Elevation of cytochrome P450-immunopositive protein and DNA damage in mussels (Mytilus edulis) transplanted to a contaminated site.

Mytilus edulis were collected from a reference site (Port Quin) and an urban/industrial contaminated site (New Brighton) in the UK during June 1999. Levels of PCBs (sigma7 congeners) and CB-138 were determined to be, respectively, 21 fold and 16 fold higher in the mussel digestive glands from New Brighton. Levels of CYPIA-immunopositive protein were 1.5 fold higher (P < 0.05) at the polluted site but the levels of DNA strand breaks were 1.3 fold higher (P<0.05) at the reference site. Mussels from Port Quin were placed in cages at both sites and both transplanted and indigenous populations sampled in September (13 weeks). Mussels transplanted from the reference site to the industrial site, reported elevated levels of CYP1A-immunopositive protein (1.4 fold; P < 0.05) and higher levels of DNA damage (1.2 fold; P < 0.05) compared to caged populations at the reference site and a PCB loading similar to the populations from the polluted site. Moreover, transplanted mussels had DNA damage 1.8 fold greater (P < 0.05) than indigenous mussels at the transplant site. These changes were small but significant when compared to the observed temporal changes in the indigenous populations.

Seasonal variation and estradiol-dependent elevation of Thames estuary eel Anguilla anguilla plasma vitellogenin levels and comparisons with other United Kingdom estuaries.

Eel Anguilla anguilla plasma vitellogenin was investigated as a biomarker of exposure to environmental compounds with estrogenic activity, along the tidal course of the Thames Estuary, UK. A. anguilla was chosen as a sentinel species because of their wide distribution, robustness in field and laboratory studies and also because they have a characterised normal intersex' condition where the gonad contains both developing male and female gonadal cells termed a Syrski organ. Following laboratory exposure to 17beta-estradiol (intraperitoneal injection), a plasma protein (approx. 211 kDa apparent molecular weight) was detected by monoclonal antibodies to vitellogenin of striped bass (Morone saxatilis). Western and dot blot analyses were developed and vitellogenin was isolated from 17beta-estradiol-treated fish to calibrate the quantification of the blots by image analysis. The limits of sensitivity for the Western and dot blots were 100 and 10 ng vitellogenin/ml, respectively. Levels of vitellogenin in Thames estuary samples were below the detection limits of the Western but not the dot blot, and showed no statistically significant site-specific (10 sites) and seasonal-specific (May, August, November) differences. Values were observed to be low, between 11 and 165 ng/ml, compared with 17-50 mg/ml for 17beta-estradiol-treated eels. Similar low levels of plasma vitellogenin were determined in fish sampled along the Tyne, Wear, Tees or Humber estuaries, or the Weston canal Liverpool, with mean plasma vitellogenin levels varying between 44 and 82 ng/ml. These levels of vitellogenin in A. anguilla plasma were observed to be consistent with the known biology of the eel. Immature females, or fish with syrski organs, reported both lower levels and smaller variation of plasma vitellogenin concentrations whereas the highest plasma vitellogenin concentrations were determined in fish above 45 cm consistent with female fish. These results indicate inter-species variation between the plasma vitellogenin concentrations of A. anguilla and other published fish studies undertaken along the same estuaries.

Hepatic 7-ethoxyresorufin O-deethylase activity in eel (Anguilla anguilla) from the Thames Estuary and comparisons with other United Kingdom Estuaries.

Hepatic microsomal 7-ethoxyresorufin O-deethylase (EROD) activities (indicative of exposure to polycyclic aromatic hydrocarbons (PAHs) and polychlorobiphenyls (PCBs)) were measured in eel Anguilla anguilla from the Thames Estuary. Fish were collected from up to 13 sites during November 1997, May and August 1998 and October 1999. Throughout this period no clear seasonal variation could be identified at every site along the Thames. However, during the summer months, fish sampled from sites in the middle to the upper estuary (Woolwich, Greenhithe and West Thurrock) reported up to 3-fold higher EROD activities compared to sites either at the upper reaches (Richmond and Brentford) at the same time of the year, or fish sampled in winter, along the entire length of the estuary. A laboratory exposure experiment demonstrated a 3-fold elevation of EROD activity 2 days after injection with beta-naphthoflavone (beta-NF). However, higher levels of activity could be determined in fish sampled from the Weston canal near the Merseyside. The lowest levels of A. anguilla EROD activity were observed in fish sampled from the upper reaches of the River Tamar, Devon, and were comparable to activities determined in fish from the Wear and Humber estuaries. A. anguilla sampled along the Thames, Tyne and Tees estuaries reported between 2.5- and 7-fold higher EROD activities compared to fish collected from the Tamar. These results indicate that a low to moderate induction of A. anguilla CYP1A had occurred (indicative of low to moderate exposure to PAHs and planar PCBs) in fish collected from the Thames, Tyne, Wear, Tees, Humber and Tamar estuaries. However, the highest level of EROD activity was observed in fish from the Weston Canal (Merseyside).

Antioxidant biomarkers in freshwater bivalves, Unio tumidus, in response to different contamination profiles of aquatic sediments.

Antioxidant systems were studied in the freshwater bivalve Unio tumidus transplanted from a control site to four different contaminated areas, in order to study the biological response according to the contamination characteristics. Reduced and oxidized glutathione (GSH, GSSG), the activities of antioxidant enzymes such as selenium-dependent and non-selenium-dependent glutathione peroxidases (SeGPx and non-SeGPx), and glutathione reductase (GRd) were measured in the gills and the digestive gland of the mussels after 15 days of exposure at different sites. Lipid peroxidation (LPO) was evaluated by means of malondialdehyde measurements (MDA). The four sites investigated were located in the valleys of Fensch (F), Moselle (M), Lot et Garonne (LG), and Sarthe (S). At each site, the bivalves were placed upstream (Up) from an identified pollution source (a cokery, a laundry, or a foundry) and downstream (Do), close to the effluent outfall (Do(1)) or farther (Do(2)). The goal was to study the antioxidant response in relation to the pollution gradient. Metals and congeners of PAHs, PCBs, and organochlorinated pesticides were analyzed in the river sediments of each station. The exposure of the bivalves at the most highly polluted sites or close to the pollution source led to a sharp depletion in some antioxidant parameters, namely GRd, SeGPx, and GSH. The decrease in enzyme activities could reach 80% for GRd and 70% for SeGPx, while GSH depletion could yield 70%, leading then in an induction of lipid peroxidation, either in the digestive gland or in the gills. The higher the MDA concentrations, the lower the activity of these three antioxidant parameters, suggesting that they could be biomarkers for toxicity. Yet, a depletion in these parameters was sometimes insufficient for cytotoxicity to be induced, since lipid peroxidation failed to appear in some cases where antioxidant depletion was clear, although not so severe. The response of the gills and the digestive gland was not always paralleled, which can be explained by differences in the bioavailability of pollutants. In some cases, a relationship was not found between the antioxidant response and the degree and the type of contamination in sediments, suggesting that the effects could result from nonidentified pollutants or/and be indicators of bio-availability.

Development of hepatic CYP1A and blood vitellogenin in eel (Anguilla anguilla) for use as biomarkers in the Thames Estuary, UK.

The potential of eel (Anguilla anguilla) as a monitoring species for the Thames Estuary, UK, was examined. Hepatic cytochrome P4501A [7-ethoxyresorufin O-deethylase (EROD) activity] and blood vitellogenin (Western analysis) were investigated as biomarkers of exposure to, respectively, organic contaminants and to contaminants showing estrogenic activity. Hepatic microsomal EROD activities in A. anguilla from seven sites in the Thames Estuary in May 1998 varied three-fold (111 +/- 24 to 355 +/- 42 pmol min-1 mg protein-1) (mean +/- S.E.M.) and showed correlation with salinity; however, the latter relationship was not maintained at other times of the year. The range of EROD activities was two- to eight-fold higher than the 37 +/- 8 pmol min-1 mg-1 for A. anguilla from the relatively clean Tamar Estuary. beta-Naphthoflavone treatment (5 mg kg-1 wet wt.; 2 days) of Thames A. anguilla produced a two-fold increase in hepatic microsomal EROD activity. Comparing the Thames EROD data with those for A. anguilla from well-characterised contaminated sites in the Netherlands (Van der Oost, R., Goksøyr, A., Celander, M., Heida, H., & Vermeulen, N. P. E. 1996. Aquatic Toxicology, 36, 189-222), the Thames is suggested to be moderately impacted by polycyclic aromatic hydrocarbons and related contaminants. 17-beta-Estradiol treatment produced the appearance of a plasma protein of 211 Kd app. mol. wt. (recognised by antibodies to vitellogenin of Morone saxatilis), but putative vitellogenin could not be detected in A. anguilla from selected sites in the Thames Estuary.

Glutathione reductase, selenium-dependent glutathione peroxidase, glutathione levels, and lipid peroxidation in freshwater bivalves, Unio tumidus, as biomarkers of aquatic contamination in field studies.

The aim of this study was to evaluate the usefulness of antioxidant parameters in the freshwater bivalve, Unio tumidus, as biomarkers of exposure to pollutants and to study their potential interest in predicting toxicity. Selenium-dependent glutathione peroxidase (Se-GPx), non-selenium-dependent glutathione peroxidase (non-Se-GPx), glutathione reductase (GRd), catalase, and superoxide dismutase (SOD) activities; reduced (GSH) and oxidized (GSSG) glutathione levels; and lipid peroxidation were measured in the gills and digestive glands of Unio. Control mussels were encaged and transplanted for 15 and 30 days to sites where the contamination of sediments was analyzed, along a river receiving domestic and industrial sources of pollution. After 15 days of exposure, all antioxidant parameters of the bivalves transferred to the most polluted sites had strongly decreased compared with control values. This was particularly true for Se-GPx and GRd activities, which were inhibited by 60 and 80% in the two tissues, and for GSH levels (80% reduction in the gills and 60% in digestive glands). These decreases were associated in the gills with lipid peroxidation (measured by malondialdehyde content) and with a high level of contamination of sediments by polycyclic aromatic hydrocarbons and polychlorinated biphenyls. In the mussels exposed at the least polluted sites, the same parameters decreased in the gills, but to a lesser extent: 50% for Se-GPx and 32% for GRd activities, and 45% for GSH levels. The gills appeared more sensitive than the digestive glands. After 30 days of exposure, while Se-GPx, GRd, and GSH remained reduced, a significant induction of non-Se-GPx and catalase activities was recorded in the gills, which reflected an adaptation of the transplanted species to their unsafe environment. All the results indicated that antioxidant defense components, namely, Se-GPx, GRd, and GSH, are sensitive parameters that could be useful biomarkers for the evaluation of contaminated aquatic ecosystems. The relationship between the degree of deficiency of antioxidant defenses and lipid peroxidation suggests that these parameters could also be biomarkers for toxicity.