Probing the xeno-oestrogenic impact of sediment cores contaminated by the pulp and paper industry: induction of aromatase cyp19a1b in late larval zebrafish
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Chemicals in effluents from the wood industry have been widely identified as sources of contamination affecting endocrine function in aquatic animals. This study examined the changes in transcript levels of key biomarkers, cytochrome P450 1A (cyp1a), brain aromatase (cyp19a1b) and vitellogenin 1 (vtg1), in late larval zebrafish exposed to sediment cores from areas upstream and downstream (two distances) from a pulp and paper mill in Finland.
Materials and methods
Bioluminescence inhibition of Vibrio fischeri by sediment elutriates was measured to determine the overall toxicity of the sediment. Then, a whole-sediment bioassay with 20-day post-fertilization zebrafish (20dpfZF) to lake sediment (0–5-cm and 20–30-cm layers) was conducted for 3 days, and changes in cyp1a, cyp19a1b and vtg1transcript levels were assessed using quantitative reverse transcription PCR (qRT-PCR) analyses.
Results and discussion
Lack of inhibition of bacterial (V. fischeri) bioluminescence by sediment elutriates indicated the non-toxic nature of the samples. Changes in gene expression of cyp19a1b (P < 0.001), but not of cyp1a and vtg1 as measured by qRT-PCR, suggested oestrogenic impact of the industrial contaminated sediment on the steroidogenic pathway of the fish. While aromatase played a significant role, more specifically for both the sites 1 and 3 km downstream, the upper layers (0–5 cm) showed higher upregulation of cyp19a1b compared to the reference sediment located 9 km upstream from the mill. There was a significantly higher upregulation of cyp19a1b (P = 0.001) by 0–5 cm compared to the that at the 20–30-cm layer at the 1-km site, indicating an oestrogenic impact of recent surface sediment near the mill compared to that in the site further downstream.
These results highlight the toxic potential of the pulp mill-contaminated sediment to aquatic biota, indicated by oestrogen-responsive cyp19a1b regulation in an immature fish (20dpfZF).
Keywordscyp19a1b Oestrogenicity PPME qRT-PCR vtg1 Whole-sediment bioassay Zebrafish
We thank Dr. Aarno Karels for help with collection of sediments in Southern Lake Saimaa. The authors also thank Mervi Koistonen for help with sediment processing and experimentation, and Dr. Eeva-Riikka Vehniäinen for numerous technical advices. Dr. Tuula Sinisalo is thanked for help with the TOC analysis. We would also like to thank the anonymous reviewers for their valuable suggestions to strengthen this paper.
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