Histopathology alterations and histochemistry measurements in mussel, Mytilus edulis collected offshore from an aluminium smelter industry (Norway).

Histopathological characteristics of specific organs express condition, and represent time-integrated impacts on the organism stemming from alterations at lower levels of biological organisation. As integrative parameters, histochemical investigations have proved to be sensitive tools to detect effects of chemical compounds. The objective of this study was to determine changes in the tissues of mussels collected at a PAH contaminated site compared to a reference site using histopathological and histochemical parameters: lipofuscin (LF) accumulation in mussel digestive gland, and lysosomal membrane stability (LMS), and using additional information provided by body burden analysis to compare the sensitivity of these parameters. The histochemical measurements for both LF and LMS gave a clear indication of a high level of stress in animals from the PAH contaminated site. This LF accumulation in lysosomes is the result of peroxidation of autophagocytosed proteins associated with protein aggregates and oxidatively damaged organelles. These measurements were able to detect the effects of PAHs, and showed a strong relationship with the body burden results.

A biomonitoring study assessing the residual biological effects of pollution caused by the HAVEN wreck on marine organisms in the Ligurian Sea (Italy).

Residual biological effects of the 1991 HAVEN oil spill off the Ligurian (Arenzano) coast were assessed in this study. Samples of the fish species Boops boops, Mullus barbatus, and Uranoscupus scaber were collected from two polluted sites near the HAVEN wreck and from an uncontaminated area. In addition to this, mussels were caged along the coast affected by the HAVEN disaster. The physiological status of fish and mussels was assessed using a battery of stress and exposure biomarkers. The PAH content of mussel and fish tissues was also analyzed. Significant biological responses were observed in lysosomal membrane stability, neutral lipid and lipofuscin accumulation and micronucleus frequency for mussels caged at two sites close to the HAVEN wreck. Chemical analyses indicated, however, that these effects are not caused by aromatic hydrocarbons. For this reason, we suggest that the aftermath of the HAVEN disaster contributes very little to coastal ecosystem pollution. This was also confirmed by the few biological effects observed in fish specimens (Boops boops) collected from surface waters. Nevertheless, it is important to point out that benthic fish displayed a stress syndrome potentially caused by aromatic hydrocarbons released from the oil tanker, as witnessed by an enhanced EROD activity and increased lipofuscin and neutral lipid lysosomal contents.

An overview of the BEEP Stavanger workshop.

Within the three-year European Research Project BEEP (Biological Effects of Environmental Pollution in marine ecosystems), a workshop was carried out at the facility of Akvamiljø a.s. with the scientific support of IRIS (International Research Institute of Stavanger) researchers. Validation of newly developed biomarkers, especially at the molecular level, was the core part of the activity. Two large scale mesocosm exposures were done and samples shared between the participants (about 30 Institute), allowing an integrated approach for studying the effects of several pollutants selected as environmentally important issues (i.e. PAHs, alkyl phenols, bisphenol A, diallyl phthalate and polybrominated diphenyl ether). In addition, a field study (Visne, Norway) has been performed for the validations of biomarkers under environmental conditions. The Stavanger workshop demonstrates the importance of collaboration between researchers to establish common ways to proceed in biomarker analysis. The common exposures encourage a joint strategy for planning biomonitoring activity. A clarification in this direction is needed to coordinate results from numerous studies about the environmental impact of pollutants which are currently carried out all over Europe and around the world. Expertise in various disciplines is required to plan and perform a successful monitoring activity and to study/predict the effect of environmental pollutants. Biologists, chemists, statisticians and environmental researchers should always be included. The Stavanger workshop also indicated the importance of developing multivariate statistical methods to enable interpretation of complex data sets. It is of interest to develop statistical tools which can integrate the results from a battery of biomarkers in order to obtain a more complete picture of environmental impact.

An ecotoxicoproteomic approach (SELDI-TOF mass spectrometry) to biomarker discovery in crab exposed to pollutants under laboratory conditions.

Ciphergen ProteinChip Technology is a proteomic tool, used for the discovery of new and sensitive biomarkers. This approach was used to evaluate the protein profile of crabs exposed to various pollutants. Two different exposure experiments were performed: spider crabs (Hyas araneus) were exposed for 3 weeks to diallyl phatalate (DAP), bisphenol A (BisA) and polybrominated diphenyl ether (PBDE-47), while shore crabs (Carcinus maeanas) were exposed to crude oil, crude oil spiked with alkylphenols (APs) and 4-nonylphenol (NP). Gender and species-related protein pattern alterations were observed and compared to controls. Results showed different responses to pollutants by the two species. Major disruption in protein peak expression was observed in samples exposed to mixtures of pollutants, i.e. oil spiked with APs. Compared to shore crab, spider crab species showed a lower degree of response in terms of number of altered protein peaks following exposure. In general, female individuals of both species showed a larger number of significantly altered proteins compared to males. Data analysis by non-metric multi-dimensional scaling (MDS) was performed. Bi-dimesional-MDS plots revealed a good separation of groups for both spider and shore crabs. In some cases, a good discrimination can also be observed between the two genders within each treatment. Results highlight the potential of crabs as sentinel organisms for the aquatic environment. The results indicate that SELDI-ToF technology is a powerful tool to discover protein expression signatures for different pollutants and sex dependent responses.

Effects of copper and cadmium exposure on functional responses of hemocytes in the clam, Tapes philippinarum.

The effects of Cu and Cd on functional responses of hemocytes in Tapes philippinarum, a Veneridae clam widespread in the lagoon of Venice, were evaluated. Bivalves were exposed for 7 days to various concentrations of CuCl(2) x 2H(2)O (0, 10, 60, and 110 microg/L) and CdCl(2) x H(2)O (0, 150, 300, and 450 microg/L). Hemocytes were collected from exposed clams, and the effects of Cu and Cd on phagocytosis, Neutral Red retention capacity, superoxide dismutase, and cytochrome oxidase activities were investigated. Hemocytes from animals exposed to Cu showed a significant decrease in phagocytic activity, whereas no inhibition was observed in cells from Cd-exposed animals. Diffusion of Neutral Red into the cytosol, an index of lysosomal membrane alteration, was enhanced by high concentrations of both metals. Different results in biochemical parameters were observed. Exposure to 60 and 110 microg Cu/L caused a significant reduction in hemocyte superoxide dismutase activity, whereas no changes resulted after Cd exposure. Increased cytochrome oxidase activity was observed in hemocytes from mollusks exposed to 60 microg Cu/L and 300 microg Cd/L; no significant differences were found in cells from bivalves exposed to 110 microg Cu/L and 450 microg Cd/L. These results suggest a relationship between heavy metal exposure and alterations in functional responses of hemocytes in T. philippinarum and indicate that the type of observed effects vary with the nature and concentration of heavy metals. Moreover, the data obtained in the analyzed clam support the possibility of using it as sentinel organism in biomonitoring studies, even if used biomarkers will be further evaluated in field conditions.