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For half a decade, the Atlantic salmon in the Baltic Sea has been facing severe health issues. Clinical signs like haemorrhage, erosions and ulcerative/necrotic skin conditions in returning adults have been reported from different Swedish rivers. These primary disease signs precede a secondary, terminal fungal infection. As initial investigations of the disease did not provide conclusive answers regarding the pathogenesis, this study was initiated to gain insight into a possible link between this so‐called Red Skin Disease and anthropogenic influences. Therefore, returning salmon were caught in rivers along the Swedish coast and different tissues were sampled. The focus was put on the measurements of a battery of biomarkers as well as biochemical and haematological parameters, which were analysed using multivariate statistics. The main findings were a severe osmotic haemodilution, an immune response and an alteration of the carbohydrate metabolism in diseased fish. Furthermore, oxidative stress does not seem to be a likely factor in the pathogenesis. Concluding, certain changes in physiological parameters were shown to be indicative for the disease patterns, while others were ruled out as significant factors. Thus, this study contributes to the understanding of the Red Skin Disease and may act as a hypothesis generator for future studies.

Estuaries are important nursery areas for many species and these habitats are often affected by anthropogenic activities. We investigate possible negative effects of pesticides and metals on penaeid shrimps in Maputo Bay, Mozambique. Shrimps and water samples were collected in three estuaries and one coastal area for biomarker and chemical analysis. Acetylcholinesterase (AChE) and glutathione-S transferase activities were analysed as biomarkers for pollutants. 37 different pesticides were analysed in water samples and shrimp muscle tissue was analysed for 10 metals. Risk assessment showed that the environmental thresholds were exceeded for several herbicides in three of four of the assessed nursery areas. Lower AChE activities were detected in shrimps captured close to an agriculture area and this location had the lowest shrimp densities. Metal analysis in shrimp showed low levels. Despite localized effects, results highlight the need to improve the regulation of pollutants in the Espirito Santo estuary in Maputo bay.

The North and Baltic Seas are heavily trafficked marine areas with extensive anthropogenic activities, including cargo and fishing vessels, waste dumping, oil platforms, industrial activities and contamination from coastal runoff. In order to evaluate the environmental health of these regions, we used the demersal fish dab (Limanda limanda) as a sentinel species. The current study used well-established biomarkers for PAH exposure and oxidative stress, measuring EROD activity, the acute antioxidant response as well as oxidation of proteins detected as protein carbonyl levels. Results show the strongest biomarker results in an area with extensive oil drilling, where dab displayed high levels of EROD activities. This was also seen in dab captured in the Baltic Sea where elevated levels of oxidized glutathione and a trend towards higher EROD activity were observed. The obtained results did, however, not indicate a coherent biomarker response. The study was conducted off shore where many areas have presumably low levels of pollutants, and we could detect minor effects using the biomarker approach.

Perch (Perca fluviatilis) has been used in biological effect monitoring in a program for integrated coastal fish monitoring at the reference site Kvädöfjärden along the Swedish east coast, which is a site characterized by no or minor local anthropogenic influences. Using a set of physiological and biochemical endpoints (i.e., biomarkers), clear time trends for “early warning” signs of impaired health were noted in the perch from this site, possibly as a result of increased baseline pollution. The data sets also showed relatively large variations among years. To identify additional temporal variation in biological parameters, global mRNA expression studies using RNA sequencing was performed. Perch collected in 2010 and 2014 were selected, as they showed variations in several biomarkers, such as the activity of the detoxification enzyme CYP1A (EROD), the plasma levels of vitellogenin, markers for oxidative stress, white blood cells count and gonad sizes. The RNA sequencing study identified approximately 4800 genes with a significantly difference in mRNA expression levels. A gene ontology enrichment analysis showed that these differentially expressed genes were involved in biological processes such as complement activation, iron ion homeostasis and cholesterol biosynthetic process. In addition, differences in immune system parameters and responses to the exposure of toxic substances have now been verified in two different biological levels (mRNA and protein) in perch collected in 2010 and 2014. Markedly higher mRNA expression of the membrane transporter (MATE) and the detoxification enzyme COMT, together with higher concentrations of bioactive naturally produced brominated compounds, such as brominated indoles and carbazoles, seem to indicate that the perch collected in 2014 had been exposed to macro- and microalga blooming to a higher degree than did perch from 2010. These results and the differential mRNA expression between the 2 years in genes related to immune and oxidative stress parameters suggest that attention must be given to algae blooming when elucidating the well-being of the perch at Kvädöfjärden and other Baltic coastal sites.

Ocean acidification and rising seawater temperature are environmental stressors resulting from the continuous increase of the atmospheric CO2 concentration due to anthropogenic activities. As a consequence, marine fish are expected to undergo conditions outside of their tolerance range, leading to physiological challenges with possible detrimental implications. Our research group has previously shown that exposure to elevated CO2 modulated the immune system of the Atlantic halibut. To further investigate this finding, we analysed non-specific immune components in blood plasma of Atlantic halibut (Hippoglossus hippoglossus) juveniles acclimated to six different temperatures (5, 10, 12, 14, 16 and 18 °C), and to water pH of 8.0 (control) or 7.6 (predicted for year 2100) for three months. Plasma ions (K+, Na+, Ca++, Cl-) and lactate concentrations were also measured. The analysis of plasma ions did not show any trends related to temperature or CO2 exposure, and the majority of the experimental fish were able to maintain ionic balance. The results show that both innate immune components (lysozyme and alternative complement system) had increased activities in response to elevated CO2, representing a CO2-related impact on the halibut's immune system. The increased activity of lysozyme and complement system is possibly part of the acclimatization process, and might be protective.

Locomotion-based behavioural endpoints have been suggested as suitable sublethal endpoints for human and environmental hazard assessment, as well as for biomonitoring applications. Larval stages of the sand goby (Pomatoschistus minutus) possess a number of attractive qualities for experimental testing that make it a promising species in behavioural ecotoxicology. Here, we present a study aimed at developing a toolkit for using the sand goby as novel species for ecotoxicological studies and using locomotion as an alternative endpoint in toxicity testing. Exposure to three contaminants (copper (Cu), di-butyl phthalate (DBP) and perfluorooctanoic acid (PFOA) was tested in the early life stages of the sand goby and the locomotion patterns of the larvae were quantified using an automatic tracking system. In a photo-motor test, sand goby larvae displayed substantially higher activity in light than in dark cycles. Furthermore, all tested compounds exerted behavioural alterations, such as hypo- and hyperactivity. Our experimental results show that sand goby larvae produce robust and quantifiable locomotive responses, which could be used within an ecotoxicological context for assessing the behavioural toxicity of environmental pollutants, with particular relevance in the Nordic region. This study thus suggests that sand goby larvae have potential as an environmentally relevant species for behavioural ecotoxicology, and as such offer an alternative to standard model species. This article belongs to the Special Issue Aquatic Organisms for Environmental Monitoring

Global climate change is predicted to result in increases in water temperature in the polar regions, but the full consequences of this for marine fish species are not understood, especially with regard to cellular mechanisms underlying oxidative stress. Warmer temperatures could potentially result in increased oxidative stress, and it is not known whether stenothermal fish can cope with this on a cellular and physiological level. In order to address this, we exposed bald notothen ( Pagothenia borchgrevinki ), a fish species endemic to Antarctica, to an increase in temperature from − 1.6 °C to 4 °C and measured the effects on oxidative stress including antioxidant defenses, oxidative damage in proteins and lipids, and transcriptional regulation of involved genes. We show that the fish responds to an acute (12 h) temperature increase with increased antioxidant defenses. However, these antioxidant defenses were similar to basal levels following long-term (3 weeks) exposure to the higher temperature and moreover, these individuals also had higher levels of oxidative damage. These results indicate that this species has the ability to alter levels of endogenous antioxidants, but that this response is transient and insufficient to protect against oxidative damage. These effects may have serious consequences for these fish in a warmer future since long-term consequences of this accumulation of damaged lipids and proteins are associated with aging and known to include decreased cellular function, disease and eventually cell death.

Chemical pollution was monitored and assessed along the Swedish west coast. 62 of 172 analyzed organic chemicals were detected in the water phase of at least one of five monitored sites. A Concentration Addition based screening-level risk assessment indicates that all sites are put at risk from chemical contamination, with total risk quotients between 2 and 9. Only at one site did none of the individual chemicals exceeded its individual environmental threshold (PNEC, EQS). The monitoring data thus demonstrate a widespread blanket of diffuse pollution, with no clear trends amongst sites. Further issues critical for the environmental chemical risk assessment include the challenges to achieve sufficiently low levels of detection especially for hormones and cybermethrin (a pyrethroid insecticide), the appropriate consideration of non-detects and the limited availability of reliable PNECs and EQS values.

Carbonyl reductase (CBR) is an enzyme involved in protection from oxidative stress. In rainbow trout (Oncorhynchus mykiss), the hepatic mRNA abundance of the two isoforms (A and B) is increased after exposure to treated sewage effluents, as well as after exposure with β-naphthoflavone (β-NF) and the pro-oxidant paraquat. In this study, we show that the same chemicals similarly increase the single known hepatic CBR mRNA level and CBR catalytic activity in the coastal living eelpout (Zoarces viviparus). Hepatic CBR mRNA abundance and catalytic activity were also compared between eelpout collected at contaminated and reference sites on the Swedish west coast, but no differences were observed. In conclusion, CBR is a potential biomarker candidate for monitoring the exposure and effects of AhR agonists and/or pro-oxidants in the marine environment, but more research is needed to investigate temporal regulation as well as dose dependency for different chemicals. The mRNA and enzymatic assays presented in this study provide two additional tools for researchers interested in expanding their biomarker battery.

Zebrafish (Danio rerio) is not only a widely used species in the Fish Embryo Toxicity (FET) test but also an emerging model in behavioural ecotoxicology. By using automatic behaviour tracking technology, locomotion of developing zebrafish (ZF) larvae can be accurately recorded and potentially used in an ecotoxicological context to detect toxicant-induced behavioural alterations. In this study, we explored if and how quantitative locomotion data can be used for sub-lethal toxicity testing within the FET framework. We exposed ZF embryos to silver ions and nanoparticles, which previously have been reported to cause neurodevelopmental toxicity and behavioural retardation in early-life stages of ZF. Exposure to a broad range of silver (Ag(+) and AgNPs) concentrations was conducted, and developmental toxicity was assessed using FET criteria. For behavioural toxicity assessment, locomotion of exposed ZF eleutheroembryos (120hpf) was quantified according to a customised behavioural assay in an automatic video tracking system. A set of repeated episodes of dark/light stimulation were used to artificially stress ZF and evoke photo-motor responses, which were consequently utilized for locomotion profiling. Our locomotion-based behaviour profiling approach consisted of (1) dose-response ranking for multiple and single locomotion variables; (2) quantitative assessment of locomotion structure; and (3) analysis of ZF responsiveness to darkness stimulation. We documented that both silver forms caused adverse effects on development and inhibited hatchability and, most importantly, altered locomotion. High Ag(+) and AgNPs exposures significantly suppressed locomotion and a clear shift in locomotion towards inactivity was reported. Additionally, we noted that low, environmentally relevant Ag(+) concentrations may cause subordinate locomotive changes (hyperactivity) in developing fish. Overall, it was concluded that our locomotion-based behaviour-testing scheme can be used jointly with FET and can provide endpoints for sub-lethal toxicity. When combined with multivariate data analysis, this approach facilitated new insights for handling and analysis of data generated by automatized behavioural tracking systems.

Protein carbonyl's and tbars as biomarkers for oxidative damage in eelpout (Zoacers vivaparus)

During 2002-2003 a dredging campaign was undertaken in Göteborg harbor, Sweden, to widen and deepen shipping lanes. A bunker oil spill occurred in the harbor in July 2003, thereby further exasperating the situation for marine life. Eelpout, Zoarces viviparus, was used as a sentinel species to monitor the impact of these events. Here, we have investigated the effects on two liver parameters, lipid peroxidation and protein carbonylation, which can serve as biomarkers for oxidative stress. Lipid peroxidation data, measured as TBARS, in eelpout from the field study showed no significant differences between reference and polluted sites, at any of the time points. These results are mirrored in a laboratory exposure to the bunker oil where no differences were seen between the control and the exposure groups. A trend towards a seasonal cycle in TBARS levels in eelpout liver was observed, with highest amounts measured during the warmer months, 97.08+/-14.45 nmol/g liver in the harbor in July compared to 41.20+/-2.66 nmol/g liver in November 2001. Protein carbonylation, measured using an ELISA method, did, however, show differences between the reference and polluted sites in the field, as well as differences between time periods (before and during dredging and following the oil spill). The laboratory exposure indicated that the formation and/or accumulation of protein carbonyls is greatly affected by exposure to this PAH rich oil. Levels in the control group were 1.76+/-0.13 nmol/mg protein while those fish exposed to the high dose had 6.23+/-0.17 nmol/mg protein. We concluded that TBARS is not an appropriate biomarker for pollutant mediated oxidative damage in eelpout while protein carbonyl formation does appear to be affected by xenobiotic exposure.

2,4,6-Trinitrotoluene (TNT) is the major explosive in ammunition dumped into lakes and the sea after World War II. To identify useful biomarkers of TNT-exposure for forthcoming fish monitoring studies at ammunition dumping sites, rainbow trout (Oncorhynchus mykiss) were intraperitoneal (i.p.) injected with TNT in peanut oil at doses of 0, 100, 200 or 400 mg TNT/kg body weight and sampled 72 h later. The study covered blood parameters, and hepatic antioxidant and detoxifying enzymes. Fish treated with TNT had an increased glutathione S-transferase (GST) activity and glutathione reductase (GR) activity, and a decreased percentage of oxidised glutathione (%GSSG) compared to the control group. In addition to increased methemoglobin, the increased glutathione and glutathione dependent enzyme activities indicate that TNT oxidises macromolecules and activates antioxidant defence systems which may be useful as general biomarkers of TNT-exposure. The fish bile was analysed for TNT and its metabolites by gas chromatography-mass spectrometry (GC-MS), and the toxicity of the bile was determined with the cladoceran Ceriodaphnia dubia. A dose-dependent increase in TNT, 2-amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) was found in the hydrolysed bile of the TNT-treated fish. These results indicate that the fish are able to detoxify and excrete TNT and suggest that the detection of TNT, 2-ADNT and 4-ADNT in bile may be suitable as a direct marker of exposure to TNT.

Effects of nanoparticles on aquatic organisms have been little studied to date and toxicological data are urgently needed for development of regulatory frameworks for these substances. Here, we report the findings of a study exposing sticklebacks to cadmium sulfide (CdS) as bulk material and quantum dots. Fish were exposed for 21 d in a flow through test system to 5, 50 or 500 microg l(-1) CdS nanoparticles (nCdS) coated in thiol terminated methyl polyethylene glycol (MPEG), bulk CdS or MPEG at 500 microg l(-1) (nominal concentrations). With the exception of the highest nCdS exposure, measured concentrations were approximately one order of magnitude below nominal. A single fish from each group (excluding MPEG) was examined using energy dispersive X-ray analysis (EDX) to localise cadmium, however, cadmium could not be detected in whole body sections. Elevated levels of oxidized glutathione were measured in the gills of fish exposed to 50 and 500 microg l(-1) nCdS. Induction of vitellogenin synthesis was not detected in any of the treatment groups. The number of males engaged in nest-building behaviour following exposure to 500 microg l(-1) nCdS was reduced and livers of 4/6 fish in the same treatment displayed hepatocellular nuclear pleomorphism. The results are discussed emphasising the fundamental importance of experimental design and the need to understand the behaviour of nanoparticles in the aqueous phase.

In the present study, long-term exposure to PCB resulted in marked induction of the xenobiotic biotransformation enzymes UDP glucuronosyl transferase, glutathione transferase and DT-diaphorase (measured as menadione reductase and DCPIP reductase) in rainbow trout 15 or 20 weeks post-injection during the experimental period. The increase of the phase II enzymes were paralleled by a marked increase of the glutathione reductase activity. These results indicate that prolonged exposure to PCB results in increased production of organo-radicals.

Studies were carried out to examine the role of aromatic hydrocarbon (AH)-quinone metabolism and DT-diaphorase (quinone oxidoreductase; EC 1.6.99.2; DTD) function in pollution-caused oxidative damage in fish. Redox cycling of quinones to produce reactive oxygen species (ROS) was studied by oxygen consumption and oxidation of the hydroxyl radical scavenger 2-keto-4-methiolbutyric acid. NADH-dependent redox cycling by hepatic microsomes of flounder (Platichthys flesus) was seen for 1,4-benzoquinone, 1,2- and 1,4-naphthoquinones, phenanthroquinone and anthraquinone. Combined with previous similar results for duroquinone, menadione and benzo[a]pyrene (BaP) quinones (Lemaire et al., 1994), this demonstrates a general potential for AH-quinone-mediated ROS production. NADPH-dependent 3H-BaP metabolism by hepatic microsomes of P. flesus produced 54% dials, 45% phenols and trace levels of quinones, whereas cumene hydroperoxide-dependent metabolism produced up to 67% quinones (metabolites resolved by HPLC). Exposure to AH-contaminated sediment can lead to enhanced hepatic lipid peroxidation (Livingstone et al., 1993) raising the possibility of an AH-mediated toxicity cycle of peroxidation leading to enhanced quinone and ROS production leading to more peroxidation and oxidative damage. Hepatic DTD purified almost to homogeneity from trout (Oncorhynchus mykiss); Sturve et al., in prep.) also catalysed NADH-dependent AH-quinone-mediated ROS production, indicating a possible function for this enzyme in quinone toxicity.

In situ biomonitoring of roundnose grenadier (Coryphaenoides rupestris) and Norway lobster (Nephrops norvegicus) and measurements of pollutants levels in caged common mussel (Mytilus edulis) were used to assess the environmental impact of contaminants in the Skagerrak and Kattegat and near the Faroe Islands. By comparing the responses of a suite of established and potential biomarkers, i.e. different liver detoxification enzymes and histopathology in roundnose grenadier in the Skagerrak, and at a reference site off the Faroe Islands, evidence was found that the deeper parts of the Skagerrak are affected by anthropogenic contaminants. Levels of polycyclic aromatic hydrocarbons (PAH) and organochlorines in caged mussels were compared with those sediment. The Skagerrak/Kattegat area was found to be more polluted than the Faroe area and the caged mussels bioaccumulated the PAHs to a higher degree than the organochlorines. This seems to indicate that the PAHs in the sediments are more bioavailable than the organochlorines and that the induced ethoxyresorufin-O-deethylase activities observed in Skagerrak roundnouse grenadier may be due to PAH exposure. The Norway lobster accumulated manganese to various degrees in the different parts of the organism. The excretion rate of manganese could not cope with the uptake rate at higher exposure concentrations. Enhanced manganese concentrations, especially in the gills and haemolymph, seem to be a useful indication of increased manganese levels in the water. Hypoxia makes a sediment-bound manganese much more bioavailable. The elevated levels of manganese in Norway lobster from the Skagerrak/Kattegat area therefore seem to reflect the increasing occurrence of hypoxic conditions in the Kattegat and coastal areas of the Skagerrak. Accumulated manganese in Norway lobster may thus serve as a biomarker of hypoxia. In conclusion, the use of a suite of different biomarkers in ecotoxicological and ecophysiological studies and analyses of contaminant levels have provided evidence of a large-scale environmental impact of pollutants and nutrients in the Skagerrak/Kattegat area. This should lead to further efforts to decrease transport into and deposition of waste compounds in the sea.

Brominated dioxins have recently been detected in Baltic Sea biota. Due to their similarities to the highly toxic chlorinated dioxins, concern has been raised about their potential biological effects. The present study investigated retention and effects of brominated dioxins in adult zebrafish, as well as maternal transfer and effects on offspring. We exposed adult zebrafish for nine weeks via feed to 2,3,7,8-tetrabromodibenzo-p-dioxin (TBDD) or to a mixture of brominated dioxins (Baltic Sea mixture), which was designed to reflect relative concentrations found in Baltic Sea biota. We studied spawning success, gonad morphology, hepatic vitellogenin gene expression, and offspring early life-stage development to investigate effects on zebrafish reproduction. Hepatic ethoxyresorufin-O-deethylase (EROD) activity and hepatic expression of a number of aryl hydrocarbon receptor (AHR)-regulated genes were studied to investigate if the brominated dioxins can activate gene transcription through the AHR pathway in zebrafish. In addition, glutathione reductase activity and expression of genes involved in adaptive responses to intracellular stress were studied to investigate potential stress effects of brominated dioxins. After nine weeks of exposure, all brominated dioxins spiked to the feed were detected in female fish and transferred to eggs. Exposure to the Baltic Sea mixture and TBDD clearly induced AHR-regulated genes and EROD activity. Exposure to TBDD reduced spawning success, altered ovarian morphology and reduced hepatic vitellogenin gene expression, which implies that TBDD has a similar effect pattern as the chlorinated analogue. Overall, our results show that dietary exposure to sublethal concentrations of brominated dioxins may impair reproductive physiology in fish and induce AHR-regulated genes.

At selected sites around the UK, the offshore sentinel flatfish species dab Limanda limanda are found to contain elevated levels of macroscopic liver tumors. Previous proteomic and metabolomic studies have demonstrated that differences exist between tumor and non-tumor tissues; however, these differing features were not identified, and little is known about the changes at the gene expression level, or whether prognostic markers are present and can be identified. A flounder Platichthys flesus custom cDNA microarray and RT-PCR were used to investigate hepatic mRNA expression in the histologically confirmed tumors, hepatocellular adenoma (HA) and hepatocellular carcinoma (HC) from dab, and in adjacent normal tissue from the same fish. Differences in gene expression were observed between tumor and normal tissues, and between tumor types. A class-prediction approach using 50 transcripts revealed sufficient group-specific expression profiles to allow segregation of samples dependent on their tumor type or the sex of the host. Vitellogenins were found to display the greatest induction (up to 500-fold induction) in some HC tumors from female fish and in both HA and HC tumors from males. To the best of our knowledge, this is the first report of the association of vitellogenin expression with tumors of wild fish.

Laser-assisted microdissection (LMD) has been developed to isolate distinct cell populations from heterogeneous tissue sections, cytological preparations, or live cell samples. Downstream applications typically include gene expression studies using real-time PCR and array platforms, diagnostic PCR, and protein expression studies. LMD techniques are now commonplace in mainstream biological research and clearly have suitable applications in the field of aquatic pathology and parasitology. The present study used LMD to isolate 2 dinoflagellate parasites (Hematodinium spp.) from formalin-fixed paraffin-embedded tissue sections from 2 crustacean hosts, Cancer pagurus and Portunus trituberculatus. DNA was isolated from LMD parasite preparations, and partial regions (up to 300 bp) of the small subunit and the first internal transcribed spacer region of the rRNA gene complex from the Hematodinium spp. were PCR amplified using diagnostic primers. The amplification products were sequenced to confirm the identity of the targeted regions. The techniques, applications, and limitations of LMD to address questions in aquatic molecular pathology and parasitology are discussed.

The use of fish in environmental monitoring has become increasingly important in recent years as anthropogenic substances, many of which function as prooxidants, are accumulating in aquatic environments. We have measured a battery of antioxidant defenses as a measure of oxidative status, as well as protein carbonylation as a measure of oxidative damage, in corkwing wrasse (Symphodus melops) captured near a disused copper mine, where water and sediment are contaminated with heavy metals, and an aluminum smelter, a site contaminated with PAHs. Results were compared to two different reference sites. Fish at the heavy metal site had lower glucose-6-phosphate dehydrogenase activity and elevated protein carbonyls (1.8 times) compared to fish from the reference site. At the PAH site, EROD was increased 2-fold, while total glutathione and methemoglobin reductase concentration, were decreased. No differences were seen in protein carbonyl levels at the PAH site. Measures of both antioxidant defenses and oxidative damage should be used when assessing effects of xenobiotics on oxidative stress in fish species.

A consequence of oil drilling at sea is the release of produced water contaminated with e.g. polycyclic aromatic hydrocarbons (PAH) and alkylphenols. In the present study, juvenile Atlantic cod were exposed to North Sea oil, nonylphenol and a combination of the North Sea oil and an alkylphenol mixture in a flow-through system. A suite of hepatic biomarkers were analysed. Exposure to North Sea oil resulted in strong induction of CYP1A protein levels and EROD activities. Exposure to nonylphenol, on the other hand, resulted in decreased CYP1A levels and EROD activities. Thus, nonylphenol appears to down-regulate CYP1A expression in Atlantic cod. Combined exposure to North Sea oil with an alkylphenol mixture resulted in lower EROD induction, compared to that in fish exposed to North Sea oil alone. This difference was not statistically significant, but still we believe that the alkylphenols have inhibited CYP1A activities in the fish which may have compromised CYP1A mediated metabolism of other xenobiotics, including PAH. CYP3A protein levels were lower, compared to controls, in fish exposed to nonylphenol and the combination of North Sea oil and alkylphenol mixture. In contrast, the oil alone had no effect on CYP3A protein content. North Sea oil exposure, alone or in combination with alkylphenols, caused oxidative stress observed as elevated levels of GSSG content and GR and CAT activities. Interestingly, exposure to nonylphenol resulted in a marked depletion of total glutathione levels. This apparent depletion may be a consequence of increased conjugation of glutathione to nonylphenol followed by excretion. An increase in conjugation enzyme GST activity was observed in the nonylphenol exposed group, although the difference was not significant. No sign of oxidative damage, measured as lipid peroxidation, was observed in any of the exposures experiments. This study suggests that North Sea oil may lead to oxidative stress and altered CYP1A and CYP3A expression. Alkylphenols, present in produced water, resulted in decreased CYP1A and CYP3A protein expression in Atlantic cod.

The present study investigated the proposed involvement of contaminant-stimulated reactive oxygen species (ROS) production in disease processes in fish. NAD(P)H-dependent ROS production of subcellular fractions was determined by the iron/EDTA-mediated oxidation of 2-keto-4-methiolbutyric acid. Hepatic cytosolic NADPH-dependent and microsomal NAD(P)H-dependent ROS production were increased 51--160% (P < 0.05) in rainbow trout (Oncorhynchus mykiss) 15 weeks after a single i.p. injection of polychlorobiphenyl (PCB) (100 mg Clophen A50 kg -1 wet wt.). Hepatic microsomal NADH-dependent ROS production was 114% higher in perch (Perca fluviatilis) from PCB-contaminated Lake Jarnsjon compared to clean Lake Vanern, Sweden. Hepatic mitochondrial NADH-dependent, cytosolic NADH-dependent and microsomal NADPH-dependent ROS production were variously elevated up to 160% in flounder (Platichthys flesus) at various sites along two pollution transects near to the ports of Rotterdam and Amsterdam, Netherlands. Overall the data indicate increased potential for ROS production in liver of fish exposed to field pollution, and support the hypothesis of oxidative stress as a mechanism of contaminant-mediated disease in fish.

Shorthorn sculpins (Myoxocephalus scorpius) were caught in four Icelandic harbours, differing in size, use and traffic. Biochemical responses in liver were measured and chemicals analysed in bile. Eyrarbakki harbour, which has not been in use for many years was chosen as a control site. Njar partial differentialvík harbour is a small fishing harbour and a marina, Sandger partial differentiali harbour is a large fishing harbour, and Reykjavík harbour is a large fishing harbour and an international transport harbour. Higher levels of DNA-adducts and cytochrome P4501A (CYP1A) in the fish from the harbours in Sandger partial differentiali, Njar partial differentialvík and Reykjavík, compared to Eyrarbakki harbour, indicate PAH exposure. This was confirmed by PAH analysis in bile. The higher activities of the antioxidant enzymes catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR) in fish caught in Sandger partial differentiali, than in fish caught in the other harbours, indicate exposure of sculpin to prooxidative compounds in Sandger partial differentiali harbour. Shorthorn sculpin seems to be a convenient species for monitoring pollution in northern coastal areas.

Sewage treatment works (STWs) are a common source of chemicals entering into the aquatic environment. In order to assess effects of these effluents on oxidative stress parameters in aquatic organisms, we caged rainbow trout at five sites: upstream, near an STW effluent, and three sites downstream in the river Viskan in western Sweden for 14 days during autumn, 2006. We then measured protein carbonyls in plasma as well as 20S proteosome activity and lipid peroxidation products, i.e. MDA and 4-HNE, in liver samples. Levels of both lipid and protein oxidative damage products were elevated in fish caged near the STW effluent while 20S activity showed no differences. This argues that complex mixtures of chemicals entering into the aquatic environment do have deleterious effects on fish. Additionally, oxidative stress parameters can serve as a biomarker in aquatic organisms.

The shorthorn sculpin Myoxocephalus scorpius is a convenient species for monitoring pollution in northern coastal areas. In this study, a series of biomarkers was measured in sculpin collected from four Icelandic harbours differing in size, use and traffic. Eyrarbakki Harbour has been inactive for many years and was chosen as a control site. Njardvik Harbour is a small fishing harbour with a marina; Sandgerdi Harbour is a large fishing harbour; and Reykjavik Harbour is a large fishing harbour and an international transport harbour. Higher levels of DNA-adducts and cytochrome P4501A in fish from Sandgerdi, Njardvik and Reykjavik harbours compared to fish from Eyrarbakki Harbour indicate polycylic aromatic hydrocarbon (PAH) exposure. This was confirmed by PAH analysis in bile. The higher activity of the antioxidant enzymes catalase, glutathione peroxidase and glutathione reductase in fish from Sandgerdi Harbour compared to fish from other harbours indicates the presence of prooxidative compounds in Sandgerdi Harbour.

In October 1997, juvenile rainbow trout Oncorhynchus mykiss were exposed to effluent water from two different sewage treatment plants (STPs) in the Goteborg area: Grabo, a small STP north of Goteborg and GRYAAB, a large STP that processes the sewage from the city of Goteborg. In Grabo, two groups of fish were caged, one upstream and one downstream. Fish were sampled after 2 and 4 weeks. In GRYAAB, fish were exposed to outlet water in aquaria. The exposure concentrations were 0, 2, 20, and 50% diluted with dechlorinated tap water. Fish were sampled after 5 days, 2, 4 and 10 weeks. The CYP1A (EROD activity) showed a time-dependent dose-response induction in the GRYAAB experiment. In Grabo, EROD activity was lower in the exposed group compared with the control. The different EROD responses caused by the two effluents may be due to the high levels of estrogens in the Grabo effluent (Parkkonen et al., (2000). Marine Environmental Research 50(1–5), this issue). The conjugating enzyme glutathione-S transferase and the antioxidant enzymes catalase, glutathione reductase and DT-diaphorase showed no significant responses.