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- A multibiomarker approach in the clam Ruditapes decussatus to assess the impact of pollution in the Ria Formosa lagoon, South Coast of PortugalPublication . Cravo, Alexandra; Pereira, C.; Gomes, Tânia; Cardoso, Cátia; Serafim, M.A.; Almeida, Cheila; Rocha, T.; Lopes, Belisandra; Company, Rui; Medeiros, A.; Norberto, R.; Pereira, R.; Araújo, O.; Bebianno, Maria JoãoThe Ria Formosa lagoon is an ecosystem whose water quality reflects the anthropogenic influence upon the surrounding areas. In this lagoon, the clam Ruditapes decussatus has a great economical importance and has been widely used as a biomonitor. A multibiomarker approach (d-aminolevulinic acid dehy- dratase, metallothionein, lipid peroxidation, acetylcholinesterase, alkali-labile phosphates, DNA damage) was applied to assess the environmental quality of this ecosystem and the accumulation of contaminants and their potential adverse effects on clams. Clams were sampled in different shellfish beds in the period between July 2007 and December 2008 and abiotic parameters (temperature, salinity, pH and dissolved oxygen of seawater and organic matter in the sediment), condition index, metals (Cd, Cu, Zn, Ni, Pb), TBTs and PAHs concentrations were measured in clam tissues. Data was integrated using Principal Component Analyses and biomarker indices: IBR (Integrated Biomarker Response) and HSI (Health Status Index). This multibiomarker approach enabled discrimination of a time and space trend between sites with different degrees of anthropogenic contamination, identifying one of them (site 2) as the most stressful and summer months as the most critical period for clams due to an increase of environmental stress (anthropogenic pressure along with extreme environmental conditions, e.g. temperature, dissolved oxygen, organic matter in the sediments, etc). The selected biomarkers provided an integrated response to assess the environmental quality of the system, proving to be a useful approach when complex mixtures of contaminants occur.
- Responses of CYP450 dependent system to aliphatic and aromatic hydrocarbons body burden in transplanted mussels from South coast of PortugalPublication . Lopes, Belisandra; Ferreira, A. M.; Bebianno, Maria JoãoMussels Mytilus galloprovincialis were cross-transplanted at South Portugal from a reference site (site 1) to a site more contaminated with hydrocarbon compounds (site 2), and vice versa, in an active biomonitoring (ABM) concept, to assess the biotransformation capacity catalyzed by the mixed function oxygenase (MFO) system. Total alkanes (TAlk), the unresolved complex mixture (UCM), and total polycyclic aromatic hydrocarbons (TPAHs) concentration increased respectively 6, 4.4 and 4.2 fold relatively to control, in mussels transplanted from site 1 to 2. In the cross-transplant, a 48, 57 and 62% depuration of TAlk, UCM and TPAHs concentrations occurred by the end of the 3-4th week. Petrogenic and biogenic (marine and terrigenous) sources of AHs, and petrogenic and pyrolitic (biomass and oil/fuel incomplete combustion) sources of PAHs were detected at both sites. CYP450, CYT b (5) and NADPH-RED in mussels transplanted from site 1 to 2 were induced from day 0 to 28, with a total increase of 35, 32 and 35%, respectively, while biochemical equilibrium to lesser environmental contamination occurs in mussels transplanted from site 2 to 1. A significant relationship between CYP450 and NADPH-RED was found with TPAH, with distinctive behavior at the two sites. MFO system components increase with exposure time at one site and decreases in the other, reflecting an adaptation to distinct environmental hydrocarbon loads. The ABM strategy proved to be useful to understand the environment real impact on the biochemical responses in mussels' local populations. In this study, CYP450 and NADPH-RED are a useful biomarker for hydrocarbon exposure.
- Multi-biomarker responses to estuarine habitat contamination in three fish species: Dicentrarchus labrax, Solea senegalensis and Pomatoschistus micropsPublication . Fonseca, V. F.; França, S.; Serafim, M.A.; Company, Rui; Lopes, Belisandra; Bebianno, Maria João; Cabral, H. N.Several biomarker responses were determined in three fish species, Dicentrarchus labrax, Solea senegalensis and Pomatoschistus microps, from two estuaries of the Portuguese coast, Ria de Aveiro and Tejo. Both estuaries have significant anthropogenic influences from multiple sources (industrial, agricultural and shipping activities), which was evident from sediment chemical characterization concerning metal (copper, zinc, nickel, lead and chromium) and polycyclic aromatic hydrocarbon (PAH) concentrations. Spatial variability in fish responses was observed across species for most biomarkers of exposure [the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), ethoxyresorufin O-deethylase (EROD) and glutathione S-transferase (GST), and metallothionein concentrations (MT)] and effect biomarkers [lipid peroxidation (LPO), RNA to DNA ratio (R:D), protein and lipid content]. In general, the interspecific differences in biomarker responses were greater than the spatial differences, due to differences in the behavior and habitat use of the species. Nevertheless, similarities were also observed considering both chemical load and biomarker responses. In highly polluted sites fish showed in general a significant antioxidant enzyme induction, associated with decreased R:D values, while fish from the least impacted site had little enzyme induction and better condition indices (high R:D and low LPO values). EROD activity was also higher for all species in the Tejo than Ria de Aveiro estuary, despite the generally higher total PAH measured in Ria de Aveiro, most likely due to a higher proportion of 4 and 6-ring PAHs, considered more toxic than low molecular weight PAHs, in the Tejo. In conclusion, this multi-biomarker approach considering multiple species provided improved understanding of the diverse responses and effects of exposure to contaminants and the effective risk it poses for different fish species.
- A multibiomarker approach in Mytilus galloprovincialis to assess environmental qualityPublication . Cravo, Alexandra; Lopes, Belisandra; Serafim, M.A.; Company, Rui; Barreira, Luísa; Gomes, Tânia; Bebianno, Maria JoãoA multibiomarker approach was carried out for the first time in the South Portuguese Coast using Mytilus galloprovincialis, to assess environmental quality, establish if there are adverse biological responses associated to different sources of anthropogenic contamination and to determine spatial and seasonal trends. For this purpose the battery of biomarkers selected was: superoxide dismutase (SOD), catalase (CAT), glutathione peroxidases (GPx total and Se dependent), Cytochrome P450 component system, Glutathione-S-transferase (GST) and acetylcholinesterase (AChE), metallothionein (MT) and lead-delta-aminolevulinic acid dehydratase (ALAD), lipid peroxidation (LPO) and Condition Index (CI) along with the determination of PAHs and metals (Cd, Cu, Ni, Pb and Zn). Results show that despite the levels of both organic and metallic contaminants in these eight spots in the South Coast of Portugal not being particularly high compared with other contaminated/polluted sites worldwide, the selected battery of biomarkers responded efficiently to the environmental changes and allowed an environmental assessment between seasons and sites. Different spatial and seasonal responses were evident along the South Coast of Portugal, meaning that the contamination is not homogeneous. This does not only reflect different competition, origin and intensity of contamination, but also different environmental conditions (e.g. temperature, salinity). Along the South Portuguese Coast site 8 was the most contaminated, while site 2 was considered the least contaminated. Despite environmental factors possibly causing difficulties in the general interpretation of biomarker data, those that better responded to environmental contamination were CYP450, SOD-mit and T-GPx for the summation SigmaPAHs, MT (digestive gland) for metals (especially Cu), ALAD for Pb and LPO for both organic and metallic contamination. These biomarkers were also positively correlated with temperature in summer, revealing this as a more stressful/critical season. In future environmental contamination assessments there is no need to analyse the components b5, P418, NADH and NADPH of phase I MFO system, and MT in the gills, since their responses are not evident.
- Modeling fish biological responses to contaminants and natural variability in estuariesPublication . Fonseca, V.F.; Vasconcelos, R.P.; França, S.; Serafim, M.A.; Lopes, Belisandra; Company, Rui; Bebianno, Maria João; Costa, M.J.; Cabral, H.N.Understanding the factors that influence biological responses to contaminants has long been a major goal in marine environmental research. Seven estuarine sites along the Portuguese coast were sampled over a year, and different biological responses of Pomatoschistus microps and Atherina presbyter were determined: superoxide dismutase, catalase, ethoxyresorufin O-deethylase, glutathione S-transferase, metallothioneins, lipid peroxidation, RNA:DNA ratio and condition factor K. Generalized linear models (GLM) were developed for each biological variable per species in relation to sediment chemical characterization (metals and polycyclic aromatic hydrocarbons concentration) and environmental conditions (month, site, water temperature, salinity, depth and mud percentage in the sediment). GLM varied in explanatory power and in the set of predictor variables included in the models. Environmental factors were frequently selected as predictor variables. Individual metals concentration and sediment quality guidelines (integrating all metals) were the major contaminants explaining biological variability. Accordingly, models for metallothioneins and lipid peroxidation had highest explanatory power. Species-specific responses and dataset size were the basis of observed differences between GLM for the two species.
- A multi-biomarker approach in cross-transplanted mussels Mytilus galloprovincialisPublication . Serafim, M.A.; Lopes, Belisandra; Company, Rui; Cravo, Alexandra; Gomes, Tânia; Serrão Sousa, Vânia; Bebianno, Maria JoãoThe present work integrates the active biomonitoring (ABM) concept in mussels Mytilus galloprovincialis from the South coast of Portugal transplanted during 28 days between two sites with different sources of contamination, and vice versa, in order to assess biological effects in these mussels. For that purpose a multibiomarker approach was used. The suit of biomarkers indicative of metal contamination were metallothioneins (MT) and the enzyme δ-aminolevulinic acid dehydratase (ALAD), for organic contamination mixed function oxidase system (MFO), glutathione-S-transferase (GST) and acetylcholinesterase (AChE), as oxidative stress biomarkers superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and lipid peroxidation (LPO). These biomarkers were used to determine an index to evaluate the stress levels in these two sites. Site A is strongly influenced by metallic contamination, with higher Cu, Cr and Pb in M. galloprovincialis, as well as higher MT levels, antioxidant enzymes activities and LPO concentrations, and lower ALAD activity. In site B organic compounds (PAHs) are prevalent and native mussels show higher activities of the MFO system components and GST. Transplanted mussels had significant alterations in some biomarkers that reflect the type of contaminants present in each site, which demonstrates the primary role of the environment in determining the physiological characteristics of resident mussels. Therefore the application of ABM using a battery of biomarkers turns out to be a useful approach in sites where usually complex mixtures of contaminants occurs. In this study the biomarkers that better differentiate the impact of different contaminants at each site were MT, CYP450, SOD and CAT.
- Using biochemical and isotope geochemistry to understand the environmental and public health implications of lead pollution in the lower Guadiana River, Iberia: A freshwater bivalve studyPublication . Company, Rui; Serafim, Angela; Lopes, Belisandra; Cravo, A.; Shepherd, T. J.; Pearson, G.; Bebianno, Maria João; Cravo, AlexandraLead is a natural component of aquatic ecosystems with no known biological role and is highly toxic. Its toxicity stems from its ability to mimic biologically important metals and to produce membrane damage through lipid peroxidation (LPO). Most lead poisoning symptoms are thought to occur by interfering with an essential enzyme, delta-aminolevulinic acid dehydratase (ALAD), the activity of which is markedly inhibited by lead. The purpose of this work was to study the levels and effects of lead pollution (responses of ALAD and oxidative stress biomarker LPO) in the freshwater bivalve Corbicula fluminea along the lower Guadiana River (Portugal and Spain); a major river system impacted by historic mining pollution and more recent anthropogenic inputs. The results show that the enzymatic activity of ALAD is negatively correlated with the total Pb concentration of the whole tissue suggesting that ALAD has considerable potential as a biomarker of lead exposure in C. fluminea. To identify the sources of lead to which bivalves have been exposed, high precision (206)Pb/(204)Pb, (207)Pb/(204)Pb, (208)Pb/(204)/Pb ratios for C. fluminea confirm that historical mining activities in the Iberian Pyrite Belt are the dominant source of lead pollution in the lower Guadiana River. The isotope patterns however exhibit marked seasonal and geographic variation in response to rainfall and river water management. Locally, other anthropogenic sources of lead have been detected in C. fluminea close to population centres, thus adding to its versatility as a freshwater bio-indicator. Overall, the study highlights the value of natural ecosystems as monitors of water quality and their importance for public health assessment and surveillance.