Eiríkur Stephensen

Biochemical indicators of pollution exposure in shorthorn sculpin (Myoxocephalus scorpius), caught in four harbours on the southwest coast of Iceland

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.

Effects of redox cycling compounds on glutathione content and activity of glutathione-related enzymes in rainbow trout liver

In fish, as in other aerobic organisms, glutathione and glutathione-related enzymes are important components in the defences against oxidative stress. To study if hepatic glutathione levels and/or activities of glutathione-related enzymes can act as indicators of oxidative stress in fish, we injected rainbow trout (Oncorhynchus mykiss) intraperitoneally with paraquat (PQ), menadione (MD), naphthazarin (DHNQ), or beta-naphthoflavone (beta-NF), all known to cause a rise in reactive oxygen species (ROS). After 2 and 5 days of exposure, we measured the activities of hepatic glutathione peroxidase (GPox), glutathione S-transferase (GST), gamma-glutamylcysteine synthetase (GCS), and glutathione reductase (GR). We also measured total glutathione (tGSH) and oxidised glutathione (GSSG) in the liver of fish treated with PQ and MD. All chemicals caused an increase in GR activity after 5 days, which ranged from 160% in fish treated with beta-NF to 1,500% in fish treated with PQ. All chemicals except beta-NF caused moderate elevation in GST activity; GPox activity was lower in fish treated with DHNQ and MD, while GCS activity increased twofold in the fish treated with DHNQ, without being affected by beta-NF, PQ or MD. After 5 days of treatment with PQ or MD, tGSH content was elevated. Our findings demonstrated that of the parameters included in the study, GR activity was the most responsive to treatment with redox cycling compounds, indicating that GR activity is a promising biomarker of such compounds and possibly indicating oxidative stress in rainbow trout.

Plasma Glutathione and Carotenoid Coloration as Potential Biomarkers of Environmental Stress in Great Tits

Measures of oxidative stress in animals may be useful biomarkers of environmental stressors, such as anthropogenic pollution. In birds, studies of oxidative stress have focused on dietary antioxidants, primarily carotenoids, which are interesting due to their multiple physiological and pigmentary functions but therefore also unspecifically related to oxidative stress. A useful complementary biomarker may be the glutathione system, commonly used in human medicine, but rarely applied to wild, terrestrial vertebrates. In this study of urban versus rural adult and nestling great tits Parus major, we investigated both the carotenoid-based yellow plumage (by reflectance spectrometry) and the plasma levels of glutathione, the latter measured as total glutathione (tGSH) and as the ratio between oxidized and reduced glutathione (GSSG:GSH), respectively. We found that urban adults had higher current oxidative stress (GSSG:GSH) and paler yellow plumage compared to rural adults, suggesting elevated stress in the urban environment. Total glutathione levels (tGSH), however, which may indicate long-term up-regulation of the GSH reservoir, did not differ between the environments. Nestlings did not show any consistent pattern between environments in either tGSH or GSSG:GSH and, among individuals, glutathione levels were uncorrelated with carotenoid coloration. The results thus suggest some population-level correspondence between the two stress biomarkers in adult birds, but more work is obviously needed to understand how the two antioxidant systems interact in different individuals and in response to different environmental disturbances.

Biomarker responses and chemical analyses in fish indicate leakage of polycyclic aromatic hydrocarbons and other compounds from car tire rubber

Rubber tire material contains toxic compounds including oils rich in polycyclic aromatic hydrocarbons (PAH), so-called highly aromatic (HA) oils, as well as other reactive additives used as antioxidants, antiozonants, and vulcanization accelerators. The toxicity of rubber tire leachates to aquatic organisms has been demonstrated before. However, previous studies have focused on lethal rather than sublethal effects. We kept rainbow trout (Oncorhynchus mykiss) in tanks with two types of tires: a tire containing HA oils in the tread or a tire free of HA oils in the tread. After 1 d of exposure, an induction of cytochrome P4501A1 (CYP1A1) was evident in both exposed groups, measured as elevated ethoxyresorufin-O-deethylase (EROD) activity and increased CYP1A1 mRNA levels. After two weeks of exposure, EROD activity and CYP1A1 mRNA were still high in fish exposed to leachate from HA oil-containing tire, whereas the effect was somewhat lower in fish exposed to leachate from HA oil-free tread tire. Compounds in the tire leachates also affected antioxidant parameters. Total glutathione concentration in liver as well as hepatic glutathione reductase, glutathione S-transferase, and glucose-6-phosphate dehydrogenase activities were markedly elevated after two weeks of exposure in both groups. The responses were greater in the group exposed to leachate from HA oil-free tread tire. Vitellogenin measurements did not indicate leakage of estrogenic compounds from the tires. Chemical analyses of bile from exposed fish revealed the presence of hydroxylated PAH as well as aromatic nitrogen compounds indicating uptake of these compounds by the fish.

Protein carbonyls and antioxidant defenses in corkwing wrasse (Symphodus melops) from a heavy metal polluted and a PAH polluted site.

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.

Effects of dredging in Göteborg harbor, Sweden, assessed by biomarkers in eelpout (Zoarces viviparus).

We used a battery of biomarkers in fish to study the effects of the extensive dredging in Göteborg harbor situated at the river Göta alv estuary, Sweden. Eelpout (Zoarces viviparus) were sampled along a gradient into Göteborg harbor, both before and during the dredging. Biomarker responses in the eelpout before the dredging already indicated that fish in Göteborg harbor are chronically affected by pollutants under normal conditions compared to those in a reference area. However, the results during the dredging activities clearly show that fish were even more affected by remobilized pollutants. Elevated ethoxyresorufin-O-deethylase activities and cytochrome P4501A levels indicated exposure to polycyclic aromatic hydrocarbons. Elevated metallothionein gene expression indicated an increase in metal exposure. An increase in general cell toxicity, measured as a decrease in lysosomal membrane stability, as well as effects on the immune system also could be observed in eelpout sampled during the dredging. The results also suggest that dredging activities in the Göta alv estuary can affect larger parts of the Swedish western coast than originally anticipated. The present study demonstrates that the application of a set of biomarkers is a useful approach in monitoring the impact of anthropogenic activities on aquatic environments.

Effects of redox cycling compounds on DT diaphorase activity in the liver of rainbow trout (Oncorhynchus mykiss)

BackgroundDT diaphorase (DTD; NAD(P)H:quinone oxidoreductase; EC 1.6.99.2) catalyses the two electron reduction of quinones, thus preventing redox cycling and consequently quinone dependent production of reactive oxygen species. In rat and mouse, a wide range of chemicals including polyaromatic hydrocarbons, azo dyes and quinones induces DTD. Bifunctional compounds, such as β-naphthoflavone (β-NF) and benzo(a)pyrene (B(a)P), induce DTD together with CYP1A and phase II enzymes by a mechanism involving the aryl hydrocarbon receptor (AHR). Monofunctional induction of DTD is mediated through the antioxidant response element and does not lead to the induction of AHR dependent enzymes, such as CYP1A. The aim of this study was to investigate the effects of prooxidants (both bifunctional and monofunctional) on the activity of hepatic DTD in rainbow trout (Oncorhynchus mykiss) in order to evaluate DTD suitability as a biomarker. We also investigated the effect of β-NF on hepatic DTD activity in perch (Perca fluviatilis), shorthorn sculpin (Myoxocephalus scorpius), eelpout (Zoarces viviparus), brown trout (Salmo trutta) and carp (Cyprinus carpio). In addition, the effect of short term exposure to prooxidants on catalase activity was investigated.ResultsIn rainbow trout, hepatic DTD activity is induced by the bifunctional AHR agonists β-NF and B(a)P and the monofunctional inducers naphthazarin, menadione and paraquat. Although exposure to both B(a)P and β-NF led to a strong 7-ethoxyresorufin-O-deethylase (EROD) induction, none of the monofunctional compounds affected the rainbow trout EROD activity. DTD was not induced by β-NF in any of the other fish species. Much higher DTD activities were observed in rainbow trout compared to the other fish species. Catalase activity was less responsive to short term exposure to prooxidants compared to DTD.ConclusionSince rainbow trout hepatic DTD activity is inducible by both monofunctional and bifunctional inducers, it is suggested that rainbow trout DTD may be regulated by the same mechanisms, as in mammals. The fact that DTD is inducible in rainbow trout suggests that the enzyme may be suitable as a part of a biomarker battery when rainbow trout is used in environmental studies. It appears as if DTD activity in rainbow trout is higher and inducible compared to the other fish species studied.

Biochemical indicators of pollution exposure in shorthorn sculpin (Myoxocephalus scorpius) caught in four Icelandic harbours

Abstract 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.