Assessment of the mechanisms of detoxification of chemical compounds and antioxidant enzymes in the digestive gland of mussels, Mytilus galloprovincialis, from Mediterranean coastal sites

In this study the effects of the main marine pollutants (metals, PAHs, PCBs and DDTs) were assessed in native mussels from the Mediterranean coast of Spain. For this purpose several biomarkers such as benzo[a]pyrene hydroxylase (BPH), DT-diaphorase (DTD), glutathione S-transferase (GST), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidases (GPs), glutathione reductase (GR), metallothionein (MT) and lipid peroxidation (LPO) were measured in the digestive gland. Results showed increased LPO levels in mussels which accumulated high loads of organic compounds and arsenic in their tissues. BPH levels correlated to the concentrations of organic compounds in mussel tissues, though the range of BPH response was low in relation to the high gradient of accumulation of organic pollutants. Increased BPH levels, concomitant to low DTD and GST activities, were detected in mussels which presented high levels of organic pollutants in their tissues. This suggests that signs of LPO present in these organisms are related to the imbalance between phase I and phase II biotransformation processes. Furthermore, the increased levels of MT and CAT detected in mussels which showed high levels of Cd in their tissues appear to reflect a coordinated response which protects against the toxicity of this metal. The application of these biomarkers in environmental assessment is discussed.     

Biological effects of diethylene glycol (DEG) and produced waters (PWs) released from offshore activities: A multi-biomarker approach with the sea bass Dicentrarchus labrax

Diethylene glycol (DEG) is largely used during oil and gas exploitation by offshore platforms. The aim of this work was to investigate if this compound induces direct molecular/cellular effects in marine organisms, or indirectly modulate those of produced waters (PWs). Sea bass (Dicentrarchus labrax) were exposed to DEG dosed alone or in combination with PWs from an Adriatic platform. A wide array of analysed biomarkers included cytochrome P450-dependent enzymatic activity, bile metabolites, glutathione S-transferases, acetylcholinesterase, peroxisomal proliferation, antioxidant defences (catalase, glutathione reductase, glutathione peroxidases, glutathione), total oxyradical scavenging capacity, malondialdehyde and DNA integrity (single strand breaks and frequency of micronuclei). Results did not reveal marked effects of DEG, while PWs influenced the biotransformation system, the oxidative status and the onset of genotoxic damages. Co-exposures caused only limited differences of biomarker responses at some experimental conditions, overall suggesting a limited biological impact of DEG at levels normally deriving from offshore activities.     

Assessment of biological effects of environmental pollution along the NW Mediterranean Sea using mussels as sentinel organisms

With the aim of assessing the biological effects of pollution along three gradients of pollution in the NW Mediterranean Sea, a biomonitoring survey was implemented using a battery of biomarkers (lysosomal membrane stability, lysosomal structural changes, metallothionein (MT) induction and peroxisome proliferation) in mussels over a period of two years as part of the EU-funded BEEP project. Mussels from the most impacted zones (Fos, Genova and Barcelona harbours) showed enlarged lysosomes accompanied by reduced labilisation period of lysosomal membranes, indicating disturbed health. MT levels did not reveal significant differences between stations and were significantly correlated with gonad index, suggesting that they were influenced by gamete development. Peroxisomal acyl-CoA oxidase (AOX) activity was significantly inhibited in polluted stations possibly due to interactions among mixtures of pollutants. In conclusion, the application of a battery of effect and exposure biomarkers provided relevant data for the assessment of biological effects of environmental pollution along the NW Mediterranean Sea.     

Are native naiads more tolerant to pollution than exotic freshwater bivalve species? An hypothesis tested using physiological responses of three species transplanted to mercury contaminated sites in the Ebro River (NE, Spain)

In the lower Ebro River exist the paradoxical convergence of relatively well preserved river dynamics with the historical presence of a chloralkali plant with a long history of mercury discharges and the recent invasion of foreign bivalves species. Here we performed a comparative study on two alien bivalves, the Zebra mussel and the Asian clam (Dreissena polymorpha and Corbicula fluminea), and one protected species of naiads (Psilunio littoralis), which is the most common species of the freshwater mussel assemblages in this river. Individuals of the three species were transplanted to three sites that included a clean unpolluted upstream site, a contaminated location next to the mercury source and a downstream one. The study focused on digestive gland antioxidant and oxidative stress responses such as antioxidant enzymes, glutathione S transferase, glutathione levels, metallothionein proteins, DNA strand breaks and lipid peroxidation levels. Results evidenced interspecies differences on accumulation levels of mercury, antioxidant defensive systems and oxidative tissue damage. The naiad species, despite of accumulating more mercury showed the greatest antioxidant defensive potential, which was characterized by having high constitutive activities of glutathione S transferase and inducible activities and levels of key antioxidant enzymes and glutathione. Exposed individuals of C. fluminea had moderate levels of metal accumulation, the highest activities of antioxidant enzymes but also high levels of lipid peroxidation. D. polymorpha mussels showed the lowest levels of mercury but the lowest antioxidant responses and consequently the highest levels of oxidative injuries in the DNA and of mortality. Our results support the hypothesis that naiad species might be more tolerant to pollution than exotic species.     

First evidence of polybrominated diphenyl ether (flame retardants) effects in feral barbel from the Ebro River basin (NE, Spain)

Polybrominated diphenyl ethers (PBDEs) are chemicals of environmental concern due to their lipophilic, persistent and bioaccumulable characteristics as well as for their potential endocrine disrupting role. Former studies carried out in a tributary of the Cinca river (Ebro basin, NE Spain) revealed high levels of PBDEs in fish due to the discharges of effluents rich in PBDEs coming from a nearby industrial park in Barbastro. In this study, several biomarkers of pollutants exposure were measured in barbel, Barbus graellsii, before (upstream) and after (downstream) the main industrial site (Barbastro) in the Vero river. The results evidenced an enhanced hepatic phase I and II metabolism (measured as reductases, glutathione S transferase and uridinediphospho-glucuronosyltransferase), and of the antioxidant enzyme glutathione peroxidase. Conversely, fishes collected from downstream reaches had their phase I CYP1A dependent ethoxyresorufin O-deethylase, antioxidant diaphorase and brain cholinesterase activities depleted. In addition, the histological study of the liver and kidney of these fish evidenced an increase of the number and size of macrophage aggregates in most individuals collected downstream. Bivariate correlated analyses showed that the above mentioned biomarkers were correlated to measured PBDE congeners, thus indicating that the observed biological effects were unlikely to be related to other environmental factors than PBDEs. Overall, the measured biochemical and histological markers provide new evidence that in field exposed fish, PBDEs levels were associated with high activities of phase I and II metabolic enzymes, oxidative stress in liver, neurotoxicity in brain and histopathological effects in both liver and kidney.     

Multiple biomarkers responses in Prochilodus lineatus allowed assessing changes in the water quality of Salado River basin (Santa Fe, Argentina)

This field study assessed water quality of Salado River basin by using a set of biomarkers in the fish Prochilodus lineatus. Multiple biomarkers were measured, including morphological indexes (condition factor, liver somatic index), hematological (red and white blood cells) and biochemical (glucose, total protein and cholinesterase activity) parameters. Besides, detoxication and oxidative stress markers (antioxidant enzymes, lipid peroxidation) were measured in liver, gills and kidney. Despite water quality assessment did not show marked differences among sites, biomarkers responses indicate that fish are living under stressful environmental conditions. According to multivariate analysis glucose, glutathione S-transferase activity, lipid peroxidation levels and the count of white blood cells are key biomarkers to contribute to discrimination of sites. So, we suggest use those biomarkers in future monitoring of freshwater aquatic systems.     

Validation of oxidative stress responses in two populations of frogs from Western Ukraine

Although amphibians are considered to be good bioindicators of environmental pollution, few data are available concerning their biochemical parameters in natural populations. We investigated seasonal (spring, summer, autumn) and spatial (wetlands in rural and urban areas) fluctuations of oxidative stress biomarkers in the liver of frog Rana ridibunda in Western Ukraine. The Centroid grouping analysis demonstrated that despite the fluctuations of separate indices, frogs from an urban site in summer and autumn are differed widely from those at the same site in spring and frogs from the rural site in all three seasons, joined in common set. In summer, suppression of Mn-superoxide dismutase and catalase activity, as well as increase in oxidized glutathione and lipid peroxidation levels demonstrate a collapse of antioxidant defense system in frogs from an urban site. The integrated oxidative stress index confirms this conclusion.     

Comparative study of biochemical and immunological biomarkers in three marine bivalves exposed at a polluted site

A battery of biochemical and immunological biomarkers used for pollution assessment were measured for first time in the clams Venus verrucosa and Callista chione and were compared with those of the mussel Mytilus galloprovincialis, a well-established indicator organism utilized in numerous environmental monitoring programs. Clams and mussel were transplanted at a polluted and a reference site or maintained at the laboratory. Among biochemical biomarkers, acetylcholinesterase did not differ at the polluted site in all species, but there was a significant difference between the mussel and the clams, glutathione S-transferase showed a clear inhibition at the polluted site in all species and a significant difference between the two clams was also indicated, while catalase activities were increased only in V. verrucosa at the polluted site and not in mussel or the other clam. Immunological biomarkers responses were also pronounced at the polluted site. Lysozyme activity was species-dependent whereas respiratory burst activity measured as luminol-dependent chemiluminescence (CL) was site and stimulus dependent, and it was evident in M. galloprovincialis and V. verrucosa and not in C. chione. Further investigation focused on biochemical and immunological biomarkers related with the oxidative mechanisms in clams will strengthen and expand their use as bioindicators for pollution assessment.     

An integrated environmental approach to investigate biomarker fluctuations in the blue mussel Mytilus edulis L. in the Vilaine estuary, France

Estuarine areas represent complex and highly changing environments at the interface between freshwater and marine aquatic ecosystems. Therefore, the aquatic organisms living in estuaries have to face highly variable environmental conditions. The aim of this work was to study the influence of environmental changes from either natural or anthropogenic origins on the physiological responses of Mytilus edulis. Mussels were collected in the Vilaine estuary during early summer because this season represents a critical period of active reproduction in mussels and of increased anthropogenic inputs from agricultural and boating activities into the estuary. The physiological status of the mussel M. edulis was evaluated through measurements of a suite of biomarkers related to: oxidative stress (catalase, malondialdehyde), detoxication (benzopyrene hydroxylase, carboxylesterase), neurotoxicity (acetylcholinesterase), reproductive cycle (vitelline, condition index, maturation stages), immunotoxicity (hemocyte concentration, granulocyte percentage, phagocytosis, reactive oxygen species production, oxidative burst), and general physiological stress (lysosomal stability). A selection of relevant organic contaminant (pesticides, (polycyclic aromatic hydrocarbons, polychlorobiphenyls) was measured as well as environmental parameters (water temperature, salinity, total suspended solids, turbidity, chlorophyll a, pheopigments) and mussel phycotoxin contamination. Two locations differently exposed to the plume of the Vilaine River were compared. Both temporal and inter-site variations of these biomarkers were studied. Our results show that reproduction cycle and environmental parameters such as temperature, organic ontaminants, and algal blooms could strongly influence the biomarker responses. These observations highlight the necessity to conduct integrated environmental approaches in order to better understand the causes of biomarker variations.     

Multi-biomarker responses in green mussels exposed to PFCs: effects at molecular,cellular, and physiological levels

Perfluorinated chemicals (PFCs) are extremely persistent and have been found extensively in the environment and wildlife. Oceans are the final sink for many persistent organic pollutants (POPs) including PFCs. However, to date, there has been a lack of studies that investigated the environmental consequences of PFCs on marine organisms. To fill in this gap, environmental toxicity of two dominant PFCs, perfluorooctane sulfonate (PFOS) and perfluorooctanoate (PFOA), was examined in a sentinel species, green mussel Perna viridis, using a series of biomarkers corresponding to different biological levels (molecular, cellular, and physiological). Correlations among these biomarkers were also investigated. The results showed that the tested compounds can induce a series adverse effect at different biological levels, including oxidative stress, DNA damage, membrane instability, suppressed filtration rate, and reduced body weight. Correlation analysis revealed that excess production of reactive oxygen species could be the major toxic pathway. An indirect mode of toxic action was also explored where adverse impacts could be secondary effects of PFC exposure. The joint analysis of biomarkers from multiple biological levels resulted in a comprehensive understanding of how PFC exposure can influence the health of organisms. The correlations of these biomarkers also provided a new perspective of the ecological consequences of PFCs.     

Can Hediste diversicolor (Nereidae, Polychaete) be considered a good candidate in evaluating PAH contamination? A multimarker approach

The aim of this study was to develop a preliminary characterization of the biological responses of Hediste diversicolor to polycyclic aromatic hydrocarbons (PAHs) under controlled laboratory conditions. In order to test the effects of PAH exposure, a battery of biomarkers was applied to the polychaetes by exposing them to sublethal concentrations of benzo[a]pyrene (0.1 and 0.5 mg L⁻¹) for 10 d under laboratory conditions. The battery of biomarkers tested included oxidative stress biomarkers (glutathione content, enzymatic activities of catalase, glutathione S-transferases, glutathione reductase, glutathione peroxidases), total oxyradical scavenging capacity (TOSC) toward peroxyl and hydroxyl radicals and activity of acyl CoA oxidase (AOX) as a marker of peroxisome proliferation measured in the entire body; lipofuscin and neutral lipid accumulations and levels of Ca²⁺-ATPase activity analyzed in the intestinal epithelium; lysosomal membrane stability and genotoxic effects measured as DNA strand breaks and frequency of micronuclei in coelomocytes. Chemical analyses were also carried out to verify the polychaete’s benzo[a]pyrene (B[a]P) bioaccumulation levels after the exposure period. The results obtained indicate that B[a]P caused significant changes in most of the parameters measured in H. diversicolor. Biological responses to the organic compound were particularly significant for the biomarkers measured in the intestinal epithelium and in coelomocytes, emphasizing that these tissues were more affected during our experimental conditions. Considering the key trophic role of this benthic species in estuarine and coastal ecosystems, this study confirmed that H. diversicolor is an appropriate bioindicator of organic contamination.     

Sub-chronic exposure to fluoxetine in juvenile oysters (<Emphasis Type="Italic">Crassostrea gigas</Emphasis>): uptake and biological effects

The bioconcentration potential of fluoxetine (FLX) and its biological effects were investigated in juvenile Pacific oyster exposed for 28 days to environmentally relevant concentrations of FLX (1 ng L^?1, 100 ng L^?1 and up to 10 μg L^?1). FLX bioaccumulated in oyster flesh resulting in 28-day bioconcentration factors greater than 2,000 and 10,000 by referring to wet and dry weights, respectively. Nevertheless, FLX did not induce oyster mortality, delayed gametogenesis, or lead to adverse histopathological alterations. At the two highest concentrations, despite non-optimal trophic conditions, FLX stimulated shell growth but only in a transient manner, suggesting a role of serotonin in the regulation of feeding and metabolism in bivalves. Those high concentrations seemed to drive bell-shaped responses of catalase and glutathione S-transferase activities throughout the exposure period, which may indicate the activation of antioxidant enzyme synthesis and then an enhanced catabolic rate or direct inhibition of those enzymes. However, no clear oxidative stress was detected because no strong differences in thiobarbituric acid-reactive substance (TBARS) content (i.e. lipid peroxidation) were observed between oyster groups, suggesting that cellular defence mechanisms were effective. These results demonstrate the importance of considering additional biomarkers of oxidative stress to obtain a comprehensive overview of the FLX-induced changes in marine bivalves exposed under realistic conditions. Considering the battery of biomarkers used, FLX appears to induce little or no effects on oyster physiology even at a concentration of 10 μg L^?1. These results do not confirm the lowest observed effect concentration (LOEC) values reported by some authors in other mollusc species.     

Mussel transplantation and biomarkers as useful tools for assessing water quality in the NW Mediterranean

Mussels, coming from an aquaculture farm located in a clean open bay, were transplanted to several stations of the bays of Nice and Cannes (NW Mediterranean) including a reference site for one month at three periods. Several biomarkers: activities of glutathione S-transferase (GST; exposure to organics), of catalase (exposure to oxidative stress) and of acetylcholinesterase (inhibited by some pesticides) and the lipid peroxidation (thiobarbituric acid reactive substances: TBARS) were measured in transplanted mussels. Cd, Cu and Zn concentrations were also measured as well as their condition index. The results demonstrated some seasonal variations in GST and catalase activities with higher levels in June compared to October. The condition index was also higher in June than in October. Principal component analyses performed with the whole set of data allowed to separate stations or groups of stations according to their responses. The mussels from the harbour of Nice were characterized by high TBARS levels and catalase activity in October 1999 whereas in the harbour of Cannes, animals presented very high copper concentrations and GST activities in June 2000. At the reference site, mussels generally presented low enzymatic activities (except AChE activity) and peroxidation levels and low heavy metal concentrations.     

Seasonal variability of antioxidant biomarkers and energy reserves in the freshwater gammarid Gammarus roeseli

In gammarids, behavioural and biochemical biomarkers are commonly used in ecotoxicological studies. In our study, we have investigated seasonal variations of several biochemical biomarkers in Gammarus roeseli, a freshwater species. Organisms were sampled monthly over a 1-year period. Gender was distinguished to measure antioxidant enzyme activities like total glutathione peroxidase (GPxtot), selenium-dependent glutathione peroxidase (SeGPx) and catalase enzymes, lipoperoxidation end-product (malondialdehyde, MDA), and energy reserves with protein and lipid contents. In the same time, usual water physico-chemical parameters were measured at the sampling site. A based-gender difference was observed for parameters related to oxidative stress. Females showed higher antioxidant enzyme activities and lower MDA level than males. Parameters related to oxidative stress and energy reserves appeared correlated with temperature and physiological status of organisms. Females GPx activities were lower in autumn and winter when no breeding occurred. In both gender, MDA levels were correlated with temperature with an increase of lipoperoxidation in summer. Lipid contents were the lowest in summer and the highest in winter, probably due to the reproductive status of organisms and their feeding behaviour. Gender-based differences of biochemical parameters suggest a specific sensitivity of males and females in ecotoxicological experiments. Moreover, organisms could be more vulnerable in summer when MDA levels are high and energy reserves low. Deleterious effect of xenobiotics would be different with gender and season.     

Assessment of biological effects of environmental pollution along the NW Mediterranean Sea using red mullets as sentinel organisms

A biomonitoring program was carried out in spring and autumn in three pollution hot-spots and sensitive areas of the NW Mediterranean Sea using red mullets (Mullus barbatus) as sentinel organisms and a battery of biomarkers together with gonad histology. In fish from anthropogenic impacted areas (Fos-sur-mer, Cortiou, Arenzano, Delta of Ebro) lysosomal membrane destabilization occurred indicating disturbed health. There were no significant differences in metallothionein (MT) levels among stations. Peroxisomal acyl-CoA oxidase (AOX) activity was highest in fish from Cortiou. Both MT levels and AOX activities were significantly correlated with gamete development. Prevalence of melanomacrophage centers were high in Cortiou in all samplings and in Fos-sur-mer in September samplings. In conclusion, the application of a battery of biomarkers in red mullets provided relevant data for the assessment of environmental pollution in the NW Mediterranean Sea but also showed the difficulties of using native fish as sentinels. For future studies caging strategies are recommended.     

Transport and acclimation conditions for the use of an estuarine fish (Pomatoschistus microps) in ecotoxicity bioassays: effects on enzymatic biomarkers

Acclimation of organisms for ecotoxicity testing is in general processed according to Organisation for Economic Co-operation and Development (OECD) and/or Environmental Protection Agency (EPA) guidelines, under controlled conditions. However, when organisms are collected in the field, their capture, transport and adaptation to laboratory conditions are factors of stress. In their natural habitat, estuarine fish are exposed to considerable fluctuations of environmental variables, while in laboratory they are acclimated to constant conditions and this can be per se a factor of stress that may influence biomarker responses. Therefore, it is important to investigate the effects of these procedures on estuarine fish performance before using them as test organisms in ecotoxicity bioassays. The goal of the present study was to investigate the effects of transporting the common goby, Pomatoschistus microps from the field (natural populations) to the laboratory and of its acclimation to laboratory conditions on the enzymes acetylcholinesterase (AChE), lactate dehydrogenase (LDH) and glutathione S-transferases (GSTs). Fish were collected in a reference site of the Minho River estuary (NW of Portugal) and the activities of the biomarkers were monitored before and after the transport of organisms to the laboratory and during the acclimation period (at 5, 10 and 15 days). The activities of all the enzymes indicated that capture and transport conditions had no effects on enzymatic activities. Furthermore, AChE, LDH and GST presented higher activities at the end of the acclimation period than at beginning, suggesting a physiological adaptation to laboratory conditions. This adaptation should be taken into consideration in the experimental design to avoid bias in the interpretation of effects of xenobiotics on biomarkers.     

Lysosomal membrane stability,phagocytosis and tolerance to emersion in the mussel Perna viridis (Bivalvia: Mytilidae) following exposure to acute,sublethal, copper

The mytilid mussel Perna viridis is distributed throughout the Indo-Pacific region and is potentially a suitable candidate for biological effects (biomarker) monitoring in the subtropics. A suite of cytological and physiological responses to acute (48-72 h) copper exposures of 50-200 microgl(-1) were assessed in order to determine the suitability of P. viridis for marine pollution monitoring. Copper elicited significant destabilisation of the haemocyte lysosomal membranes and also impaired phagocytosis. Survival during emersion following exposure to copper was not related to the experimental copper exposures suggesting that higher metal concentrations may be required to interfere with anaerobic enzymes responsible for suppression of metabolism. Based on this preliminary study, cytological biomarkers evaluated in the haemocytes extracted from P. viridis should prove an effective non-destructive means of assessing metal pollution throughout the mussels subtropical range.     

Mercury in the Mediterranean,part I: spatial and temporal trends

The present paper provides an overview of mercury studies performed in the Mediterranean Sea region in the framework of several research projects funded by the European Commission and on-going national programmes carried out during the last 15 years. These studies investigated the temporal and spatial distribution of mercury species in air, in the water column and sediments, and the transport mechanisms connecting them. It was found that atmospheric concentrations of Hg compounds, particularly oxidised Hg species observed at five coastal sites in the Mediterranean Sea Basin, are significantly higher compared with those recorded at five coastal sites distributed across N Europe, most probably due to natural emissions. Hg levels in water are comparable to other oceans. Anthropogenic and natural point sources show locally limited enrichments, while natural diffusive sources influence Hg speciation over larger areas. Results and statistic comparison of mercury species concentrations within Mediterranean compartments will be presented and discussed.     

Biological detection and analysis of mercury toxicity to alfalfa (Medicago sativa) plants

Mercury has become one of the major causes of toxic metal pollution in agricultural lands. Accumulation of mercury by plants may disrupt many cellular functions and block growth and development. To assess mercury toxicity, we performed an experiment focusing on the responses of alfalfa (Medicago sativa) to Hg(2+)-induced oxidative stress. Alfalfa plants were treated with 0-40microM HgCl(2) for 7d. The concentrations of Hg(2+) were positively correlated with the generation of O2- and H(2)O(2) in leaves. Treatment with Hg(2+) increased the activities of NADH oxidase and lipoxygenase (LOX) and damaged the biomembrane lipids. To understand biochemical responses under Hg stress, activities of several antioxidant enzymes, superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) were assayed. Analysis of SOD activity by non-denaturing polyacrylamide gel electrophoresis revealed five isoforms in leaves, but they showed different patterns. Also, eight isoenzymes of APX and seven of POD in leaves were detected. However, only one isoform of CAT was visualized. The total activities of APX, POD and CAT were generally enhanced. We also measured several antioxidative metabolites such as ascorbate and glutathione (GSH), and found that both differentially accumulated in leaves. These results indicate that the increased levels of O2- and H(2)O(2) under Hg stress were closely linked to the improved capacity of antioxidant enzymes. The data not only provide the important information for better understanding of the toxic and tolerance mechanisms, but as well can be used as a bio-indicator for soil contamination by Hg.     

Chemical and biomarker responses for site-specific quality assessment of the Lake Maggiore (Northern Italy)

Since the 1990s, the Lake Maggiore (Northern Italy) has been recognized as an aquatic environment contaminated by DDTs and other persistent organic pollutants, but to date just few studies were carried out to investigate the effects of pollution to aquatic organisms. The aim of this study was the application of a stepwise approach based on chemical data, a suite of biomarkers and the integration of their responses into a biomarker response index (BRI) to evaluate the site-specific quality assessment in different sampling stations of Lake Maggiore, one of the largest European lakes. We used as biological model the freshwater bivalve Zebra mussel (Dreissena polymorpha). Several hundred bivalve specimens were sampled on May 2011 from eight sampling sites located along the lake shoreline. We measured levels of DDTs, PCBs, HCHs, HCB, and PAHs accumulated in D. polymorpha soft tissues by GC/MSn, while the activities of catalase, superoxide dismutase, glutathione peroxidase, and glutathione S-transferase, as well as the lipid peroxidation and protein carbonyl content were evaluated in homogenates from native bivalves as oxidative stress indices. Moreover, DNA damage was investigated by the alkaline precipitation assay. Significant imbalances of enzymatic activity were found in mussels from most of the sampling sites, as well as notable increases of damage to macromolecules. Health status of mussels from Baveno was greatly affected by lake pollution, probably due to high levels of DDTs measured in this site, while a wide variability in biomarker responses was found in all the other stations. The application of a BRI allowed distinguishing impacts of pollution to bivalves, confirming mussels from Baveno as the most threatened and revealing that also the health status of bivalves from Suna, Brissago, Pallanza, and Laveno is affected. These evidences suggest the usefulness of a specific index to integrate all the biomarker endpoints in order to provide a correct environmental risk assessment.