Ecotoxicological assessment of sediment, suspended matter and water samples in the upper Danube River. A pilot study in search for the causes for the decline of fish catches

GOALS, SCOPE AND BACKGROUND: Fish populations, especially those of the grayling (Thymallus thymallus), have declined over the last two decades in the upper Danube River between Sigmaringen and Ulm, despite intensive and continuous stocking and improvement of water quality since the 1970s. Similar problems have been reported for other rivers, e.g. in Switzerland, Great Britain, the United States and Canada. In order to assess if ecotoxicological effects might be related to the decline in fish catch at the upper Danube River, sediment, suspended matter and waste water samples from sewage treatment plants were collected at selected locations and analyzed in a bioanalytical approach using a battery of bioassays. The results of this pilot study will be used to decide if a comprehensive weight-of-evidence study is needed. METHODS: Freeze-dried sediments and suspended particulate matters were extracted with acetone in a Soxhlet apparatus. Organic pollutants from sewage water were concentrated using XAD-resins. In order to investigate the ecotoxicological burden, the following bioassays were used: (1) neutral red assay with RTL-W1 cells (cytotoxicity), (2) comet assay with RTL-W1 cells (genotoxicity), (3) Arthrobacter globiformis dehydrogenase assay (toxicity to bacteria), (4) yeast estrogen screen assay (endocrine disruption), (5) fish egg assay with the zebrafish (Danio rerio; embryo toxicity) and (6) Ames test with TA98 (mutagenicity). RESULTS AND DISCUSSION: The results of the in vitro tests elucidated a considerable genotoxic, cytotoxic, mutagenic, bacteriotoxic, embryotoxic and estrogenic burden in the upper Danube River, although with a very inhomogeneous distribution of effects. The samples taken from Riedlingen, for example, induced low embryo toxicity, but the second highest 17beta-estradiol equivalent concentration (1.8 ng/L). Using the fish egg assay with native sediments, a broad range of embryotoxic effects could be elucidated, with clear-cut dose-response relationships for the embryotoxic effects of contaminated sediments. With native sediments, embryotoxicity was clearly higher than with corresponding pore waters, thus corroborating the view that--at least for fish eggs--the bioavailability of particle-bound lipophilic substances in native sediments is higher than generally assumed. The effect observed most frequently in the fish egg assay was a developmental delay. A comparison of our own results with locations along the rivers Rhine and Neckar demonstrated similar or even higher ranges of ecotoxicological burdens in the Danube River. CONCLUSIONS: The complex pattern of ecotoxicological effects caused by environmental samples from the Danube River, when assessed in an in vitro biotest battery using both acute and more specific endpoints, showed that integration of different endpoints is essential for appropriate hazard assessment. Overall, the ecotoxicological hazard potential shown has indeed to be considered as one potential reason for the decline in fish catches at the upper Danube River. However, based on the results of this pilot study, it is not possible to elucidate that chemically induced alterations are responsible for the fish decline. RECOMMENDATIONS AND PERSPECTIVES: In order to confirm the ecological relevance of the in vitro results for the situation in the field and especially for the decline of the grayling and other fishes, further integrated investigations are required. For linking the weight of evidence obtained by in vitro assays and fish population investigations, the application of additional, more specific biomarkers (e.g. vitellogenin induction, EROD and micronucleus assay) has been initiated in fish taken from the field as well as in situ investigations.