Integrated use of biomarkers (superoxide dismutase, catalase and lipid peroxidation) in mussels Mytilus galloprovincialis for assessing heavy metals’ pollution in coastal areas from the Saronikos Gulf of Greece
Introduction
In recent years there is increasing interest in studies of oxidative toxicity by various pollutants in aquatic organisms (Livingstone, 1998). Aerobic organisms, in the process of evolution, have adapted to the increased concentrations of oxygen which is very important to their metabolism and growth but equally damaging to biomolecules because of its oxidative potential. The pro-oxidant/antioxidant balance in biological systems and the scavenging of reactive oxygen species (ROS) are crucial for cellular homeostasis (Livingstone, 2001, Valavanidis et al., 2006). Aquatic organisms are highly susceptible to oxidative effects from environmental pollutants, especially when they can generate or enhance the production of ROS, such as transition metals (Viarengo, 1989).
Aquatic organisms, especially marine bivalves, exhibit a variety of changes in enzymatic antioxidant defences after exposure to pollutants with oxidative potential (Regoli et al., 2002a, Regoli et al., 2002b). Prominent among these antioxidant defence system are superoxide dismutase (SOD) converting the superoxide anion into hydrogen peroxide, catalase (CAT) converting H2O2 into water and the glutathione S-transferase (GST), phase II detoxifying enzyme. All these enzyme antioxidant defences are used as biomarkers of oxidative damage (Regoli and Principato, 1995). Antioxidant defence enzymes are induced by various environmental pollutants under pro-oxidant conditions, initially their concentrations increase to counteract oxidative stress but prolonged exposure causes their depletion, leading to oxidative damage to basic biological molecules, such as lipid peroxidation, protein and DNA damage (Bebianno et al., 2005).
Metals, such as Cd, Ni, Cr, Pb and Hg are toxic in aquatic organisms mainly because of the oxidative potential whereas other metals, such as Fe, Zn, Cu, Se and Mn are essential for their metabolism but become toxic when their concentrations are excessive (Chang et al., 1996). Numerous studies investigated the bioaccumulation and oxidative damage caused by heavy metals in the gills and digestive glands of mussels and in fish (Canesi et al., 1998, Arabi and Alaeddini, 2005).
Bivalve mollucs, such as mussels and oysters are filter-feeding sedentary species prone to the accumulation of pollutants. They have been used extensively in recent years as sentinel organisms and sensitive bioindicators for pollutants associated with ROS generation and oxidative damage (Frenzilli et al., 2004). Seasonal and spatial variability of antioxidant defence enzymes, metallothionein concentrations, protein damage and other parameters in mussels have been used in numerous studies for marine monitoring pollution studies (Orbea et al., 2002, Manduzio et al., 2003). An international “Mussel Watch” was established many decades ago to monitor various pollutants in the world ocean, especially metals, PAHs, radioactive elements and other pollutants of concern (Goldberg and Bertine, 2000). Mussels, such as Mytilus galloprovincialis are used in the Mediterranean countries to evaluate contaminated coastal environments (Lionetto et al., 2001). The present “Mussel Watch” programme in Greece use mainly the mussel M. galloprovinciallis as a bioindicator of heavy metals (mainly as metallothionein content) contamination in various harbors and estuarine ecosystems which are influenced by anthropogenic activities (Kalpaxis et al., 2004, Zangrandi et al., 2005, Catsiki and Florou, 2006).
The Saronikos Gulf is a marine enclosed area near Athens and Piraeus directly influenced by the sewage outfall of the two metropolitan cities, industrial discharges (shipyards) and intensive shipping activities of the port of Piraeus and surrounding harbors. The start of the operation (1995) of the wastewater treatment plant of the city of Athens in Psyttalia was a turning point, contributing to the decreasing levels of pollutants discharged.
Marine pollution monitoring of the Saronikos Gulf started in 1986 within the framework of the MED-POL (MAP/UNEP National Monitoring Program) in 16 monitoring stations (NCMR, 1997, HCMR, 2003, Papathanasiou and Zenetos, 2005). Heavy metal levels of the Saronikos Gulf were measured by researchers of the University of Athens and the results were published (Scoullos and Pavlidou, 1997, Scoullos et al., 2007). Biomarkers such as metallothionein (MT) levels and acetylocholinesterase activity (AchE) were measured in M. galloprovincialis in relation to heavy metals, but results are very limited (Cotou et al., 2001, Tsangaris et al., 2004).
This is the first study of systematic biomonitoring for three years, using as bioindicators indigenous mussels M. galloprovincialis and as biomarkers the seasonal variations of SOD and CAT activities and LP concentrations in the gills and mantle in three polluted sites, in relation to concentrations of certain heavy metals.
Section snippets
Site description
In the first year of our project we investigated various sites in the Saronikos Gulf, taking samples and measuring trace metal concentrations in mussels. We aimed for indigenous mussels and seasonal availability for their collection. Indigenous mussels M. galloprovincialis were sampled in the period 2004–2006 in three sites of the Saronikos Gulf. Sites S1 and S2 are from the Elefsis Bay area, a highly polluted area, while site S3 is located in the Aegina island (in the middle of the Gulf),
Results and discussion
Total protein concentration in mussel’s gills and mantle was expressed as mg g−1 wet mass. Measurements were: mantle (M): 1.80–2.18 (min.–max.) mg g−1 wet mass and gills (G): 1.82–2.09 mg g−1 wet mass.
Conclusions
It is recognized in recent years that risk assessment of environmental pollution cannot be based solely on chemical analysis because does not provide a clear indication of toxic effects of pollutant on the aquatic biota (Livingstone, 2001). Although higher aquatic organisms are necessary for an unambiguous conclusion of the ecosystem health, mussels can be used as sentinel organisms for the evaluation of an early warning response to long-term ecological damage.
The present work, as far as we
Acknowledgement
This work was funded partially by the University of Athens Research Fund. We would like to thank Dr. Catsiki of the Hellenic Marine Research Centre for her help and advice concerning the collection of mussels. Also, we would like to thank Assoc. Prof. Vasilios Roussis of the Pharmacy Department of the University of Athens for his advice concerning experimental methods and the treatment of mussels.
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