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Once in the marine environment, fishing nets and cables undergo weathering, breaking down into micro and nano-size particles and leaching plastic additives, which negatively affect marine biota. This study aims to unravel the ecotoxicological impact of different concentrations of leachate obtained from abandoned or lost fishing nets and cables in the mussel Mytilus galloprovincialis under long-term exposure (28 days). Biochemical biomarkers linked to antioxidant defense system, xenobiotic biotransformation, oxidative damage, genotoxicity, and neurotoxicity were evaluated in different mussel tissues. The chemical nature of the fishing nets and cables and the chemical composition of the leachate were assessed and metals, plasticizers, UV stabilizers, flame retardants, antioxidants, dyes, flavoring agents, preservatives, intermediates and photo initiators were detected. The leachate severely affected the antioxidant and biotransformation systems in mussels' tissues. Following exposure to 1 mg·L-1 of leachate, mussels' defense system was enhanced to prevent oxidative damage. In contrast, in mussels exposed to 10 and 100 mg·L-1 of leachate, defenses failed to overcome pro-oxidant molecules, resulting in genotoxicity and oxidative damage. Principal component analysis (PCA) and Weight of Evidence (WOE) evaluation confirmed that mussels were significantly affected by the leachate being the hazard of the leachate concentrations of 10 mg·L-1 ranked as major, while 1 and 100 mg·L-1 was moderate. These results highlighted that the leachate from fishing nets and cables can be a threat to the heath of the mussel M. galloprovincialis., This work was conducted under the framework of the JPI Oceans RESPONSE project (FCT JPIOCEANS/0005/2020), funded by the European Union through the Science and Technology Foundation (FCT) of Portugal. The authors thank FCT for the funds attributed to CIMA, University of Algarve UIDP/00350/2020 (doi.org/10.54499/UIDP/00350/2020), and LA/P/0069/2020 ((doi.org/10.54499/LA/P/0069/2020) granted to the Associate Laboratory ARNET.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g−1 dry weight. The method was successfully applied to wild mussels.

This study investigated the ecotoxicological effects of differently sized (4-6 µm and 20-25 µm) low-density polyethylene (LDPE) microplastics (MPs), with and without adsorbed benzo-a-pyrene (BaP), in clam Scrobicularia plana. Biomarkers of oxidative stress (superoxide dismutase-SOD; catalase-CAT), biotransformation (glutathione-S-transferases-GST), oxidative damage (lipid peroxidation-LPO) and neurotoxicity (acetylcholinesterase-AChE) were analysed in gills and digestive glands at different time intervals for a total of 14 days of exposure. In order to have a better impact perspective of these contaminants, an integrated biomarker response index (IBR) and Health Index were applied. Biomarker alterations are apparently more related to smaller sized (4-6 µm) MPs in gills and to virgin LDPE MPs in the digestive gland according to IBR results, while the digestive gland was more affected by these MPs according to the health index., Funding agencies:National funding agencies in the framework of JPI Oceans (Fundacao para a Ciencia e Tecnologia (FCT), Portugal JPIOCEANS/0005/2015FCT through University of Algarve UID/00350/2020

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g−1 dry weight. The method was successfully applied to wild mussels.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g−1 dry weight. The method was successfully applied to wild mussels.

Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to be the main pharmaceutical class accumulated in seafood. Among them, ibuprofen (IBU) is of special concern as it is used worldwide to treat common pain, does not require a medical prescription, it is often taken in a high daily dose, and has been reported to cause potential adverse effects on aquatic organisms. IBU is highly transformed into hydroxy- and carboxy-metabolites and/or degradation products generated not only after its administration but also during wastewater treatment or in the environment. These compounds can be present in the environment at higher concentrations than IBU and present higher toxicity. In this work, a low-cost and affordable routine analytical method was developed and validated for the first-time determination of IBU and its main metabolites in mussels. The method is based on ultrasound-assisted extraction (UAE), clean-up by dispersive solid-phase extraction (d-SPE) and analytical determination by liquid chromatography-tandem mass spectrometry. Box-Behnken experimental design was used for method optimisation to better evaluate the influence and interactions of UAE and d-SPE variables. Extraction recoveries were in the range from 81 to 115%. Precision, expressed as relative standard deviation, was lower than 7%. Method detection limits were in the range from 0.1 to 1.9 ng g−1 dry weight. The method was successfully applied to wild mussels.

This study investigated the ecotoxicological effects of differently sized (4-6 µm and 20-25 µm) low-density polyethylene (LDPE) microplastics (MPs), with and without adsorbed benzo-a-pyrene (BaP), in clam Scrobicularia plana. Biomarkers of oxidative stress (superoxide dismutase-SOD; catalase-CAT), biotransformation (glutathione-S-transferases-GST), oxidative damage (lipid peroxidation-LPO) and neurotoxicity (acetylcholinesterase-AChE) were analysed in gills and digestive glands at different time intervals for a total of 14 days of exposure. In order to have a better impact perspective of these contaminants, an integrated biomarker response index (IBR) and Health Index were applied. Biomarker alterations are apparently more related to smaller sized (4-6 µm) MPs in gills and to virgin LDPE MPs in the digestive gland according to IBR results, while the digestive gland was more affected by these MPs according to the health index., Funding agencies:National funding agencies in the framework of JPI Oceans (Fundacao para a Ciencia e Tecnologia (FCT), Portugal JPIOCEANS/0005/2015FCT through University of Algarve UID/00350/2020

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Whilst both plastic production and inputs at sea have increased since the 1950s, several modelling studies predict a further increase in the coming years in these respective quantities. We compiled scientific literature on trends in marine litter, consisting largely of plastic and microplastics in the ocean, understanding that monitoring programs or assessments for these aspects are varied, frequently focusing on limited components of the marine environment in different locations, and covering a wide spectrum of marine litter types, with limited standardization. Here we discuss how trends in the amounts of litter in the marine environment can be compared with the information provided by models. Increasing amounts of plastic are found in some regions, especially in remote areas, but many repeated surveys and monitoring efforts have failed to demonstrate any consistent real temporal trend. An observed steady state situation of plastic quantities in many marine compartments and the fate and transport of plastic in the marine environment remain areas for much needed further research.

Microplastics (MPs) lipophilic nature and widespread distribution raises concerns due to their increasing presence in the marine environment and their ability to adsorb organic contaminants, as being potential vehicles for transport and potential source of accumulation of organic contaminants by marine organisms. The organic UV-filter, oxybenzone (BP-3) is a constituent of sunscreens and personal care products, entering the marine environment either by direct contact with swimmers or by wastewater effluents. In this study the ecotoxicological effects of exposure to low-density polyethylene (LDPE) microplastics with and without adsorbed BP-3 were investigated in the peppery furrow shell clam, Scrobicularia plana. LDPE microplastics with a size range of 11-13 μm were previously contaminated with an environmentally relevant concentration of BP-3 (82 ng g-1). S. plana individuals were exposed to a concentration of 1 mg L-1 of microplastics with and without BP-3 adsorbed in a water-sediment exposure system for 14 days. Clams were sampled at the beginning of the experiment and after 3, 7, and 14 days of exposure. Multiple biomarkers were analysed to investigate the effect of exposure in different clam tissues, gills, digestive gland, and haemolymph. Antioxidant (superoxide dismutase, catalase, glutathione peroxidase) and biotransformation (glutathione-S-transferases) enzyme activities, oxidative damage (lipid peroxidation), genotoxicity (single and double strand DNA breaks), and neurotoxicity (acetylcholinesterase activity) were assessed along with two biomarker indexes to assess the overall health status. Results indicate that after 7 days of exposure MPs with adsorbed BP-3 induced oxidative stress and damage, when compared to exposure to virgin MPs and control treatments. Neurotoxic effects were also noted in MPs with adsorbed BP-3 after 14 days exposure, while some evidence points to increased genotoxicity with exposure time. Overall results indicate that gills were mo, Funding Agencies:JPI Oceans FCT JPIOCEANS/0005/2015EMERGEMIX project 02/SAICT/2017Portuguese Foundation for Science and Technology UID/MAR/00350/2013

The increasing use of rare earth elements (REEs) in diverse technological applications has augmented the demand and exploitation of these worldwide, leading to a higher input of REEs + Yttrium (Y) in the marine environment. The present study investigated the ecotoxicity of Lanthanum (La) and Y to Mytilus galloprovincialis developing embryos and juveniles. This was achieved by quantifying the embryogenesis success after 48 h, and survival of juveniles after 96 h of exposure to different concentrations of La and Y. Results show that both La and Y are more toxic to developing embryos and larvae than to juveniles of M. galloprovincialis. Predicted no-effect concentration (PNEC) values were also derived for the embryo development as a preliminary approach to assess the environmental risk for these compounds to marine organisms. Results revealed that La is more toxic than Y. The high sensitivity of the early developmental stages to these compounds highlight the relevance of including these stages when evaluating the toxicity of chemicals where little information is available. Although older life stages may be more tolerant to toxicants, the population survival will be compromised if new recruits are not viable, with implications to the whole ecosystem health and functioning of the impacted area. Information on the ecotoxicity of chemicals with expanded technological use and that may be released during deep-sea mining activities is urgent in order to help estimate environmental impacts.

The increasing use of rare earth elements (REEs) in diverse technological applications has augmented the demand and exploitation of these worldwide, leading to a higher input of REEs + Yttrium (Y) in the marine environment. The present study investigated the ecotoxicity of Lanthanum (La) and Y to Mytilus galloprovincialis developing embryos and juveniles. This was achieved by quantifying the embryogenesis success after 48 h, and survival of juveniles after 96 h of exposure to different concentrations of La and Y. Results show that both La and Y are more toxic to developing embryos and larvae than to juveniles of M. galloprovincialis. Predicted no-effect concentration (PNEC) values were also derived for the embryo development as a preliminary approach to assess the environmental risk for these compounds to marine organisms. Results revealed that La is more toxic than Y. The high sensitivity of the early developmental stages to these compounds highlight the relevance of including these stages when evaluating the toxicity of chemicals where little information is available. Although older life stages may be more tolerant to toxicants, the population survival will be compromised if new recruits are not viable, with implications to the whole ecosystem health and functioning of the impacted area. Information on the ecotoxicity of chemicals with expanded technological use and that may be released during deep-sea mining activities is urgent in order to help estimate environmental impacts.

The vent blood-red commensal polynoid polychaete Branchipolynoe seepensis is commonly found in the pallial cavity of the vent mussel Bathymodiolus azoricus, the dominant bivalve species along the Mid-Atlantic-Ridge (MAR) and is known to be kleptoparasitic. Mussels were collected from three hydrothermal vent fields in the MAR: Menez Gwen (850 m depth, MG2, MG3 and MG4), Lucky Strike (1700 m depth, Montségur-MS and Eiffel Tower-ET) and Rainbow (2300 m depth). Polychaetes were absent in all Menez Gwen vent mussels, while the highest percentage was detected in mussels from Lucky Strike, where more than 70% of the mussels had at least one polychaete in their mantle cavity, followed by Rainbow with 33% of mussels with polychaetes. Total metal concentrations (Ag, Cd, Co, Cu, Fe, Mn, Ni and Zn) were determined in polychaetes whole body and in the mussel tissues (gills, digestive gland and mantle). To understand the possible metal interactions between symbiont and host, the activity of antioxidant defence (catalase (CAT), metallothioneins (MTs)), biotransformation enzymes (glutathione-s-transferases (GST)) activities and lipid peroxidation (LPO) were determined in polychaete whole soft tissues and in mussel tissues (gills, digestive gland and mantle). Metal concentrations in polychaetes and mussels tissues indicated that the accumulation patterns were species specific and also influenced by, and possibly dependent upon, the inter- and intra-variation of vent physico-chemistry between hydrothermal fields. Despite not detecting any strong correlations between metal and enzymes activities in polychaetes and mussels, when in presence of polychaetes, mussels presented less metal concentrations in the gills and digestive gland and lower activity of enzymatic biomarkers. This leads to infer that the polychaete plays a role on the detoxification process, and the interaction between the polychaete mussel association is probably an adaptation to metals concentrations at the vent sites

Antineoplastic drugs used in chemotherapy were detected in aquatic environment: despite the very low concentrations (ng L−1 to ug L−1), due to their potent mechanism of action they could have adverse effects on non-target aquatic organisms particularly under chronic exposure. Cisplatin (CDDP) is one of the most effective anticancer drug currently in use but information on its ecotoxicological effects is very limited. In this study, Mytilus galloprovincialis was used to investigate the toxic effects related to CDDP exposure. Mussels were exposed to cisplatin (100 ng L−1) for 14 days: antioxidant (superoxide dismutase, catalase, total and selenium-dependent glutathione peroxidase) and phase II (glutathione-S-transferase) enzymes activities, oxidative damage (lipid peroxidation), genotoxicity (DNA damage) and neurotoxicity (acetylcholinesterase) was evaluated. Results indicate that CDDP at tested concentration induce changes in the antioxidant capacity, oxidative stress in target organs (digestive gland and gills) as well as DNA damage in mussel hemocytes and neurotoxicity representing a risk for non-target organisms.

Hydrothermal vents are extreme environments rich in biodiversity. The polychaete Branchipolymoe seepensis subsists in the palleal cavity of the mussel vent Bathymodiolus azoricus. These scale-worms absorb metal particles deposited in mussel gills surface and their presence has been associated with local environmental characteristics. The goal of this work was to establish a host-symbiont relationship between these two species using oxidative stress as an endpoint in response to metal accumulation.

Hydrothermal vent biology has been studied in an attempt to understand the mechanisms involved in vent organisms that permit their survival and reproduction in extreme conditions. Symbiosis is very important for the subsistence of these deep-sea organisms. The scale-worm Branchipolymoe seepensis lives in the palleal cavity of the mussel vent Bathymodiolus azoricus, the dominant species in the MAR. Polychaetes are known to absorb particles in excess deposited in mussel gills and at the same time get protection. The aim of this study was to understand the metal accumulation in this symbiosis relationship. Mussels B. azoricus were collected from two sites from Lucky Strike Hydrothermal vent (1700m), Montségur-MS and Eiffel Tower-ET and from Rainbow-Rb hydrothermal vent (2300m) where B. seepensis is present. To compare the interaction between both species, mussels were also collected from Menez Gwen hydrothermal vent (MG2, MG3 and MG4) (850m) where polychaetes are not present. Induction of metallothionein (MT) and lipid peroxidation (LPO) levels were higher in polychaetes from Rb and MS vent sites, while in mussels were higher in the gills followed by the digestive gland and mantle in all vent fields. Enzymatic activities (CAT and GST) showed that mussels had higher CAT activity in digestive gland, followed by gills and mantle, while GST levels were higher in gills. In polychaetes , both CAT and GST activities were higher in ET. Metal concentrations (Ag, Cd, Co, Cu, Fe, Mn and Zn) in the cellular and subcellular fractions determined in mussels tissues and polychaetes whole body by ICP-OES suggest intra-and inter-variation of fluid chemistry between hydrothermal vent sites. The subcellular metal partitioning in polychaetes was higher in insoluble and soluble (thermolabile and heat-stable) fractions from ET, MS and Rb, respectively. In mussel tissues, me tal partitioning was higher in gills and digestive gland. Overall, the data suggest that the presence of the scale-worm

Lucky Strike hydrothermal vent field is the largest known vent in the Mid-Atlantic Ridge (MAR), characterized by fluids that reach temperatures of 330ºC at 1,700m deep and by high amounts of metals (e.g. Cd, Cu, Fe, Hg, Mn and Zn). The vent polychaete Branchipolynoe seepensis lives in the palleal cavity of the mussel dominant species Bathymodiolus azoricus, cleaning the gills surface from particles in excess. This scale-worm is found more frequently inside the host mussel compared to other scale-worms, probably related with local environmental characteristics. Polychaetes are a very good trace-metal bioindicators, accumulating metals directly related with local metal bioavailability. The aim of this study was to understand the changes in protein expression of the membranar subproteome of polychaetes B. seepensis in response to metal concentration characteristics of Lucky Strike vent sites. Mussels were collected from two sites from Lucky Strike Hydrothermal vent field, Montségur-MS and Eiffel Tower-ET, during BIOBaz mission in August 2013 and polychaetes separated from the mussels. Subcellular fractionation was performed by sequential centrifugation of polychaetes tissues and 2D-SDS-PAGE applied to the membranar subproteome. Metal concentrations were also determined in polychaetes cellular and subcellular fractions by ICP-OES. Protein expression signatures show significant differences in 471 protein spots for B. seepensis between the two vent sites, with 11 specific for Montségur and 382 specific for Eiffel Tower. Common proteins to both sites with the higher change in expression (2-fold or higher) were selected to be identified by MALDI-TOF. Metal concentrations in the cellular (insoluble) and subcellular (thermo labile and heat-stable compounds) fractions were higher in ET and MS, respectively. Data suggests that metal concentrations from both sites reflect the environmental fluctuations between the two microhabitats within the same vent field and are related with

Lucky Strike is the biggest hydrothermal vent field located in the Mid-Atlantic Ridge (MAR) at 1, 700m de pth, characterized by fluid chemistry with high metal concentrations (e.g. Cu, Fe, Mn and Zn), low pH and temperatures of 330ºC. The polychaete Branchipolynoe seepensis live in the palleal cavity of the mussel dominant species Bathymodiolus azoricus. B.seepensis is known by clean the mussel gills surface from particles in excess and its presence is above 85% in Lucky Strike vent field. The aim of this study was to understand, at a membranar subproteome level, if the presence of B.seepensis in vent mussel B.azoricus is a response to high metal concentrations in Lucky Strike vent sites (Montsegur-MS and Eiffel Tower-ET). Mussels and polychaetes were collected from Lucky Strike hydrothermal vent field (MS and ET), in August 2013, aboard the vessel “Pourquoi Pas?”. Subcellular fractionation of polychaetes and mussel gills was performed and 2D-SDS-PAGE applied to the membranar subproteome. Metal concentrations (e.g. Ag, Cd, Cu, Fe and Zn) of both species were also determined in cellular and subcellular fractions by ICP-OES. Protein expression signatures showed significant differences, suggesting intra variation in Lucky Strike field. For B. seepensis 471 different proteins were detected between the two vent sites, 11 specific for MS and 382 specific for ET and 71 and 67 specific proteins for B.azoricus for MS and ET, respectively, out of a total of 176. Common proteins to both sites with the higher change in expression (2-fold or higher) were identified by MALDI-TOF. Metal concentrations in the cellular (insoluble) and subcellular (heat-stable compounds) fractions were higher for both species in ET and MS, respectively. Data suggests that the different metal concentrations found in these species from both sites reflect the environmental chemical fluctuation between the two microhabitats within the same vent field and are related with the changes at the subproteome le

Bioindicators can reflect the effects of pollutants on their metabolism, being widely used to assess environmental stress. In this sense, the crab Procambarus clarkii has been previously proposed to monitor the contamination in Doñana National Park (southwest Spain) using conventional biomarkers. In this work, a metabolomic approach based on direct infusion mass spectrometry, which allows an easy and quick study of a large number of metabolites in a single run, was used for pollution assessment of this area, considering the biological response of this organism to contamination. In addition, metal accumulation in crab tissues was determined. Thus, the integrated analysis of metabolomic and metallomic data enabled the study of metabolic response of the organism against pollution. Several metabolites were discovered as potential biomarkers of pollution, such as decreased levels of carnosine, alanine, niacinamide, acetoacetate, pantothenic acid, ascorbate, glucose-6-phosphate, arginine, glucose, lactate, phospholipids, and tryglicerides, as well as elevated levels of acetyl carnitine, phosphocholine, choline, and uric acid. In this way, metal-induced toxicity could be related to metabolic impairments, principally oxidative stress, metabolic dysfunction, and dyslipidemia.

The coastal ecosystems are impacted as consequence of human activities. The inputs of contaminants can affect to marine organisms and reduce the biodiversity. Within the input of contaminants to coasta l areas, pharmaceuticals have provoked social concern. In fact, some of them have been proposed to be included in the WFD of the EU. Nevertheless, the information about the levels of pharmaceuticals in coastal ecosystem is lacking. To know, the occurrence of pharmaceuticals in coastal areas of the southwest of the Iberian Peninsula, a survey was carried out in the superficial sediments of ten sampling sites. For many pharmaceuticals their concentrations were below the than limit and the pharmaceuticals occu rrence was different depending on coastal areas. A preliminary assessing of environmental risk was carried out.

Aquatic environments in the south of Portugal are characterized by estuaries, lagoons and saltmarshes that have their environmental situation critically dependent on the impact of contaminants as consequence of mining activities, increase of population densities, agricultural and industrial activities. The evaluation of contaminant impacts on marine organisms and consequent coastal water quality is crucial and requires efficient analytical methodologies combined with new management tools. In order to assess environmental risk and deterioration in these ecosystems analytical tools were developed under the framework of Interreg IIIA project Rise to characterize and diagnostic the presence of contaminants. A biomonitoring survey was carried out in three different ecosystems (Arade river, Ria Formosa lagoon and Guadiana river) to assess the quality of water, sediment and biota (clams Scrobicularia plana and polychaetes Nereis diversicolor). This involved analyses of: a) water: abiotic parameters (temperature, salinity, pH, and dissolved oxygen); b) sediments: metals (Cd, Cu, Ni, Zn, Cr, Hg, As, Fe, Mn, Co, Li, Al and Pb); c) biota: condition index, metals (Cd, Cu, Ni, Zn, Cr, Fe, Mn, Co, Al, As and Pb), and a battery of biomarkers (lipid peroxidation, antioxidant enzymes (CAT, SOD and GPX), metallothionein and alkali-labile phosphates). Biological answers were integrated using the Integrated Biomarker Response (IBR) and compared with metal levels in sediments and organism tissues. The IBR applied in this biomonitoring survey allowed a discrimination of stress levels among sites and species related differences, and can be a useful tool in environmental quality assessment studies.

The present study considers potential environmental problems in Doñana National Park using bioindicators that reflect the biological response to the presence of contaminants. Invertebrates have been frequently proposed for this aim [1]. The red swamp crayf ish Procambarus clarkii lives in a wide range of environmental conditions, even in highly polluted waters and it was introduced into the marshes of the lower Guadalquivir River (SW, Spain) in 1974, colonizing quickly all the aquatic systems around in the a bsence of natural predators, reaching the Doñana National Park. Therefore, the crab can be used as bioindicator to measure the effects of pollutants in this important ecological reserve. In this context, the omics techniques are suitable tools to evaluate the global biological response to contamination. Metabolomics studies the complete set of low molecular weight compounds in living organisms, which may be considered as the final product of interactions between gene and protein expression and the cellular environment. Thus, metabolomics aims to determining a sample’s profile at a specified time under specific environmental conditions for a better compression of biological systems [2]. Metallomics is focused on the study of metal -­‐ species related to cells, or gans and tissues, using multidimensional analytical approaches in which inductively coupled plasma mass spectrometry plays a fundamental role. Due to the importance of metals in biological systems, metallomics provides complementary information about the b iochemical changes associated with pollution [3]. In this study, crab specimens were sampled in different areas of Doñana National Park, for the dissection of digestive glands, gills and abdominal muscles, extraction and analysis. To assess heavy metal co ntamination, total metal content was performed, since it is one of the most important pollutants in marine origin organisms in this area. In addition, to evaluate the effects of these pollutants in th

[EN] The environmental quality of the southwest Iberian coast was assessed in different areas (Ria Formosa Lagoon, Guadiana Estuary and Cádiz Bay) focusing on metal contamination (Cd, Cu, Ni, Pb and Zn) in Nereis diversicolor whole tissues. In addition, associated toxicological effects were assessed using a multibiomarker approach combining several conventional biomarkers. The set of biomarkers selected included antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase), metallothionein and lipid peroxidation. N. diversicolor from the Ria Formosa Lagoon, Guadiana River and Cádiz Bay responded differently to metal contamination with different biomarker responses showing a clear site trend, suggesting different sources and/or magnitudes of contamination. Cadmium was a source of oxidative stress in polychaetes, mainly in Cádiz Bay, with a significant influence on antioxidant enzymes and enhancement of lipid peroxidation. The highest MT concentrations were in the Ria Formosa Lagoon and in the Guadiana River, where there was a direct relationship with high nickel concentrations. Biomarker responses of N. diversicolor are useful tools for environmental quality assessment on the southwest Iberian coast, and more specific metal biomarkers should be included in future assessments., [ES] B iomarcadores en N ereis diversicolor (P olychaeta : n ereididae ) como herramientas de gestión de la evaluación del medio am B iente en el suroeste de la P enínsula i B érica . – El estudio de la calidad ambiental de los ecosistemas costeros del SO de la Península Ibérica (Ría Formosa, estuario del Guadiana y bahía de Cádiz) en relación a la contaminación metálica (Cd, Cu, Ni, Pb y Zn) se ha llevado a cabo mediante el análisis del contenido de metales en Nereis diversicolor . Además, los efectos ecotoxicológicos fueron evaluados mediante una aproximación “multibiomarcadores”, en donde se combinan varios biomarcadores convencionales. La batería de biomarcadores seleccionadas han sido: las enzimas antioxidantes (catalasa (CAT), superóxido dismutasa (SOD) y glutatión peroxidasa (GPX), metalotioneína (MT) y peroxidación lipídica (LPO). Los ejemplares de N. diversicolor recolectados en las distintas zonas, mostraron una respuesta diferente a la contaminación metálica, lo que sugiere diferentes fuentes y/o magnitud de la contaminación. Los niveles de cadmio estan directamente relacionados con las actividades SOD y CAT y los niveles de LPO. En el caso del níquel, sus niveles se hallaban correlacionados positivamente con los de MT en la Ría Formosa y en el río Guadiana. La integración de las respuesta de los biomarcadores en N. diversicolor ha demostrado ser una herramienta útil para la evaluación de la calidad ambiental de los ecosistemas costeros del SO de la Península Ibérica.

The assessment of environmental issues is generally performed by the use of bioindicators that provide the biological response to contaminants. Mammals and invertebrates have been frequently proposed for this aim, and the study of conventional biomarkers supplied a deeper knowledge about the toxicological mechanisms. The great number of substances and metabolites involved in these complex mechanisms has driven the use of omics, which are the new tool to evaluate the global biological response to contamination. Metallomics is the most recent omic focused in the study of metal-binding biomolecules using instrumental couplings, generally based on the use of SEC-ICP-MS and organic mass spectrometry. In the present study we apply this approach to compare the biological response of Scrobicularia plana collected from polluted and non-polluted areas from the south-west of Spain and Portugal. Digestive gland of the clam was extracted to obtain the cytosolic fraction and the chromatographic profiles of the extracts reveal the presence of low molecular mass Cu and Zn molecules about 300 Da that can be related to the Cu- and Zn-glutathiones and As metabolites such as arsenobetaine and arsenocholine. The up-regulation of Cu-biomolecules is higher in Cadiz area against Portugal zone while Zn-biomolecules present an opposite behaviour. In several samples has been observed the presence of a remarkable Cu-peak with molecular mass of about 67 kDa. Metal biomolecules up- or down-regulated under the action of contaminants are being identified.

The southwest of the Iberian Peninsula is characterized by the presence of diverse aquatic environments, estuaries and marshes which environmental situation depends critically on the impact of pollutant substances as consequence of mining activities, increase of population densities, agricultural and industrial activities . A great number of contaminants are treated and eliminated in the waste water treatment plants, but recently the presence of new emergent pollutants has been detected which chemical nature limits the action of the treatment systems . In order to assess environmental risk and deterioration in these ecosystems analytical tools were developed under the framewor k of Interreg IIIA project Rise sub projects Endocrinbiotox and Farmacotox to characterize and diagnostic the presence of emergent pollutants . Samplings were carried in marshes and aquatic ecosystems of the cross- border region of Spain and Portugal, namely Bay of Cádiz, River Guadiana, Ria Formosa Lagoon and Arade River . Core sediments and benthic organisms (clam Scrobicularia plana and polychaete Nereis diversicolor) were sampled . Accordingly, molecular biomarkers based on the characterization of antioxidant enzymes and oxidative damages in biomolecules were analyzed, as well as neurotoxicity and endocrine disruption . Biological answers were integrated with the determination of contaminants in organisms’ tissues and sediments . Additionally, a proteomic approach was applied to gather comprehensive information about protein changes in organisms’ tissues induced by distinct contaminants to identify new biomarkers to be used in environmental quality assessment . The integration of biomarker responses and sediment characterization can be a useful tool for marine pollution monitoring in southwest of the Iberian Peninsula.

Aquatic environments in the south of Portugal support several high biodiversity ecological ecosystems that have their environmental situation critically dependent on the contributions of mining activities, increase of population densities, agricultural and industrial activities. The evaluation of contaminant impacts on marine organisms and consequent quality of coastal zones is crucial and requires efficient analytical methodologies combined with new management tools. In order to assess the environmental risk of contamination in these ecosystems data obtained from several Interreg IIIA framework projects (RED CONTAMAR, PROTEOBIOMAR and RISE) was gathered and analysed. Accordingly, the environmental quality of two important aquatic ecosystems of south of Portugal, the Ria Formosa Lagoon and the Guadiana Estuary, was assessed carrying out an integrated approach combining conventional oxidative stress biomarkers (SOD, CAT, GPX, and LPO) with proteomic analysis. Clams were collected in the Ria Formosa and in the Guadiana Estuary, in two winter periods (2007 and 2010). The biomarker approach indicated site and temporal related differences. As for proteomic analysis, several differences were detected in protein expression signatures between sites and winter periods. The highest changes in protein expression profiles (PEPs) corresponded to clams from the Guadiana Estuary possibly reflecting differences between contaminant loads as well as their interactions in complex mixtures. The comparison of PEPs through time could lead to the development of new biomarkers to be used in environmental quality assessment and its integration with biomarker responses in S. plana can be a useful tool for marine pollution monitoring in these ecosystems.

Aquatic environments in the south of Portugal, like estuaries and marshes, support vital habitats for organisms and have their environmental situation critically dependent on the contributions of pollutant substances. The evaluation of contaminant impacts on marine organis ms and consequent quality of the coastal zone is crucial and requires efficient analytical methodologies combined with new tools. Mudflat sediments are the definitive sink for most of the chemicals introduced in aquatic environments by anthropogenic sources. Consequently, benthic species like the polychaete Nereis diversicolor that live in close contact with sediments are particularly exposed to chemical stress. In this study, the environmental quality of two important aquatic ecosystems of south of Portugal (Ria Formosa Lagoon and Guadiana Estuary) was assessed carrying out an integrated approach that combined conventional oxidative stress biomarkers (SOD, CAT, GPX, and LPO) with proteomic analysis. Polychaetes were collected in the Ria Formosa and in the Guadiana Estuary, in two winter periods (2007 and 2010). The biomarker approach indicated site and period related differences , showing a slight improvement of one site in the Ria Formosa and a decline in the other. The Guadiana Estuary presented a higher oxidative stress level for polychaetes although not markedly different between winters. As for proteomic analysis, several significant differences were detected in protein expression signatures between sites. The highest changes in protein expression profi les (PEPs) corresponded to spots from the Guadiana Estuary possibly reflecting differences between contaminant loads as well as their interactions in complex mixtures. The comparison of PEPs through time could lead to the development of new biomarkers to b e used in environmental quality assessment and its integration with biomarker responses in N. diversicolor can be a useful tool for marine pollution monitoring in these ecosystems

There is little information regarding pollutant levels in farmed fish, and the risks associated to consumption. This study was designed to assess levels of exposure to metals, organochlorinated compounds, polycyclic aromatic hydrocarbons (PAHs) and alkylphenols (APEs) in farmed sea bass Dicentrarchus labrax from five aquacultures located in Southern Europe. Additionally, several biochemical responses (metallothionein, 7-ethoxyresorufin O-deethylase, vitellogenin) were determined as complementary tools. The obtained data indicate that pollutants exposure in farmed fish is similar to the levels reported in wild specimens from the area. Nonetheless, some biochemical responses were observed in the studied organisms, viz. metallothionein induction in Cu exposed organisms, and 7-ethoxyresorufin O-deethylase (EROD) and vitellogenin induction in PAHs and APEs exposed ones. The study further supports the usefulness of the biomarker approach as a first screening method to discriminate between basal and high levels of exposure in cultured fish. © 2007 Elsevier Ltd. All rights reserved.