Description: Chemical weapons dumped into the sea after World War II possess growing concern for the marine environment; metal shells of different chemical munitions lying on the bottom are severely corroded and dangerous contents pollute the sediments. Chemical warfare agents (CWAs), such as mustard gas and various arsenic-based compounds (e.g., Clark I and Adamsite) and their degradation products have been detected in noticeable concentrations in sediments at the major dumping sites at the Baltic Sea. Blue mussel caging approach was applied to assess environmental impact of thousands of tons of CWAs at the main dumping site at the Bornholm Basin. Due to the patchy occurrence of the CWAs in the sediments mussel caging method was chosen to deploy the organisms exactly at sites where high CWA concentrations were detected in sediments and to one reference site. Biomarkers representing different biological functions including antioxidant defence, biotransformation, neurotoxicity, lysosomal membrane stability, geno- and cytotoxicity, cellular energy allocation and condition index were investigated. Moreover, tissue concentrations of different CWAs and the possible metabolic derivatives of these compounds were analysed in mussels together with “classical” contaminants, such as polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) and trace metals. Further, mussel cages were equipped with salinity, temperature and oxygen sensors to follow the fluctuations in the environmental parameters during the two and half month caging time. Environmental parameters indicated mixing of the near bottom water in the lower water layer where the cages were deployed (at 60 meters). Significant biomarker responses were observed at the two contaminated sites compared to the reference site indicating CWA induced effect on molecular and cellular level; however the possible anaerobic conditions and lower food availability in deep waters led to decreased bioenergetic status of the caged mussels. Results of the still on-going chemical analysis will be employed together with the biomarker responses and environmental data in the multi-level integrated impact assessment of the area.

Description: After World War II, large amounts of chemical weapons stored on German territory were dumped in the Baltic Sea by order of the allied forces . In addition of being a cheap method of disposal, the belief was that the vast amounts of waters in the oceans would neutralize and absorb the dangerous substances. At least 40.000 tonnes of chemical munitions containing an estimated 13.000 tonnes of chemical warfare agents were dumped in the Baltic Sea, primarily in the Bornholm Basin. Other official dumping sites were the Little Belt area and the Gotland Deep. There is evidence, however, that also other areas besides the offical CWA dumpsides are contaminated by chemical warfare agents (CWA). CWA were dumped as artillery shells, aircraft bombs or in containers; partly entire ships loaded with munitions were sunk. Today, munitions are in different stages of decomposition. Metal shells are corroding and contents are leaking into the environment at a rate that has not been measured so far, posing a possible risk for the Baltic Sea ecosystem. In previous studies, several CWAs of major concern for biota, such as inorganic arsenic and organo-arsenic compounds, have been found in the sediments within and around dumpsites (Missiaen et al. 2010). Unaware of this risk, human sea-bottom activities, such as bottom trawling, constructions of pipelines and cables as well as windfarms are increasingly claiming space within the contaminated areas. The aim of the present study was to increase knowledge on the bioavailability and biological effects of CWAs on fish, using a suite of biomarkers in an integrated approach.

Description: Blue mussels (Mytilus trossulus) were transplanted in cages for three months in two Swedish coastal areas in the Bothnian Sea (northern Baltic Sea) to investigate the interactions between analysed environmental chemicals and biological responses. A wide array of biological parameters (biomarkers) including antioxidant and biotransformation activity, geno-, cyto- and neurotoxic effects, phagocytosis, bioenergetic status and heart rate were measured to detect the possible effects of contaminants. Integrated Biomarker Response index and Principal Component Analysis performed on the individual biological response data were able to discriminate between the two study areas as well as the contaminated sites from their respective local reference sites. The two contaminated sites outside the cities of Sundsvall (station S1) and Gävle (station G1) were characterised by different biomarker response patterns. Mussels at station S1 showed a low condition index, increased heart rate recovery time and phagocytosis activity coinciding with the highest tissue concentrations of some trace metals, polycyclic aromatic hydrocarbons and organotins. At station G1 the highest organochlorine pesticide concentration was recorded as well as elevations in glutathione S-transferase activity, thiamine content and low lysosomal membrane stability. Significant variability in the geno- and cytotoxic responses and bioenergetic status was also observed at the different caging stations. The results obtained suggest that different chemical mixtures present in the study areas cause variable biological response patterns in organisms.

Description: CHEMSEA, Chemical Munitions Search and Assessment, is a flagship project of the Baltic Sea Region Program and is partly financed by the European Union. The main focus of the project is to locate the dumped chemical warfare agents (CWAs) and sample the surrounding environment to assess the possible threat. The project lasts from the fall 2011 until 2014. CHEMSEA is a transnational collaboration including project partners and associated organizations, including governmental agencies and international organizations. Ocean waters are in a constant flux, making the study of effect of CWAs on fish and other marine biota a challenging task. For the evaluation of the risks of dumped CWAs, triphenylarsine (TPA), sulphur mustard (H), Adamsite (DM) and Clark I (DA) are thought to pose the highest realistic risk to marine biota. The exact effects of these chemicals are not known and no information is available for the detoxification rates of CWAs in fish tissues or other marine organism, such as mussels. During CHEMSEA project, cod and mussels were chosen for chemical analysis. Cod was sampled from different sites of Baltic Sea, covering official and suspected dumpsites as well as reference sites in the western and eastern Baltic Sea. Mussels were studied both in situ caging experiments and in vivo exposure experiments. For in situ experiment, cages were deployed at two different depths at two selected hotspot sites and one reference site in the Bornholm dumping area. Based on hydrographical data, the cages were placed at 35 m and 65 m depths at all stations. The poor oxygen conditions prevailing in the main CW dumping area made caging closer to the sea bottom unfeasible. In vivo experiment, mussels were exposed to mixtures of the arsenic-containing CWAs DA, DM and the tear gas α-chloroacetophenone. The main aim was to evaluate biological responses in mussels induced by CWA mixtures at environmentally relevant concentrations. For chemical analysis, cod urine, bile and muscle tissues were chosen in addition to the whole blue mussels from caging experiments and exposure studies. Sulphur mustard hydrolyses quickly into thiodiglycol (TDG) in aqueous environment. TDG was analysed from cod urine and bile and also from whole mussels. TDG could also be in its oxidised form as thiodiglycol sulfoxide so it was reduced to TDG at the beginning of sample preparation. Analyses were performed using combined gas chromatography–tandem mass spectrometry (GC–MS/MS) and the TDG was detected as its heptafluorobutyrylimidazole (HBFI) derivative. DM, DA and TPA were analysed as their oxidation products using liquid chromatography–tandem mass spectrometry (LC–MS/MS) from cod muscle and mussel samples. The sample preparation for these analyses is laborious and contains homogenization, multiple extractions and filtration. TDG was detected from notable amount of fish urine samples as background level and from few samples as higher level. For arsenic-containing compounds, only oxidized TPA was found from one fish muscle sample. Nothing was detected from caged mussels. However, high concentrations of oxidized DM and DA were measured from exposed mussels. Detailed results and conclusions will be discussed.