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 β-naphthoflavone (β-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).     

Mutation analysis of ras gene in the liver of European eel (Anguilla anguilla L.) exposed to benzo[a]pyrene

Ras is regarded as one of the most important genes involved in carcinogenesis. Such genes have been characterised in several fish species and the presence of ras mutations have already been described in fish populations from hydrocarbon contaminated areas and following experimental exposure to specific contaminants. The aims of this study were to evaluate the DNA integrity by comet assay, to isolate the normal ras gene of Anguilla anguilla and analyse for the presence of ras gene mutations or changes in gene expression levels induced after one month of benzo[a]pyrene (BaP) experimental exposure. The A. anguilla ras cDNA isolated revealed a 189 amino acid protein and alignment with other vertebrate ras proteins revealed conservation of functionally important regions. Following experimental exposure to BaP, an increase in DNA damage was found by comet assay. However, no point mutations or changes in ras gene expression levels were detected when compared to control samples. In contrast to the majority of fish ras gene sequences, a high degree of polymorphic variation was detected in the A. anguilla ras gene.     

Variation of antioxidant enzyme activities of sprat (Sprattus sprattus) larvae and organic contaminant levels in mixed zooplankton from the southern North Sea.

Sprat (Sprattus sprattus) larvae and mixed zooplankton were collected from sites in the southern North Sea across three transects running north, north-west and west from the Elbe and Weser estuaries (Germany). Sprat larval sub-cellular fractions (13,500 g supernatants) were assayed for the antixoxidant enzymes catalase (EC 1.11.1.6) and superoxide dismutase (SOD; EC 1.15.1.1), and levels of polychlorobiphenyls (PCBs), p,p'-DDE and polycyclic aromatic hydrocarbons (PAHs) were determined in the mixed zooplankton (fish larval diet). Higher levels (p < 0.05) of SOD and catalase activities were observed at sites closest to estuaries corresponding to sites with the highest levels (p < 0.05) of total PCBs and p,p'-DDE. Antioxidant enzymes activities decreased in samples collected from sites further from the estuaries across a northern and north-western transect; however this was not observed across a western transect. Larval antioxidant enzyme activities are discussed in relation to potential processes affecting them including plankton contaminant level distributions.     

Induction of Hepatic Cytochrome P-450 1A in Pikey Bream (Acanthopagrus berda) Collected from Agricultural and Urban Catchments in Far North Queensland

A variety of sources of organic contaminants to the Great Barrier Reef lagoon and near-shore environment exist including boating activity, agriculture and urban run-off. Cytochrome P-450 1A activity as measured by ethoxy-resorufin O-deethylase (EROD) activity has been widely used as an indicator of the exposure of fish to organic contaminants such as polychlorinated biphenyls (PCB), polycyclic aromatic hydrocarbons (PAH) and some organochlorine pesticides. This study demonstrates the successful application of EROD measurements in a common Australian tropical estuarine fish species, Acanthopagrus berda (Pikey Bream), to identify areas under potential stress from organic contaminants. Fish were captured from four creeks draining agricultural land, a creek draining urban land and two creeks with less disturbed catchments. Significant induction of cytochrome P450-1A was observed in fish captured from Ross Creek (urban catchment, 7.4-fold) and Cromarty Creek (agricultural catchment, 6.4-fold). Increased activity was also observed in fish captured from other creeks draining agricultural land (Plantation Creek, Victoria Creek, Seymour River, 1.9–2.6-fold) as compared to those captured from creeks in undisturbed catchments (Baldy Creek, Fisher Creek, 67–114 pmol/min/mg protein).     

Hepatic ethoxyresorufin O-deethylase (EROD) activity in flounder (Platichthys flesus) from contaminant impacted estuaries of the United Kingdom: continued monitoring 1999-2001

The determination of hepatic ethoxyresorufin O-deethylase (EROD) has been used to assess the induction of the mixed function oxygenase system (MFO) of flounder (Platichthys flesus) in UK estuaries. Induction of the MFO system denotes possible exposure to certain organic contaminants (e.g. polycyclic aromatic hydrocarbons, polychlorinated biphenyls) and its measure has been incorporated in national monitoring programmes. This study presents EROD monitoring data from 5 UK estuaries taken between 1999 and 2001 and builds on data from previous years. The results reveal that for all sampled estuaries EROD values have been significantly (p < 0.05) elevated on the majority of occasions in comparison with the reference estuary, the Alde in Suffolk, UK. However, the limited temporal scale of the reported monitoring does not allow any conclusions to be drawn with respect to trends in the data. Possible factors influencing the data (size, gender, seasonality, reproductive status, etc.) are discussed and recommendations for continued monitoring are made.     

Circulation and mixing in estuary outflows

The outflow of brackish waters from the Tees estuary on an ebb tide forms a plume which is initially spread by the action of gravity. Under quiescent conditions, the rate of vertical mixing of the brackish field is relatively slow and an area of low salinity may form in the surface waters as a result of the superimposition of the outflow plumes from successive tides. A field area of some 70 km² has been observed to be enclosed by the 34.2 × 10^?3 contour.Averaged over a tidal period, the estuary outflow from the Tees forms a broad plume of low-salinity water which extends to seaward of the 20-m contour. The movement of the field thereafter depends on the residual coastal drift which runs either to the northwest or to the southeast with a typical speed of 0.015 m s^?1. The tidally averaged vertical mixing coefficient in this offshore field was estimated to be about 10^?4 m^?2 s^?1, which is similar to that off the Wear estuary and an order of magnitude smaller than that calculated for the waters off the Humber.     

Biomarkers of exposure and effect in flounder (Platichthys flesus) exposed to sediments of the Adriatic sea.

The modifications of several biomarkers were investigated in flounder (Platichthys flesus) when exposed in the laboratory to sediment samples collected from the Northern Adriatic Sea. Besides clean sand used as a control substrate, fish were exposed to sediments sampled offshore the delta of the Po River, the harbour of Trieste, and from the industrial harbour of Venice (Porto Marghera). After six days of exposure, the enzyme activities ethoxyresorufin-O-deethylase (EROD), UDP glucuronyltransferase (UDPGT), glutathione S-transferase (GST), glutathione reductase (GR) and glutathione peroxidase (GPx), were assayed in fish liver. In addition, the contents of reduced glutathione (GSH), nonprotein thiols (SH), total sugars and extractable lipids were also determined in hepatic tissue, as well as the number of micronucleated hepatocytes and biliary concentrations of fluorescent aromatic compounds (FAC). Despite some variability within treatment groups, differences among exposed organisms were evident and consistent with known contaminant levels of sampled areas. Microsomal activities (EROD, UD-PGT) and FAC levels were the most sensitive exposure indicators. Variations in the other biomarkers showed only tendencies which although not statistically significant were generally consistent with the contamination pattern.     

The distribution and relative toxic potential of organic chemicals in a PCB contaminated bay

For reliable monitoring of environmental improvement, the PCB contaminated bay Orserumsviken on Sweden's Baltic coast was investigated prior to remediation. We examined PCB congener patterns and the relative toxic potential of PCBs in extracts of three matrices: bottom sediment, settling particulate matter and the dissolved phase (obtained from semi-permeable membrane devices). Congener patterns were similar in all matrices. Three fractions were isolated from the extracts: (1) aliphatic and monocyclic aromatic compounds (MAC-fraction), (2) dicyclic aromatic compounds (DAC-fraction), including PCBs, and (3) polycyclic aromatic compounds (PAC-fraction), including PAHs. Total extracts and fractions were injected into newly fertilised rainbow trout eggs. At larval stage, hepatic EROD activities were quantified. Though high, the PCB contamination could explain only an estimated 1-5% of the total EROD induction. The order of EROD induction potential was: total extract > PAC-fraction > DAC-fraction > MAC-fraction in all matrices, suggesting that in Orserumsviken PACs made a larger contribution to the EROD induction potential than PCBs and other DACs.     

Biomarkers of contaminant exposure: results of a field study with flounder (Platichthys flesus) from the southern Baltic Sea

The results of the present study are based on enzyme biomarker measurements in flounder (Platichthys flesus), a flatfish species that is widely distributed in Baltic coastal waters. The fish were collected from known pollution gradients and from reference areas regarded as relatively free of anthropogenic input. Muscular cholinesterases (ChEs), hepatic glutathione S-transferase (GST), and hepatic ethoxyresorufin-O-deethylase (EROD) activities were measured in each sampled specimen of flounder. Generalized linear models (GLM) were used to analyze the dependence of the enzyme activity on sampling year and area as well as on the biological parameters of the fish. Statistically significant differences in the activities of the measured biomarkers were observed between reference and contaminated sites. ChEs and GST activities differed with gender. The results of this study suggest that the location and year of sampling have a significant impact on the activity of all the measured biomarkers.     

Bioaccumulation and biomarker responses of trace metals and micro-organic pollutants in mussels and fish from the Lagoon of Venice,Italy

The aims of this study were to evaluate levels and effects of trace metals (Cd, Cr, Cu, Fe, Mn, Pb, Zn), PAHs and PCBs in mussels and fish native to the Lagoon of Venice. To this end, bioaccumulation and various biomarkers of generic stress and exposure (malondialdehyde, MDA and metallothioneins, MT; ethoxyresorufin O-deethylase activity, EROD; fluorescent bile metabolites and somatic indices in fish; lysosomal membrane stability, LMS; lipofuscins, LF; neutral lipids, NL and survival-in-air, SOS in mussels) were measured in organisms seasonally collected from two differently influenced areas.In mussels, metal bioaccumulation levels at the two sites were low and fluctuated similarly, exhibiting decreasing levels in summer, like MDA and MT. Micro-organic pollutants and the probably related biological response of LMS did not show either significant site differences or seasonal trends. LF and NL revealed inconclusive patterns. The results of the survival-in-air test were quite erratic, showing that they were associated with the high variability of both natural and physiological parameters temperature, food, reproduction, and body reserve cycle.In fish, both inorganic and organic micropollutant levels were low, and differences were detected between sites only for PCBs. The pattern exhibited by micro-organic contaminants was clearly related to the reproductive cycle at both sites; a relationship with PCBs, EROD and MDA was established at the site where their levels were generally higher. Bile metabolites (3-OH benzo(a)pyrene, 1-OH pyrene) were associated with PAH contents.     

Comparison of the partitioning of trace metals in the Humber and Mersey estuaries

Studies into the partitioning of trace metals between the dissolved phase and suspended solids have been performed on two major UK estuaries, the Mersey and the Humber. Aqueous phase trace metal concentrations were determined, along with partitioning data (K_p) determined from both laboratory equilibration experiments and by chemical leaching techniques.

Dissolved trace metals in the Humber essentially exhibited conservative behaviour (or a slight positive deviation). In the Mersey, however, removal of zinc, chromium, and, in particular, copper and cadmium occurred in the low salinity region.

Sequential extraction data gave an indication of the nature of trace metal bindings to suspended sediment, and revealed total extractable trace metals on the Mersey suspended sediment to be a factor of two greater than that found in the Humber. Similarly, partition coefficients in the Mersey were approximately an order of magnitude greater than in the Humber. Chloro-complexation was shown to control the partitioning of cadmium within estuaries. Copper, however, exhibited a distinct trend of increasing K_p with increasing salinity.     

Sublethal effects of cupric ion activity on the grazing behaviour of three calanoid copepods

This research (1) characterized the effects of sublethal cupric ion activities on the grazing behavior of two estuarine copepods (Acartia tonsa, Acartia hudsonica) and one nearshore, neritic copepod (Temora longicornis) and (2) compared the sensitivity of short-term sublethal behavioral assays with that of longer-term acute toxicity tests. A nitrilotriacetate-trace-metal-ion buffer system at 27‰. S was used to quantify and control the free cupric ion activity. Acute toxicity tests were used to determine the mortality of A. tonsa and T. longicornis over 72 h within the approximate cupric ion activity range of 10⁻¹³ to 10^−9.5 M. 24 h survival was not affected within the approximate cupric ion activity range of 10⁻¹³ to 10^−9.7 M, the range used for subsequent grazing activity experiments after 24 h exposure to Cu. Grazing activity was significantly diminished at cupric ion activities of ≈ 10⁻¹⁰ M for A. tonsa and T. longicornis, and at ≈ 10⁻¹¹ M for A. hudsonica. A hormetic pattern of response in feeding activity was observed with A. tonsa and T. longicornis. Grazing activity was found to be a sensitive measure of sublethal Cu stress compared with the acute toxicity tests. Grazing activity was affected at environmentally relevant cupric ion activities.     

A multibiomarker approach in barramundi (Lates calcarifer) to measure exposure to contaminants in estuaries of tropical North Queensland

A suite of biomarkers were measured in barramundi (Lates calcarifer) from five North Queensland estuaries along a perceived pollution gradient. The biomarkers selected were 7-ethoxyresorufin-O-deethylase (EROD), cytochrome P450, fluorescent aromatic compounds (FACs), DNA integrity, RNA:DNA ratio, cholinesterase activity (ChE), condition factor and hepatosomatic index. The resulting database was subjected to uni- and multi-variate analyses in order to assess the most suitable biomarkers to assess pollution in North Queensland estuaries and to classify the environmental quality of the sites. Principal components analysis (PCA) on the biochemical markers revealed that EROD, EROD/P450, DNA damage and to a lesser extent ChE and FACs were found to be responsive to contaminants in the environment while cytochrome P450, condition factor and the hepatosomatic index were found to be less responsive biomarkers. This study has demonstrated the utility of applying a multibiomarker approach in conjunction with traditional analysis of contaminants in providing valuable information in environmental risk assessment.     

Chlorinated hydrocarbons in marine fish and shellfish of Nigeria

The concentration ranges of chlorinated hydrocarbons in marine fish were HCB (0.03–9.5 ppb), Lindane (0.02–5.3 ppb), Endosulphan (0.21–4.9 ppb), DDT (0.50–18.6 ppb), Aldrin (0.05–54.6 ppb) and PCB (4.78–225 ppb). Fish contained higher concentrations of Aldrin, Heptachlor, HCB and Lindane than shellfish, while a reverse trend was observed for DDT and PCBs. The concentrations of residues obtained were found to be lower than those reported in literature for industrialized countries. Predator fish species were found to concentrate more residues in muscle tissue than plankton feeders. The DDT/PCB values were less than 1 indicating a predominance of industrial activities over agricultural activities as the source of contamination of the marine environment. The fish Galeoides decadactylus is a potential bio-indicator for chlorinated hydrocarbons pollution monitoring in the study area.     

Petroleum and PAH contamination of the Black Sea

Concern has been expressed regarding the extent of contamination of the Black Sea. Analyses of coastal sediments taken from throughout the region indicate, however, that levels of petroleum hydrocarbons (2-300 microg g(-1) dry wt total hydrocarbons) are generally comparable to those encountered in the Mediterranean and are lower than concentrations reported for highly contaminated areas such as the Gulf, Hong Kong, Taiwan and New York Bight. Highest concentrations of total hydrocarbons (>100 microg g(-1) dry wt) were associated with discharges from Odessa, Sochi and the River Danube. Chronic/degraded petroleum was the major contributor at these sites. Samples from the Ukrainian coastline were comparatively clean (<10 microg g(-1) dry wt total hydrocarbons). Major contributions of fresh oil (as indicated by sigma n-C14-34) occur through the River Danube. Concerning total PAH, concentrations (7-638 ng g(-1) dry wt) compare to relatively unpolluted locations in the Mediterranean and are much lower than levels reported for polluted UK estuaries (e.g. Mersey, Tyne, Thames). Both pyrolytic and petrogenic PAH are present in most samples, although petroleum derived PAH are dominant at Sochi and pyrolytic sources are prevalent in the Bosphorus region. The absence of a correlation between total hydrocarbons and PAH (R2 = 0.04) indicates different primary sources for the two.     

EROD activity, serum SDH and PAH biliary metabolites in sand flathead (Platycephalus bassensis) collected in Port Phillip Bay, Australia

In order to evaluate the health status of fish inhabiting Port Phillip Bay, Australia, southern sand flathead (Platycephalus hassensis, N = 133) were collected at six stations throughout the Bay. Fish had a similar serum sorbitol dehydrogenase (SDH) activity level (p = 0.12), indicating that they were not experiencing hepatocellular injuries. Ethoxyresorufin-O-deethylase (EROD) activity was generally lower in the non-urbanized and non-industrialized southern part of the Bay. The highest EROD activity was observed in Hobson Bay, the closest station from Melbourne city. Naphthalene-type biliary metabolites were also highest in Hobson Bay with intermediate levels found in Corio Bay where refineries are present. An opposite trend was observed with the pyrene-type bile metabolites, the highest levels being observed in Corio Bay while intermediate levels were found in Hobson Bay. The ratio of naphthalene-type to benzo(a)pyrene (B(a)P)-type metabolites indicate that relatively to other sites sampled in Port Phillip Bay, Corio Bay is subjected to enriched petroleum hydrocarbons of pyrolytic origin. Temporal trends indicate that the availability of xenobiotics to fish remained unchanged over the 1990s.     

Factors affecting longitudinal dispersion in estuaries of different scale

Traditionally, the overall diluting capacity of an estuary is characterized using a coefficient of longitudinal dispersion, K_xe, which is given by the ratio of the dispersive flux of a dissolved substance to its tidally averaged longitudinal gradient. A steady-state model, which assumes a balance between the dispersive and non-dispersive fluxes and an exponential increase in estuary cross-sectional area towards the sea, has been used to derive expressions for the axial salinity distribution and the dispersive flux of salt. The model was set up assuming either a constant dispersion coefficient along the estuary or one that increased with distance towards the sea. By comparing salinity predictions with data from five UK estuaries, estimates were made of the maximum dispersive salt flux and the corresponding maximum salinity gradient for each system. The results indicated that there was an approximately linear relation between the fluxes and gradients, and the slope of a line plotted through the origin provided an estimate of a common K_xe for all five estuaries. The magnitude of K_xe was found to be about 90 m² s^–1 with a standard deviation of approximately ±32 m² s^–1. It is concluded that a representative value of 100 m² s^–1 for K_xe is a reasonable first choice when setting up a cross-sectionally averaged estuary model. The results also showed that larger systems, such as the Thames, had lower salinity gradients and lower dispersive salt fluxes, whilst smaller estuaries displayed the opposite characteristics. The model was used to predict the variation in the non-advective flux of salt along an estuary. The distribution was found to be similar to the corresponding flux distribution estimated from observations at the seaward end of the Tees estuary, despite appreciable spatial variations in the individual flux components. Allowing for a small decrease in freshwater flow, the model indicated that there was a decrease in the maximum dispersive flux between neap and spring tides. It is argued that such a reduction in flux can result in a seaward shift in the salinity distribution to a region of greater cross-section, where the freshwater transport per unit area again balances the reduced upstream dispersive flux, as found in the neap to spring response in the Tees estuary.Responsible Editor: Hans Burchard     

An integrated technique to quantify sewage,oil and PAH pollution in estuarine and coastal environments

An analytical protocol is described which allows parallel quantification of sewage, oil and PAH pollution on the same sample, thus maximizing the information gained for the effort expended. Capillary gas chromatography-flame ionization detection (GC-FID), now a routine technique in many laboratories, has been selected as the method for quantification. The protocol described is evaluated, and analyses of sediments from estuaries of the Rivers Mersey, Dee and Tamar, UK, are given as examples of how to interpret results achieved using the technique.     

Isolation and characterization of Aeromonas species from an eutrophic tropical estuary

Isolation and characterization of Aeromonas species were undertaken in the Ebrié lagoon (Abidjan, Côte d’Ivoire) over a one year period. Overall 63% of 501 water samples were identified as positive for Aeromonas spp. A. sobria, A. hydrophila and A. caviae represent respectively 49.20%, 20.63% and 30.15% of the positive samples. Thirty nine of the positive samples were recovered from surface water samples; while 24 bottom water samples were positive for Aeromonas. Aeromonas species are most frequently (82.53) present in the Ebrié lagoon urban area during the rainy and flood seasons when the salinity is low and below 10‰. A lower isolation frequency is noticed during the dry season when water salinity is over 10‰. Finally, a higher isolation frequency occurred in samples with highest counts of Escherichia coli.     

Esterase activities and lipid peroxidation levels in offshore commercial species of the NW Mediterranean Sea

There is a lack of information on monitoring neurotoxicity in offshore commercial species. To help fill this gap, we sampled hake (Merluccius merluccius) and Norway lobster (Nephrops norvegicus) in fishing grounds off the coast of l’Ametlla de Mar (NW Mediterranean) in June 2005 at a depth of 100 m and 400 m. Additionally, at 400 m depth, two other fish species, Micromesistius poutassou and Phycis blennoides were included. Neurotoxicity markers such as Colinesterases (ChEs), namely acethyl- (AChE), butyryl- (BChE), propionyl- (PrChE) and carboxilesterase (CbE) were measured in muscle. Lipid peroxidation (LP), a marker of oxidative damage, was also included. The results are discussed in relation to the animal’s sex, size and fishing depth. A comparison of esterases and LP levels between muscle and liver of hake and between muscle and hepatopancreas of Norway Lobster was made.

AChE was dominant in muscle and CbE in hepatopancreas. No differences between fish species were seen for AChE. However, N. norvegicus, presented lower levels of ChEs and LP. A size-dependence in ChEs was seen for M. merluccius, with larger animals showing significantly lower activities (p < 0.05). Sex-dependence was seen in N. norvegicus for most esterases, except AChE, with males displaying higher activities (p < 0.05). A sampling-depth effect was also seen in the crustacea, with animals from 100 m generally presenting lower esterase activities and higher LP levels.