection of emerging contaminants (UV filters, UV stabilizers and musks) in marine mussels from Portuguese coast by QuEChERS extraction and GC–MS/MSPublication . Picot Groz, M.; Martinez Bueno, M.J.; Rosain, D.; Fenet, H.; Casellas, C.; Pereira, Catarina; Maria, Vera L.; Bebianno, Maria João; Gomez, E.The UV filters and musk fragrances have come into focus because these compounds are contained and increasingly used not only in sunscreen products but also in many products of daily use, such as cosmetics, skin creams, plastics or varnish. In view of this, the main objective of the present work was to develop and validate a method for the determination of three UV filters, two UV stabilizers and four musks in mussel samples (Mytilus galloprovincialis). The procedure combined a QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) extraction method with an analysis by gas chromatography-tandem mass spectrometry (GC-MS/MS). The methodology allowed the determination of target analytes at trace concentration levels (ng/g), with mean recoveries ranging from 91 to 112%. A monitoring study was conducted in four beaches in the Portuguese coast which are impacted by recreational activities and outflow of treated waste water effluents in rivers. The results are used to assess the occurrence of UV filters in comparison with UV stabilizers and musk fragrances which indicate other activities than bathing.