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- Biomarkers in nereis diversicolor (Polychaeta: Nereididae) as management tools for environmental assessment on the southwest Iberian coastPublication . Gomes, Tânia; Gonzalez-Rey, Maria; Rodriguez-Romero, Araceli; Trombini, Chiara; Riba, Immaculada; Blasco, Julian; Bebianno, Maria JoãoThe environmental quality of the southwest Iberian coast was assessed in different areas (Ria Formosa Lagoon, Guadiana Estuary and Cadiz 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 Cadiz 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 Cadiz 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.
- Occurrence of pharmaceutical compounds and pesticides in aquatic systemsPublication . Gonzalez-Rey, Maria; Tapie, Nathalie; Le Menach, Karyn; Dévier, Marie-Hélène; BUDZINSKI, Hélène; Bebianno, Maria JoãoThis paper deals with the detection and quantification of APIs and other priority substances in the Arade River estuary (Portugal) providing the usefulness of Polar Organic Compound Integrative Samplers (POCIS). Thirteen APIs were detected whose variation was site and time dependent. Caffeine was at the highest concentration (804±209 ng/L) followed by theophylline (184±44 ng/L). Other APIs were analgesic, anticonvulsant, non-steroidal anti-inflammatory drugs, anti-lipidemic, anxiolytic and antidepressants. Twenty pesticides comprising atrazine, diuron, isoproturon, terbutryn and simazine included in the Water Framework Directive priority list were also site and time dependent. Carbendazim occurred at the highest concentration (45±18 ng/L at site 1) but atrazine, diuron, isoproturon and simazine levels were below the Environmental Quality Standards. Although the summer impact was unclear, the results highlighted POCIS suitability for profiling these contaminants. This is to our knowledge the first study concerning APIs and pesticides in this area.
- Chemical stressors in the Guadiana RiverPublication . Bebianno, Maria João; Gonzalez-Rey, Maria; Veiga-Pires, C.The Guadiana River,the fourth major river of the Iberian Peninsula,represents an important aquatic system from a socio,geo and ecological point of view for Portugal and Spain. Important economic activities such as agriculture,tourism and aquaculture are present along with several economic facilities such as harbour (commercial and fishing),marina and small shipyards for construction and repair of small vessels.All these activities generate wastes some of them toxic to the aquatic environment.
- Does non-steroidal anti-inflammatory (NSAID) ibuprofen induce antioxidant stress and endocrine disruption in mussel Mytilus galloprovincialis?Publication . Gonzalez-Rey, Maria; Bebianno, Maria JoãoIbuprofen (IBU) is one of the most sold over-the-counter non-steroidal anti-inflammatory drugs (NSAID) and widely detected in the aquatic ecosystems. Nevertheless, the information regarding IBU effects in biota is still sparse. The goal of this study was to assess IBU potential effect as oxidative stress and endocrine disruption inducer in mussel Mytilus galloprovincialis applying a battery of biomarkers. Over two weeks of exposure to IBU (250 ngL(-1)), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), phase II glutathione S-transferase (GST) activities and lipid peroxidation (LPO) levels were determined in the digestive gland and alkali-labile phosphates (ALP) were carried out in sex-differentiated mussels' gonads. The results confirm a transitory induction of antioxidant activities responses concomitant to lipid peroxide formation outline and an increase of ALP levels over time, particularly in exposed males which may lead to mussels' reproductive fitness impairment highlighting a higher impact of IBU as an endocrine disruptor than as a short-term reactive oxygen species (ROS)-generator.
- Does selective serotonin reuptake inhibitor (SSRI) fluoxetine affects mussel Mytilus galloprovincialis?Publication . Gonzalez-Rey, Maria; Bebianno, Maria JoãoFluoxetine (FLX) the active pharmaceutical ingredient (API) in Prozac(®) is a widely prescribed psychoactive drug which ubiquitous occurrence in the aquatic environment is associated to a poor removal rate in waste-water treatment plant (WWTP) systems. This API acts as a selective serotonin reuptake inhibitor (SSRI) frequently reported to cause disrupting effects in non-target species. The objective of this study includes a multibiomarker response evaluation on mussel Mytilus galloprovincialis during two weeks exposure to 75 ng L(-1) FLX assessing antioxidant enzymes activities--superoxide dismutase (SOD), catalase (CAT) and glutathione-S-transferase (GST); lipid peroxidation (LPO), acetylcholinesterase (AChE) neurotoxic response and endocrine disruption through alkali-labile phosphates (ALP) indirect measurement of vitellogenin-like proteins. Results show transient tissue-specific enzymatic responses and damage affecting mostly mussel gills. However, the clear ALP levels inhibition throughout time in both sex-differentiated gonads gives evidence to FLX reinforced action as an endocrine disruptor rather than an oxidative or neurotoxic inducer.
- 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.
- Effects of active pharmaceutical ingredients in mussel mytilus galloprovincialisPublication . Gonzalez-Rey, Maria; Bebianno, Maria JoãoMore than 3000 types of active pharmaceutical ingredients (APIs) are applied in Human and veterinary medicine practice. These compounds are considered an emergent class of environmental contaminants with the ability to cause damage and unexpected effects to aquatic organisms, namely in species of high commercial value. APIs are ubiquitous in the environment being frequently detected in influents and effluents of waste water treatment plants (WWTPs), surface waters and more distressingly in the public tap water in concentrations ranging from ng to μg.L-1. Considering these premises, the present thesis focused on APIs detection in the Arade river water, the impact of summer period in APIs’ concentration alterations applying the passive sampler device, POCIS (polar organic compound integrative sampler), as well as, the assessment of the effects caused by non-steroidal anti-inflammatory drugs (NSAID) ibuprofen (IBU) and diclofenac (DCF) and antidepressant selective serotonin reuptake inhibitor (SSRI) fluoxetine as single and mixture exposures along with a classical contaminant copper (Cu) on a non-target species, mussel Mytilus galloprovincialis. For this purpose, a multibiomarker approach was applied namely including biomarkers of oxidative stress (antioxidant enzymes activities of superoxide dismutase – SOD, catalase – CAT, glutathione reductase – GR and Phase II glutathione-S-transferase), damage - lipid peroxidation (LPO), neurotoxic effects (through the activity of acetylcholinesterase enzyme - AChE) and endocrine disruption (through vitellogenin-like proteins measurement applying the indirect method of alkali-labile phosphate - ALP) after exposure of mussel species’ to selected APIs at environmental relevant concentrations. The main results highlighted the occurrence of 19 APIs in the river Arade from several distinct therapeutic classes. Stimulant caffeine, antiasthmatic theophylline, NSAID ibuprofen and analgesic paracetamol presented the highest concentrations. Summer impact was inconclusive due to each API transient concentration in each month. The multibiomarker results revealed distinct responses towards each selected API (as single exposure or as mixtures) that were tissue and time dependent. Several multistressor interactions were proposed for each biomarker. The results also revealed APIs potential to induce oxidative stress, LPO, neurotoxicity and endocrine disruption even at extremely low concentrations on a species extremely vulnerable to APIs presence highlighting the urgency on the development of methodologies able to prevent its entrance in the aquatic environment.
- Incidence of intersex in male clams Scrobicularia plana in the Guadiana Estuary (Portugal)Publication . Gomes, Tânia; Gonzalez-Rey, Maria; Bebianno, MariaEndocrine disrupting chemicals (EDCs) known to be present in the marine environment, have great potential to influence reproductive status in aquatic organisms. A widespread incidence of intersex linked with EDCs exposure has been reported in molluscs and fish species. The species Scrobicularia plana is inherently gonochoristic; nonetheless, the feminisation of male clams was described in estuaries from the United Kingdom. Thus, this study reports the presence of oocytes in the testis of male clams of S. plana collected in the Guadiana River (Portugal). Samples were collected between June and September (2007), and gonad tissue analysed by light microscopy. Intersex with several degrees of intensity was detected during the first three months of sampling, in which the percentage of male clams affected varied between 5.9, 58.3 and 71.4%. Little is known about the causes of intersex in S. plana, being the urban, industrial and agricultural discharges possible major sources of induction. In the Guadiana River, the presence of several EDCs may be directly linked to the incidence of the intersex condition detected in Scrobicularia plana.
- Effects of active pharmaceutical ingredients mixtures in mussel Mytilus galloprovincialisPublication . Gonzalez-Rey, Maria; Mattos, J.J.; Piazza, C.E.; Bainy, A.C.D.; Bebianno, Maria JoãoActive pharmaceutical ingredients (APIs) are emergent environmental contaminants widely detected in surface waters as result of incomplete waste water treatment plant (WWTP) removal processes and improper disposal. The assessment of potential effects of APIs on non-target organisms is still scarce since besides presenting multiple chemical structures, properties and modes of action, these compounds occur as complex mixtures. This study comprises a 15-day exposure of mussels Mytilus galloprovincialis to mixtures (at environmentally relevant nominal concentrations) of non-steroidal inflammatory drugs ibuprofen (IBU) and diclofenac (DCF) (250 ng L(-1) each) and selective serotonin reuptake inhibitor (SSRI) fluoxetine (FLX) (75 ng L(-1)) (MIX 1) along with the addition of classical pro-oxidant copper (Cu) (5 μg L(-1)) (MIX 2). The goals included the assessment of oxidative stress, neurotoxic and endocrine effects on this sentinel species applying both a multibiomarker and gene expression (here and later gene expression is taken as synonym to gene transcription, although it is acknowledged that it is also affected by, e.g. translation, and mRNA and protein stability) analysis approaches. The results revealed a swifter antioxidant response in digestive glands than in gills induced by MIX 1, nevertheless the presence of Cu in MIX 2 promoted a higher lipid peroxidation (LPO) induction. Neither mixture altered acetylcholinesterase (AChE) activity, while both triggered the formation of vitellogenin-like proteins in females confirming the xenoestrogenic effect of mixtures. All these results varied with respect to those obtained in previous single exposure essays. Moreover, RT-PCR analysis revealed a catalase (CAT) and CYP4Y1 gene expression down- and upregulation, respectively, with no significant changes in mRNA levels of genes encoding superoxide dismutase (SOD) and glutathione-S-transferase (GST). Finally, this study highlights variable tissue and time-specific biomarker responses and gene expression alterations, which along with several interactions between each mixture component on each biomarker confirm the susceptibility of mussels to API mixtures.
- Differential gene transcription, biochemical responses, and cytotoxicity assessment in Pacific oyster Crassostrea gigas exposed to ibuprofenPublication . Serrano, Miguel A. S.; Gonzalez-Rey, Maria; Mattos, Jacó J.; Flores-Nunes, Fabrício; Mello, Álvaro C. P.; Zacchi, Flávia L.; Piazza, Clei E.; Siebert, Marília N.; Piazza, Rômi S.; Alvarez-Muñoz, Diana; Rodriguez-Mozaz, Sara; Barceló, Damià; Bebianno, Maria João; Gomes, Carlos H. A. M.; Melo, Cláudio M. R.; Bainy, Afonso C. D.Pharmaceuticals, such as anti-inflammatory nonsteroidal drugs, are frequently detected in aquatic ecosystems. Studies about the effects of these substances in nontarget organisms, such as bivalves, are relevant. The aim of this study was to evaluate the effects on antioxidant status caused by ibuprofen (IBU) in oysters Crassostrea gigas exposed for 1, 4, and 7 days at concentrations 1 and 100 μg L(-1). Levels of IBU in tissues of oysters, as well as cell viability of hemocytes, were measured. The transcription of cytochrome P450 genes (CYP2AU2, CYP356A1, CYP3071A1, CYP30C1), glutathione S-transferase isoforms (GST-ω-like and GST-π-like), cyclooxygenase-like (COX-like), fatty acid binding protein-like (FABP-like), caspase-like, heat shock protein-like (HSP70-like), catalase-like (CAT-like), and the activity of catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione S-transferase (GST) were also evaluated in the gills of oysters. The highest levels of IBU were observed in animals exposed to 100 μg L(-1). A significant upregulation of CYP2AU1, CYP356A1, CYP3071A1, GST-ω-like, GST-π-like, COX-like, and FABP-like was observed in oysters exposed to IBU under different experimental conditions. Oysters exposed to 1 μg L(-1) for 7 days showed a significantly higher transcription of CYP2AU2, CYP356A1, CYP3071A1, GST-ω-like, and GST-π-like but lower GR activity. In conclusion, C. gigas exposed to environmentally relevant concentrations of IBU (1 μg L(-1)) exhibited increased transcription of certain genes and alterations on antioxidant and auxiliary enzymes, which could, in the the long term, cause damages to exposed organisms.