Jari Parkkonen

Ethinyloestradiol — an undesired fish contraceptive?

Environmental oestrogens are natural or synthetic substances present in the environment, which imitate the effects of endogenous oestrogen. Oestrogenic substances were identified by gas chromatogra ...

Triclosan, a commonly used bactericide found in human milk and in the aquatic environment in Sweden.

High levels of the commonly used, effective bactericide Triclosan was found in three out of five randomly selected human milk samples. It was also found in the bile of fish exposed to municipal wastewater and in wild living fish from the receiving waters of the three wastewater treatment plants.

Therapeutic levels of levonorgestrel detected in blood plasma of fish : results from screening rainbow trout exposed to treated sewage effluents

Pharmaceuticals are found in surface waters worldwide, raising concerns about effects on aquatic organisms. Analyses of pharmaceuticals in blood plasma of fish could provide means to assess risk for pharmacological effects, as these concentrations could be compared with available human therapeutic plasma levels. In this study we investigated if fish exposed to sewage effluents have plasma concentrations of pharmaceuticals that are approaching human therapeutic levels. We also evaluated how well the bioconcentration of pharmaceuticals into fish blood plasma can be predicted based on lipophilicity. Rainbow trout were exposed to undiluted, treated sewage effluents at three sites in Sweden for 14 days. Levels of 25 pharmaceuticals in blood plasma and effluents were analyzed with liquid chromatography-mass spectrometry/mass spectrometry and gas chromatography-high resolution mass spectrometry. The progestin pharmaceutical levonorgestrel was detected in fish blood plasma at concentrations (8.5-12 ng mL(-1)), exceeding the human therapeutic plasma level. In total 16 pharmaceuticals were detected in fish plasma at concentrations higher than 1/1000 of the human therapeutic plasma concentration. Twenty-one pharmaceuticals were detected in either plasma or effluent, and 14 were detected in both compartments, allowing plasma bioconcentration factors to be determined. For 11 of these, theoretically calculated and experimentally measured values were in reasonably good agreement. However a few drugs, including levonorgestrel, did not bioconcentrate according to the screening model used. This study shows that rainbow trout exposed to sewage effluents have blood plasma levels of pharmaceuticals similar to human therapeutic concentrations, suggesting a risk for pharmacological effects in the fish. There is a particular concern about effects of progestin pharmaceuticals. For levonorgestrel, the measured effluent level (1 ng/L) was higher than water levels shown to reduce the fertility of fish.

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.

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.

Estrogenicity of domestic and industrial effluents in Sweden

Estrogenicity in Swedish wastewaters was surveyed. Estrogenicity was examined using a recombinant yeast cell test and analyses of the yolk protein precursor in the blood of caged juvenile rainbow trout at sites close to wastewater outlets or in continuous flow tanks of undiluted wastewater. Estrogenic effects corresponding to <0.1 to 15 ng estradiol equivalents were found in municipal effluents. In 9 of 12 industrial effluents, estrogenic effects were below the detection limit of the screen test. None of the six effluents from the pulp and paper industry, from a steel works, and from two chemical industries had a detectable estrogenic effect. Two assays were used for analysis of plasma vitellogenin, a heterologous enzyme-linked immunosorbent assay and a homologous radio immunoassay. Exposure to undiluted municipal wastewater resulted in increased levels of vitellogenin in the plasma of juvenile trout. Using the more sensitive RIA test, increased levels of plasma vitellogenin were detected in cage exposure of rainbow trout in municipal effluent receiving waters. Textile industry effluents mixed with domestic wastewater were estrogenic probably due to contributions from the domestic effluent. In some effluents, toxicity may have masked estrogenic effects.

Contraceptive pill residues in sewage effluent are estrogenic to fish

Abstract The estrogenicity of sewage effluent water to fish has been described in several countries. Natural oestrogens have been confirmed as major causing agents in several British rivers. In a recent study (Larsson et al., Aquatic Toxicology, 1999, 91–97) we identified estrogenic substances by GC/MS in effluent water from a Swedish sewage treatment works receiving mainly domestic wastewater. Substances found include the synthetic estrogen used in contraceptives 17α-ethinylestradiol (4.5 ng/l), the natural estrogens estrone (5.8 ng/l) and 17β-estradiol (1.1 ng/l) and the weaker non-steroidal estrogens 4-nonylphenol (840 ng/l) and bisphenol A (490 ng/l). Ethinylestradiol exceeded levels shown to be estrogenic to fish by 45 times. The estrogenicity of the effluent water was investigated by introducing juvenile rainbow trout (Oncorhynchus mykiss) in cages downstream of the sewage treatment works. All estrogens indicated above were present in the bile of the fish, and the estrogen-inducible protein, vitellogenin, was found in large amounts in the plasma as determined by ELISA and Western blotting. At present we are studying effects further downstream on caged as well as wildcaught fish. Our studies suggest that a widely used synthetic estrogen affects the endocrine systems of fish exposed to sewage effluent water.

A gene to organism approach—assessing the impact of environmental pollution in eelpout (Zoarces viviparus) females and larvae

Abstract A broad biomarker approach was applied to study the effects of marine pollution along the Swedish west coast using the teleost eelpout (Zoarces viviparus) as the sentinel species. Measurements were performed on different biological levels, from the molecular to the organismal, including measurements of messenger RNA (mRNA), proteins, cellular and tissue changes, and reproductive success. Results revealed that eelpout captured in Stenungsund had significantly higher hepatic ethoxyresorufin O‐deethylase activity, high levels of both cytochrome P4501A and diablo homolog mRNA, and high prevalence of dead larvae and nuclear damage in erythrocytes. Eelpout collected in Göteborg harbor displayed extensive macrovesicular steatosis, whereby the majority of hepatocytes were affected throughout the liver, which could indicate an effect on lipid metabolism. Results also indicate that eelpouts collected at polluted sites might have an affected immune system, with lower mRNA expression of genes involved in the innate immune system and a higher number of lymphocytes. Biomarker assessment also was performed on livers dissected from unborn eelpout larvae collected from the ovary of the females. No significant differences were noted, which might indicate that the larvae to some extent are protected from effects of environmental pollutants. In conclusion, usage of the selected set of biological markers, covering responses from gene to organism, has demonstrated site‐specific biomarker patterns that provided a broad and comprehensive picture of the impact of environmental stressors. Environ Toxicol Chem 2015;34:1511–1523.

Biomarker responses in eelpouts from four coastal areas in Sweden, Denmark and Germany

To increase our understanding of possible chemical impacts on coastal fish populations in the Baltic Sea, Kattegat and Skagerrak, the viviparous eelpout (Zoarces viviparus) was used as sentinel species in two major sampling campaigns (spring and autumn) in 16 different coastal sites. Condition factor (CF), liver somatic index (LSI), gonad somatic index (GSI) were measured and the activity of the hepatic enzymes ethoxyresorufin-O-deethylase (EROD), glutathione reductase GR), glutathione S-transferase (GST), catalase (CAT) and muscular activity of acetylcholinesterase (AChE) were assessed. PAH metabolites in bile were also analyzed. The most notable finding in the data set was the low EROD activity in eelpouts collected at the relatively polluted region in Germany compared to the other regions, which could be due to an inhibition of the CYP1A-system or to adaptation to chronic exposure of pollutants in this area. Additionally, low AChE activity was noted in the German region in the autumn campaign and low AChE activity detected in the Danish region in the spring campaign. These differences suggest possible season-specific differences in the use and release of AChE-inhibiting chemicals in the Danish and German regions. Clustering of biomarkers on site level indicated a relationship between CF and GSI and suggested that sites with a high CF contained eelpout that put a larger effort into their larvae development. Clustering of the oxidative stress markers GR, GST and CAT on the individual level reflected a possible coordinated regulation of these enzymes. Overall, the results support the importance of taking into account general regional differences and seasonal variation in biomarker activity when monitoring and assessing the effects of pollution. Despite the expected seasonal variation for most of the measured endpoint, several markers (GSI, EROD and CF) vary similarly between all selected sites in both spring and autumn. This suggests that the differences between sites for these endpoints are independent of season.

Biomarker responses in rainbow trout (Oncorhynchus mykiss) exposed to sewage treatment plant effluent

Abstract 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.