CHEMSEA conducted toxicity studies to investigate the biological uptake of CWA under varying environmental conditions. These tests included both laboratory activities and field measurements of concentrations and effects of CWA in representative fish species and benthic fauna caught in the dumping area. Effects in fish were studied using various biomarkers which serve to characterize health status and by utilizing data available from fish stock assessments and fish health databases.
Additionally, for a period of two months, CHEMSEA deployed specifically designed cages at Bornholm Basin - where the highest concentrations of chemical munitions have been measured - containing blue mussels. Through laboratory analyses, the accumulation of chemical substances in their tissues were then measured as well as their biological effects (which can be assessed through biomarkers for cytotoxicity, immunotoxicity, oxidative stress, bioenergetics or genotoxicity).
Macro and meiofauna
In the CHEMSEA project three sampling sites were selected to investigate the macro- and meiofauna commmunities: Bornholm Deep, Gotland Deep and Gdansk Deep. Fauna collected from these locations was compared with fauna from a reference area located between the study regions in an area of similar depth, i.e. deeper than 80m. In total six scientific cruises were conducted in different seasons between 2011 and 2013.
The first results show that macrofaunal and meiofaunal communities in CW dumping sites are very poor in terms of abundance and number of taxa in comparison with the reference area. A complete lack of any representatives of macrozoobenthos in all investigated dumping sites was noted. In total 48 genera of nematodes belonging to 21 families were noted in all investigated areas. A first Baltic Sea record of viviparous nematodes (Halomonhystera) was observed. All listed species are resistant to low oxygen concentrations in the external environment. The direct effects of CWA on benthic faunal communities, however, are difficult to determine due to hypoxic or even anoxic conditions. Although the lack of oxygen near the bottom at sampling sites located below 80-100 m depth is probably the main factor in determining the occurrence of living organisms, in combination with this, the low nematode density and diversity at dumping sites can also be regarded as the consequence of the higher toxicity levels in the sediments.
Cod (Gadus morhua L.)
Cod, the most abunding gadoid species in the Baltic Sea and one of major economic importance, is a demersal and bentho-pelagic species and thus is at risk of being in direct contact with dumped CWA. Some of its major spawning grounds are located in the deep basins at Bornholm and Gotland. FI, AWI, SYKE and NRC assessed the health status of cod at CW dumpsites and in reference areas in the Baltic Sea. Sampling took place during four campaigns between December 2011 and September 2013 onboard the fishery R/V Walther Herwig III. Cod was sampled by means of pelagic (at the dumpsites) and bottom trawling (at the reference sites), covering official and suspected dumpsites (Bornholm, Gotland, Belt Sea, Gdansk Deep) as well as a number of reference sites in the western (Arkona Sea) and eastern Baltic Sea (outside the Gulf of Gdansk). In addition to studies on grossly visible external and internal diseases and parasites and biometric characteristics carried out directly onboard the vessel, a large number of tissue samples were taken for subsequent analyses of contaminant concentrations as well as for the analysis of biochemical, physiological, pathological, neurotoxic and genotoxic responses to CWA exposure.
The data revealed no significant overall health effects in cod from CW dumpsites compared to the reference areas. However, when looking at single parameters some differences were noted, possibly reflecting responses to CWA exposure. As an example, tissue samples of the blood generating head kidneys of cod from 2011 showed more alterations and less compact tissue architecture in individuals caught at the dumping sites compared to those from the reference area. While the mean Fish Disease Index (FDI) value at the Bornholm dumpsite was high in 2011, it was significantly lower in 2012 and did not indicate differences between CWA dumpsites and the reference areas. Thus, it cannot be excluded that health risks posed by CWA vary on the short and the long term scales.
Furthermore, analyses at lower organisational levels, such as organ, tissue, cellular and subcellular levels using a suite of biomarkers revealed some stress responses in cod from CW dumpsites.
Blue mussels (Mytilus edulis L.)
Using the mussel as a model organism, the aim of the study was to use various biomarker methods representing different biological functions and levels of biological organisation to allow prediction of potential risks of dumped chemicals to aquatic organisms.
In the experiment, blue mussels (Mytilus spp.) were deployed at four sites with two cages at each site (35m and 65m depth). Four hundred mussels were placed in each cage. In addition, the cages were equipped with passive samplers to record the accumulation of selected organic contaminants and hydrography sensors with automatic loggers measuring temperature, salinity and oxygen. After retrieval of the cages, the exposed organisms were dissected onboard, all the samples were frozen and later on delivered to several laboratories. The samples were analyzed for accumulation of chemical substances and, even more importantly, their true biological effects (biomarkers).
First results show highest biomarker responses at both hotspot sites at 65 m. Markedly lower lysosomal membrane stability observed in 65 m cages at hotspot stations indicated significantly impaired haemocyte function compared to all other stations. A somewhat lower LMS value at the 65 m reference station compared to 35 m stations possibly shows the effect of increased general stress due to the e.g. lower oxygen at the 65 m. Environmental parameters recorded at those depths showed that the salinity and the concentration of dissolved oxygen fluctuated, indicating mixing of the near bottom water in the sub halocline water layer Thus, this makes it likely that mussels at 65 m were exposed to CWAs and other contaminants present in the sediment or near bottom water resulting in the observed biological effects. However, it has to be kept in mind that mussels were also exposed to lower oxygen concentrations that could, at least for a short period of time, induce the change from aerobic to anaerobic metabolism and contribute to the variation in the observed biological responses.
Water fleas (Daphnia magna)
Water fleas are also an indicator genus to test the effects of toxins on an ecosystem. For the CHEMSEA project water fleas were exposed at the Swedish Defence Research Agency (FOI) to slurries and water extracts from sediment samples from the Gotland Deep CW dumping site. The experiment was set up as a static acute toxicity test.
Two experiments were run on sediment extracts using Daphnia magna as test animal: testing the effect on survival and galactosidae activity (Daphnia IQ-test) and testing the effects on juvenile growth. The results of the laboratory experiments showed that the slurries of the sediments were not toxic to the animals. In the experiment on sediment extracts two of the extracts affected both the survival and the galactosidase activity. In the growth study one extract decreased the growth of the juveniles significantly. However, the analysis of the sediment samples indicates that none of the three toxic samples actually showed traces of compounds related to the presence of CWA. Which compound in the three sediments samples actually affected the water fleas is yet unknown.
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