Browsing by Author "Company, Rui"
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- 2-D difference gel electrophoresis approach to assess protein expression profiles in Bathymodiolus azoricus from Mid-Atlantic Ridge hydrothermal ventsPublication . Company, Rui; Antúnez, Oreto; Bebianno, Maria João; Cajaraville, Miren P.; Torreblanca, AmparoHydrothermal vent mussels Bathymodiolus azoricus are naturally exposed to toxic chemical species originated directly from vent chimneys. The amount of toxic elements varies significantly among vent sites along the Mid-Atlantic Ridge and B. azoricus must be able to adapt to changes in hydrothermal fluid composition, temperature and pressure. The aim of this work was to study changes in the proteome in the "gill-bacteria complex" of mussels B. azoricus from three hydrothermal vent sites with distinct environmental characteristics using 2-D Fluorescence Difference Gel Electrophoresis (2-D DIGE). Results showed that 31 proteins had different expression profiles among vent sites and both cluster and principal component analysis confirm a clear separation of mussels between sites. This suggests the existence of specific parameters grouping individuals from the same hydrothermal site. Protein spots of the more abundant differentially expressed proteins were excised, digested with trypsin and identified by mass spectrometry. All identified proteins (actin, ubiquinone, S-adenosylhomocysteine hydrolase, cysteine peptidases, chaperonin and catalase) have been related previously with oxidative stress conditions and are known to be affected by ROS inducing stressors, including metals. Results point out to specific adaptations at the proteome level of B. azoricus depending on the level of toxicants present in their environment.
- Adaptation of the antioxidant defence system in hydrothermal-vent mussels (Bathymodiolus azoricus) transplanted between two Mid-Atlantic Ridge sitesPublication . Company, Rui; Serafim, Angela; Cosson, Richard; Fiala-Medioni, Aline; Dixon, David R.; Bebianno, Maria JoãoThe vent mussel Bathymodiolus azoricus is the dominant member of the Northern Mid-Atlantic Ridge (MAR) hydrothermal megafauna, and lives in an environment characterized by temporal and spatial variations in the levels of heavy metals, methane and hydrogen sulphide, substances which are known to increase reactive oxygen species levels in the tissues of exposed organisms. To evaluate the effects of two contrasting hydrothermal environments on the antioxidant defence system of this vent mussel species, a 2-week transplant experiment was carried out involving mussels collected from the relatively deep (2300 m), and chemical rich, Rainbow vent field. These were transplanted to the shallower (1700 m), and relatively less toxic, Lucky Strike vent field. To achieve this objective, levels of superoxide dismutase, catalase (CAT), total glutathione peroxidase (GPx), selenium-dependent glutathione peroxidase and lipid peroxidation (LPO) were measured in the gills and mantle tissues of resident and transplant mussels before and after the transplant experiment. With the exception of CAT, the gills of the transplanted mussels had significantly higher antioxidant enzyme activity compared with the basal levels in the donor (Rainbow) and recipient (Lucky Strike) populations; whereas the antioxidant enzyme levels in the mantle tissues of the transplants reflected the baseline levels of activity in the native Lucky Strike mussels after 2 weeks. In contrast, LPO levels were significantly higher in both tissue types in the transplants than in either the source or the recipient populations, which suggested a response to hydrostatic pressure change (note, the transplant animals were brought to the surface for transportation between the two vent fields). The fact that the Rainbow mussels survived the transplant experience indicates that B. azoricus has a very robust constitution, which enables it to cope behaviourally, physiologically and genetically with the extreme conditions found in its naturally contaminated deep-sea environment.
- Adaptation to metal toxicity: a comparison of hydrothermal vent and coastal shrimpsPublication . Gonzalez-Rey, Maria; Serafim, Angela; Company, Rui; Bebianno, Maria JoãoRainbow vent field is one of the most metal-contaminated hydrothermal sites on the Mid-Atlantic Ridge near the Azores region. Two hydrothermal shrimp species dominate the fauna at the Rainbow site along with the mussel Bathymodiolus azoricus. Although the levels of essential and non-essential metals in these shrimps have been studied, the biological consequences of a metal-rich environment are still largely unknown. Therefore, the aim of this study was to determine the levels of metal-binding proteins - metallothioneins (MT) and the activities of antioxidant enzymes - superoxide dismutase, catalase, total glutathione peroxidase and selenium-dependent glutathione peroxidase in two hydrothermal vent shrimps (Mirocaris fortunata and Rimicaris exoculata) collected from the Rainbow site and to compare them with two coastal shrimps (Palaemon elegans and Palaemonetes varians) from a south Portugal lagoon (Ria Formosa) to evaluate their different adaptation strategies towards metals in their environment. Results show significant differences in MT levels and antioxidant enzymatic activities between vent and coastal shrimps and also between shrimp species collected from the same site. This suggests that biochemical responses in both vent and coastal shrimps are affected not only by the environmental characteristics but also by inter-specific differences. Nevertheless, these responses apparently confer successful adaptation for survival in a metal-extreme environment.
- Antioxidant biochemical responses to long-term copper exposure in Bathymodiolus azoricus from Menez-Gwen hydrothermal ventPublication . Company, Rui; Serafim, Angela; Cosson, Richard P.; Fiala-Médioni, Aline; Camus, Lionel; Colaço, Ana; Serrão-Santos, Ricardo; Bebianno, Maria JoãoCopper (Cu) is essential to various physiological processes in marine organisms. However, at high concentrations this redox-active transition metal may enhance the formation of reactive oxygen species (ROS) and subsequently initiate oxidative damage. High concentrations of Cu may increase oxidative damage to lipids, proteins and DNA. Bathymodiolus azoricus is a Mytilid bivalve very common in hydrothermal environments near the Azores Triple Junction continuously exposed to high metal concentrations, including Cu, emanating from the vent fluids. The knowledge of antioxidant defence system and other stress related biomarkers in these organisms is still scarce. The aim of this work was to study the effect of Cu (25 microg l(-1); 24 days exposure; 6 days depuration) on the antioxidant stress biomarkers in the gills and mantle of B. azoricus. The expression of stress related biomarkers was tissue-dependent and results suggest that other factors than metal exposure may influence stress biomarkers, since little variation in antioxidant enzymes activities, MT concentrations, LPO and total oxyradical scavenging capacity (TOSC) occurred in both control and Cu-exposed mussels. Moreover, there is a general tendency for these parameters to increase with time, in both control and Cu-exposed mussels, suggesting that reactive oxygen species (ROS) formation is not metal dependent, and may be related with poor physiological conditions of the animals after long periods in adverse conditions compared to those in hydrothermal environments.
- Antioxidant defence systems in the deep-sea mussel Bathymodiolus azoricus from Mid-Atlantic Ridge hydrothermal ventsPublication . Company, Rui; Bebianno, Maria JoãoA descoberta das fontes hidrotermais em 1977 foi um dos eventos oceanográficos mais importantes do século 20. Estes sistemas marinhos localizados nas cordilheiras médio-oceânicas, são caracterizados pela presença de compostos considerados tóxicos noutros ambientes, como o sulfureto de hidrogénio, metano, dióxido de carbono e em particular metais que provém dos fluidos hidrotermais e por isso são geralmente considerados um dos ambientes mais tóxicos do planeta.
- Antioxidant systems and lipid peroxidation in from Mid-Atlantic Ridge hydrothermal vent fieldsPublication . Bebianno, Maria João; Company, Rui; Serafim, Angela; Camus, L.; Cosson, R.; Fialamedoni, A.Enzymatic defenses involved in protection from oxygen radical damage were determined in gills and mantle of Bathymodiolus azoricus collected from three contrasting Mid-Atlantic Ridge (MAR) hydrothermal vent fields (Menez-Gwen, Lucky Strike and Rainbow). The activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidases (GPx) (total and Se-dependent), and levels of total oxyradical scavenging capacity (TOSC), metallothioneins (MT) and lipid peroxidation (LPO) were determined in B. azoricus tissues and the impact of metal concentrations on these antioxidant systems and lipid peroxidation assessed. SOD, CAT, TOSC, MTs and LPO levels were higher in B. azoricus gills while glutathione peroxidases (total and Se-dependent) were higher in the mantle, and with the exception of CAT, were of the same order of magnitude as in other molluscs. TOSC levels from Menez-Gwen indicate that the vent environment at this site is less stressful and the formation of ROS in mussels is effectively counteracted by the antioxidant defense system. TOSC depletion indicates an elevated ROS production in molluscs at the other two vent sites. Cytosolic SOD, GPx and LPO were more relevant at Lucky Strike (Bairro Alto) where levels of essential (Cu and Zn) and toxic metals (Cd and Ag) were highest in the organisms. CAT activity and LPO were predominant at the Rainbow vent site, where an excess of Fe in mussel tissues and in vent fluids (the highest of all three vent sites) may have contributed to increased LPO. Therefore, three distinct pathways for antioxidant enzyme systems and LPO based on environmental metal speciation of MAR vent fields are proposed for Bathymodiolus gills. At Menez-Gwen, TOSC towards peroxyl and hydroxyl radicals and peroxynitrite are predominant, while at Lucky Strike cytosolic SOD activity and GPx are the main antioxidant mechanisms. Finally at Rainbow, catalase and lipid peroxidation are dominant, suggesting that resistance of mussels to metal toxicity at these vent fields decreases in the sequence Menez-Gwen > Lucky Strike and Rainbow.
- Biomarkers of exposure to metal contamination and lipid peroxidation in the benthic fish Cathorops spixii from two estuaries in South America, BrazilPublication . Azevedo, J. S.; Serafim, M. A.; Company, Rui; Braga, E. S.; Fávaro, D. I.; Bebianno, Maria JoãoBiomarkers as lipid peroxidation, metallothionein and delta-aminolevulinic acid dehydratase were determined in Cathorops spixii to compare the biological responses of this fish from estuaries with distinct anthropogenic influence. Three areas were selected in two estuaries in accordance with the levels of contamination for the polluted (Santos/São Vicente) and with the hydrodynamic characteristics for the non-polluted (Cananéia) estuary. Water characteristics and mercury levels in C. spixii confirmed a high human influence in the polluted system. In general, the biomarkers showed differences between the estuaries, suggesting disturbances in the specific cell mechanisms due to the presence of multiple xenobiotics in the contaminated system. Therefore, these biomarkers are recommended to promote more accurate information about the exposure to pollutants. Additionally, the study of the effect of the multiple xenobiotics on resident species such as the benthic fish C. spixii can favor a better assessment of the environmental quality of these systems.
- Changes in protein expression of pacific oyster Crassostrea gigas exposed in situ to urban sewagePublication . Flores-Nunes, Fabrício; Gomes, Tânia; Company, Rui; Moraes, Roberta R. M.; Sasaki, Silvio T.; Taniguchi, Satie; Bicego, Márcia C.; Melo, Cláudio M. R.; Bainy, Afonso C. D.; Bebianno, Maria JoãoThe composition and concentration of substances in urban effluents are complex and difficult to measure. These contaminants elicit biological responses in the exposed organisms. Proteomic analysis is a powerful tool in environmental toxicology by evidencing alterations in protein expression due to exposure to contaminants and by providing a useful framework for the development of new potential biomarkers. The aim of this study was to determine changes in protein expression signatures (PES) in the digestive gland of oysters Crassostrea gigas transplanted to two farming areas (LIS and RIB) and to one area contaminated by sanitary sewage (BUC) after 14 days of exposure. This species is one of the most cultivated molluscs in the world. The identified proteins are related to the cytoskeleton (CKAP5 and ACT2), ubiquitination pathway conjugation (UBE3C), G protein-coupled receptor and signal transduction (SVEP1), and cell cycle/division (CCNB3). CKAP5 showed higher expression in oysters kept at BUC in comparison with those kept at the farming areas, while ACT2, UBE3C, SVEP1, and CCNB3 were suppressed. The results suggest that these changes might lead to DNA damage, apoptosis, and interference with the immune system in oyster C. gigas exposed to sewage and give initial information on PES of C. gigas exposed to sanitary sewage, which can subsequently be useful in the development of more sensitive tools for biomonitoring coastal areas, particularly those devoted mainly to oyster farming activities.
- Comparative petroleum hydrocarbons levels and biochemical responses in mussels from hydrothermal vents (Bathymodiolus azoricus) and coastal environments (Mytilus galloprovincialis)Publication . Serafim, Angela; Lopes, Belisandra; Company, Rui; Ferreira, A. M.; Bebianno, Maria JoãoAliphatic hydrocarbons and PAHs in the whole soft tissues of Bathymodiolus azoricus from three Mid-Atlantic Ridge hydrothermal vents (Menez-Gwen, Lucky Strike and Rainbow), and Mytilus galloprovincialis from three contaminated coastal sites in South Portugal were analysed, and its effects on the digestive gland microsomes mixed-function oxygenase system (MFO) were assessed. Aliphatic hydrocarbons levels were present in the same magnitude in both coastal and hydrothermal environments, while the UCM (unresolved complex mixture) for coastal mussels were higher than in vent mussels. In general, significantly higher PAHs concentrations were found in coastal mussels, compared to B. azoricus where low molecular weight PAHs (2-3 rings) represented the majority of PAHs contrarily to what was observed in M. galloprovincialis. The MFO components were present in both mussel species, and were detected in vent mussels for the first time. However this system seems to have different roles in species from these contrasting environments. In coastal mussels MFO responded to hydrocarbon contamination while response in hydrothermal organisms appeared to be related mainly to endogenous factors.
- Comparison of thiol subproteome of the vent mussel Bathymodiolus azoricus from different Mid-Atlantic Ridge vent sitesPublication . Company, Rui; Torreblanca, Amparo; Cajaraville, Miren; Bebianno, Maria João; Sheehan, DavidDeep-sea hydrothermal mussels Bathymodiolus azoricus live in the mixing zone where hydrothermal fluid mixes with bottom seawater, creating large gradients in the environmental conditions and are one of the most studied hydrothermal species as a model of adaptation to extreme conditions. Thiol proteins, i.e. proteins containing a thiol or sulfhydryl group (SH) play major roles in intracellular stress defense against reactive oxygen species (ROS) and are especially susceptible to oxidation. However, they are not particularly abundant, representing a small percentage of proteins in the total proteome and therefore are difficult to study by proteomic approaches. Activated thiol sepharose (ATS) was used for the rapid and quantitative selection of proteins comprising thiol- or disulfide-containing subproteomes. This study aims to isolate thiol-containing proteins from the gills of B. azoricus collected in distinct hydrothermal vents and to study the thiol-containing subproteome as a function of site-specific susceptibility to ROS. Results show that ATS is a powerful tool to isolate the thiol-containing sub-proteome and differently-expressed protein spots showed significant differences among the three vent sites, supporting previous findings that specific environmental conditions are crucial for ROS formation and that B. azoricus have different susceptibilities to oxidative stress depending on the vent site they inhabit.