Effects on food intake of Gammarus spp. after exposure to PFBA in very low concentrations.

Per- and polyfluoroalkyl substances (PFAS) are a group of thousands of highly persistent anthropogenic chemicals widely used in many industries. Therefore, they are, ubiquitously present in various types of environments. Despite their omnipresence, ecotoxicological studies of most PFAS are scarce, and those available often assess the effects of long chain PFAS. In this study, we present the results of an exposure experiment in which wild aquatic amphipod Gammarus spp. was exposed to the short chain perfluorinated substance perfluorobutanoic acid (PFBA) at very low and environmentally relevant concentrations of 0, 10 and 100 ng/L. The exposure lasted for 12 days, and food intake and non-reproductive behavior were analyzed. Exposure to 10 and 100 ng/L PFBA resulted in a lower consumption of food during exposure but no effect on behavior was found.

Persistence of norfluoxetine in marine mussels.

The concentration of pharmaceuticals in coastal waters is tending towards increasing due to a shift of the human population into coastal zones. In parallel, the number of prescriptions of antidepressants, mainly selective serotonin reuptake inhibitors (SSRI), is constantly growing. Most of the SSRI is metabolised into active compounds; for instance, norfluoxetine (NFLU) is the main active metabolite of fluoxetine. In this study, we tested the bioaccumulation and depuration of NFLU in Mytilus trossulus at two environmentally relevant concentrations (100 and 500 ng/L, after six days of exposure and five days of depuration at 10 °C). The concentration of NFLU in the mussels' tissue seems not to be directly proportional to the exposure concentration. The levels of NFLU in the mussels' tissues after the depuration period were comparable to the levels detected at the end of exposure. This indicates that NFLU is not efficiently removed by the mussels and points to a potential risk for consumers of such marine organisms.

Somatic evolution of marine transmissible leukemias in the common cockle, Cerastoderma edule.

Transmissible cancers are malignant cell lineages that spread clonally between individuals. Several such cancers, termed bivalve transmissible neoplasia (BTN), induce leukemia-like disease in marine bivalves. This is the case of BTN lineages affecting the common cockle, Cerastoderma edule, which inhabits the Atlantic coasts of Europe and northwest Africa. To investigate the evolution of cockle BTN, we collected 6,854 cockles, diagnosed 390 BTN tumors, generated a reference genome and assessed genomic variation across 61 tumors. Our analyses confirmed the existence of two BTN lineages with hemocytic origins. Mitochondrial variation revealed mitochondrial capture and host co-infection events. Mutational analyses identified lineage-specific signatures, one of which likely reflects DNA alkylation. Cytogenetic and copy number analyses uncovered pervasive genomic instability, with whole-genome duplication, oncogene amplification and alkylation-repair suppression as likely drivers. Satellite DNA distributions suggested ancient clonal origins. Our study illuminates long-term cancer evolution under the sea and reveals tolerance of extreme instability in neoplastic genomes.

Oxytetracycline-induced inflammatory process without oxidative stress in blue mussels Mytilus trossulus.

Potentially harmful compounds including pharmaceuticals are commonly found in marine waters and sediments. Amongst those, antibiotics and their metabolites are detected worldwide in various abiotic (at concentrations as high as µg/L) and biotic matrices at ng/gram of tissue, posing a risk to non-target species exposed to them such as blue mussels. Amongst those, oxytetracycline (OTC) belongs to the most detected antibiotics in the marine environment. In this work, we concentrated on studying the potential induction of oxidative stress, activation of cellular detoxification processes (including Phase I and Phase II xenobiotic biotransformation enzymes) and multixenobiotic resistance pumps (Phase III) as well as changes in the aromatisation efficiency in Mytilus trossulus exposed to 100 µg/L OTC. Our results show that 100 µg/L OTC concentration did not provoke cellular oxidative stress and did not affect the expression of genes involved in detoxification processes in our model. Moreover, no effect of OTC on aromatisation efficiency was found. Instead, phenoloxidase activity measured in haemolymph was significantly higher in OTC exposed mussels than in those from the control (30.95 ± 3.33 U/L and 17.95 ± 2.75 U/L, respectively). OTC exposed mussels were also characterised by a tissue-dependant activation of major vault protein (MVP) gene expression (1.5 times higher in gills and 2.4 times higher in the digestive system) and a decreased expression of the nuclear factor kappa B-a (NF-κB) gene (3.4 times lower in the digestive system) when compared to those from the control. Additionally, an elevated number of regressive changes and inflammatory responses in tissues such as gills, digestive system and mantle (gonads) was observed underlining the worsening of bivalves' general health. Therefore, instead of a free-radical effect of OTC, we for the first time describe the occurrence of typical changes resulting from antibiotic therapy in non-target organisms like M. trossulus exposed to antibiotics such as OTC.

Bivalves Transmissible Neoplasia: Biochemical Aspects of Contagious Cancer in a Clam Macoma Balthica.

BACKGROUND/AIMS: Occurring in marine invertebrates infectious haemic neoplasia (bivalves transmissible neoplasia, BTN) arises from genome instabilities leading to multilevel malfunctions and unregulated cell division of presumably haemocyte precursors. As its biochemical characterisation remains unknown, we here present the first data describing selected aspects of the physiology and biochemistry of the disease a in model clam Macoma balthica. We chose free amino acids (FAA) composition, mitochondrial respiration and enzymatic activity, oxidative stress enzymes activities and corticosteroids profile as markers of this contagious cancer. METHODS: Selected markers were measured in neoplastic and healthy clams and two tissue types, haemolymph and solid tissue. FAA composition was assessed in the haemolymph samples using high performance liquid chromatography-mass spectrometry (LC/MS). Mitochondrial respiration analysis was performed on haemocytes using oxygen electrodes integrated system Seahorse XFp. Mitochondrial enzymes activities were measured using spectrophotometry (cytochrome oxidase, COX) and commercial kit (succinate dehydrogenase, SDH). Total Antioxidant Capacity (TAC), Acetylocholinesterase (AChE), Protein Carbonyl Content (CBO) and Malondialdehyde (MDA) levels were measured in the solid tissue using analytical kits, and glutathione (GSH) was measured spectrophotometrically. Corticosteroids profile, measured in the solid tissue, was obtained with Liquid Chromatography-Electrospray Ionization-Mass Spectrometry (HPLC-ESI-MS/MS) technique. RESULTS: In both clam groups nine FAAs were detected with Asp, Glu, Pro, Ser constituting over 90% of total FAA content. Significantly higher Gln level was detected in BTN positive clams. In neoplastic clams, an impairment of mitochondrial metabolism was observed as a decrease in mitochondrial oxygen consumption and lower cytochrome c oxidase activity. In the neoplastic clams significantly higher concentration of low molecular weight antioxidants was found. Finally, we report high level of corticosterone and lower levels of dehydrocorticosterone, cortisol and cortisone in healthy clams and elevated cortisol level in BTN individuals. CONCLUSION: Neoplastic clams are characterized by altered mitochondrial metabolism, with a potential key role of glutamine (Gln) in cancer cells energy production. Despite low aerobic respiration, BTN cells have efficient antioxidative response to elevated concentration of ROS. Elevated cortisol level in BTN-positive clams may indicate an important role of this corticosteroid in cancer biochemistry. Thus, we here provide the first results of selected physiological and biochemical aspects of BTN, making an important step in studying cancer epidemiology in wildlife.

A multi-biomarker approach to assess toxicity of diclofenac and 4-OH diclofenac in Mytilus trossulus mussels - First evidence of diclofenac metabolite impact on molluscs.

Although the presence of pharmaceuticals in the environment is an issue widely addressed in research over the past two decades, still little is known about their transformation products. However, there are indications that some of these chemicals may be equally or even more harmful than parent compounds. Diclofenac (DCF) is among the most commonly detected pharmaceuticals in the aquatic environment, but the potential effects of its metabolites on organisms are poorly understood. Therefore, the present study aimed to evaluate and compare the toxicity of DCF and its metabolite, 4-hydroxy diclofenac (4-OH DCF), in mussels using a multi-biomarker approach. Mytilus trossulus mussels were exposed to DCF and 4-OH DCF at 68.22 and 20.85 µg/L (measured concentrations at day 0), respectively, for 7 days. In our work, we showed that both tested compounds have no effect on most of the enzymatic biomarkers tested. However, it has been shown that their action can affect the protein content in gills and also be reflected through histological markers. ENVIRONMENTAL IMPLICATION: Studies in recent years clearly prove that pharmaceuticals can negatively affect aquatic organisms. In addition to parent compounds, metabolites of pharmaceuticals can also be a significant environmental problem. In the present work, the effects of diclofenac and its main metabolite, 4-hydroxy diclofenac, on marine mussels were evaluated. Both compounds showed negative effects on mussels, which was primarily observed through histological changes. The present study therefore confirms that not only diclofenac, but also its main metabolite can have negative effects on aquatic organisms.

Long-term stability of diclofenac and 4-hydroxydiclofenac in the seawater and sediment microenvironments: Evaluation of biotic and abiotic factors.

Studies in recent years have shown that significant amounts of diclofenac (DCF) and its metabolites are present in marine coastal waters. Their continuous flow into the environment may be associated with numerous negative effects on both fauna and flora. Although more and more is known about the effects of pharmaceuticals on marine ecosystems, there are still many issues that have not received enough attention, but are essential for risk assessment, such as long term stability. Furthermore, interaction of pharmaceuticals with sediments, which are inhabited by rich microbial, meiofaunal and macrobenthic communities need investigation. Therefore, we undertook an analysis of the stability of DCF and its metabolite, 4-hydroxy diclofenac, in seawater and sediment collected from the brackish environment of Puck Bay. Our 29-day experiment was designed to gain a better understanding of the fate of these compounds under experimental conditions same as near the seafloor. Diclofenac concentration decreased by 31.5% and 20.4% in the tanks with sediment and autoclaved sediment, respectively during 29-day long experiment. In contrast, the concentration of 4-OH diclofenac decreased by 76.5% and 90.2% in sediment and autoclaved sediment, respectively. The concentration decrease of both compounds in the sediment tanks resulted from their sorption in the sediment and biodegradation. Obtained results show that marine sediments favour DCF and 4-OH DCF removal from the water column.

Horizontal transmission of disseminated neoplasia in the widespread clam Macoma balthica from the Southern Baltic Sea.

Disseminated neoplasia (DN) is one of the most challenging and unrecognised diseases occurring in aquatic fauna. It has been diagnosed in four bivalve species from the Gulf of Gdansk (Southern Baltic Sea) with the highest frequency in Macoma balthica (formerly Limecola balthica), reaching up to 94% in some populations. The aetiology of DN in the Baltic Sea has not yet been identified, with earlier studies trying to link its occurrence with environmental pollution. Taking into account recent research providing evidence that DN is horizontally transmitted as clonal cells between individuals in some bivalve species, we aimed to test whether DN is a bivalve transmissible neoplasia (BTN) in the population of M. balthica from the Gulf of Gdansk highly affected with cancer. We examined mitochondrial cytochrome c oxidase I (mtCOI) and elongation factor 1α (EF1α) sequences of genomes obtained from haemolymph and tissues of neoplastic and healthy individuals. Sequence analysis resulted in detection of an independent transmissible cancer lineage occurring in four neoplastic clams that is not present in healthy animals. This study describes the first case of BTN in the clam M. balthica (MbaBTN), providing further insights for studies on this disease.

Presence of pharmaceuticals and their metabolites in wild-living aquatic organisms - Current state of knowledge.

In the last decades an increasing number of studies has been published concerning contamination of aquatic ecosystems with pharmaceuticals. Yet, the distribution of these chemical compounds in aquatic environments raises many questions and uncertainties. Data on the presence of selected pharmaceuticals in the same water bodies varies significantly between different studies. Therefore, since early 1990 s, wild organisms have been used in research on environmental contamination with pharmaceuticals. Indeed, pharmaceutical levels measured in biological matrices may better reflect their overall presence in the aquatic environments as such levels include not only direct exposure of a given organisms to a specific pollutant but also processes such as bioaccumulation and biomagnification. In the present paper, data concerning occurrence of pharmaceuticals in aquatic biota was reviewed. So far, pharmaceuticals have been studied mainly in fish and molluscs, with only a few papers available on crustaceans and macroalgae. The most commonly found pharmaceuticals both in freshwater and marine organisms are antibiotics, antidepressants and NSAIDS while there is no information about the presence of anticancer drugs in aquatic organisms. Furthermore, only single studies were conducted in Africa and Australia. Hence, systematization of up-to-date knowledge, the main aim of this review, is needed for further research targeting.

Exposure of Mytilus trossulus to diclofenac and 4'-hydroxydiclofenac: Uptake, bioconcentration and mass balance for the evaluation of their environmental fate.

Diclofenac (DIC) is one of the most widely consumed drugs in the world, and its presence in the environment as well as potential effects on organisms are the subject of numerous recent scientific works. However, it is becoming clear that the risk posed by pharmaceuticals in the environment needs to be viewed more broadly and their numerous derivatives should also be considered. In fact, already published results confirm that the transformation products of NSAIDs including DIC may cause a variety of potentially negative effects on marine organisms, sometimes showing increased biological activity. To date, however, little is known about bioconcentration of DIC and DIC metabolites and the role of sex in this process. Therefore, the present study for the first time evaluates sex-related differences in DIC bioconcentration and estimates bioconcentration potential of DIC metabolite, 4-OH DIC, in the Mytilus trossulus tissues. In the experiment lasting 7 days, mussels were exposed to DIC and 4-OH DIC at concentrations 68.22 and 20.85 µg/L, respectively. Our study confirms that DIC can be taken up by organisms not only in its native form, but also as a metabolite, and metabolised further. Furthermore, in the present work, mass balance was performed and the stability of both studied compounds under experimental conditions was analysed. Obtained results suggest that DIC is more stable than its derivative under the tested conditions, but further analyses of the environmental fate of these compounds are necessary.

Cellular level response of the bivalve Limecola balthica to seawater acidification due to potential CO2 leakage from a sub-seabed storage site in the southern Baltic Sea: TiTank experiment at representative hydrostatic pressure.

Understanding of biological responses of marine fauna to seawater acidification due to potential CO2 leakage from sub-seabed storage sites has improved recently, providing support to CCS environmental risk assessment. Physiological responses of benthic organisms to ambient hypercapnia have been previously investigated but rarely at the cellular level, particularly in areas of less common geochemical and ecological conditions such as brackish water and/or reduced oxygen levels. In this study, CO2-related responses of oxygen-dependent, antioxidant and detoxification systems as well as markers of neurotoxicity and acid-base balance in the Baltic clam Limecola balthica from the Baltic Sea were quantified in 50-day experiments. Experimental conditions included CO2 addition producing pH levels of 7.7, 7.0 and 6.3, respectively and hydrostatic pressure 900 kPa, simulating realistic seawater acidities following a CO2 seepage accident at the potential CO2-storage site in the Baltic. Reduced pH interfered with most biomarkers studied, and modifications to lactate dehydrogenase and malate dehydrogenase indicate that aerobiosis was a dominant energy production pathway. Hypercapnic stress was most evident in bivalves exposed to moderately acidic seawater environment (pH 7.0), showing a decrease of glutathione peroxidase activity, activation of catalase and suppression of glutathione S-transferase activity likely in response to enhanced free radical production. The clams subjected to pH 7.0 also demonstrated acetylcholinesterase activation that might be linked to prolonged impact of contaminants released from sediment. The most acidified conditions (pH 6.3) stimulated glutathione and malondialdehyde concentration in the bivalve tissue suggesting potential cell damage. Temporal variations of most biomarkers imply that after a 10-to-15-day initial phase of an acute disturbance, the metabolic and antioxidant defence systems recovered their capacities.

Bioaccumulation of phenolic endocrine disruptors in the clam Rangia cuneata: Storage in shells and influence of size and sex.

This study aimed to investigate the sequestration of phenolic endocrine disrupting compounds (EDCs) such as bisphenol A (BPA), 4-t-octylphenol (4-t-OP), and 4-nonylphenol (4-NP) in the shells of the mature clam Rangia cuneata from the Vistula Lagoon (southern Baltic Sea) and to determine the influence of sex and shell length on bioaccumulation of these contaminants. Even though there is broad interest in EDCs influences on aquatic organisms, these basic parameters are poorly understood, yet necessary for assessing environmental risks for clams. Average proportions of the total body burden (ng/individual) deposited in shells of R. cuneata were more than 70% for BPA and 4-NP and up to 32% for 4-t-OP. These results indicate that shell storage can be an important route for elimination of specific EDCs. Relationships between EDCs concentrations and the size and sex of R. cuneata indicate that females and large individuals experience greater exposures to the adverse effects of these pollutants than males and smaller clams. This effect could have significant impacts on population ecology and ultimately affect the entire ecosystem, in which bivalves play an important role. In the context of using clams to assess water pollution, the co-variation of EDCs concentrations with the size and sex of bivalves could influence the quality of monitoring data, unless accounted for in sampling design and data analysis.

Effects of environmentally relevant concentrations of diclofenac in Mytilus trossulus.

The presence of pharmaceuticals in the marine environment is a growing problem of global importance. Although awareness of the significance of this issue is increasing, many questions related to the ecotoxicology of pharmaceuticals remain unclear. Diclofenac is one of the drugs most commonly detected in the marine environment and its potential toxicity has been previously highlighted, thus its impact on organisms deserves a special attention. Therefore, in this study, a thorough analysis of the effects of diclofenac on a condition and tissue level of a model representative of marine invertebrates - Mytilus trossulus - was performed. During the 25-day experiment, divided into exposure and depuration phases, bivalves were exposed to two environmentally relevant drug concentrations of 4 and 40 µg/L. The study showed that mussels absorb diclofenac in their tissues and the highest recorded concentration was 1.692 µg/g dw on day 8. Moreover, the content of diclofenac metabolites (4-OH and 5-OH diclofenac) was also examined, but they were not detected either in water or in tissues. Although exposure to low diclofenac concentrations did not significantly affect the condition index of organisms, changes in numerous histopathological parameters were noted. Performed histological examination provided additional valuable information on the influence of drugs on the functioning of invertebrates. Nevertheless, applicability of histopathological techniques in ecotoxicology of drugs requires additional evaluation in future studies.

Toxic effects of NSAIDs in non-target species: A review from the perspective of the aquatic environment.

The presence of pharmaceuticals in the aquatic environment, both in marine and freshwater reservoirs, is a major concern of global environmental protection. Among the drugs that are most commonly used, NSAIDs tend to dominate. Currently, being aware of the problem caused by drug contamination, it is extremely important to evaluate the scale and the full spectrum of its consequences, from short-term to long-term effects. The influence on non-target aquatic animals can take place at many levels, and the effects can be seen both in behaviour and physiology, but also in genetic alterations or reproduction disorders, affecting the development of entire populations. This review summarises all the advances made to estimate the impact of NSAIDs on aquatic animals. Multicellular animals from all trophic levels, inhabiting both inland waters, seas and oceans, have been considered. Particular attention has been paid to chronic studies, conducted at low, environmentally-relevant concentrations, to estimate the real effects of the present pollution. The number of such studies has indeed increased in recent years, allowing for a better insight into the possible consequences of pharmaceutical pollution. It should be stressed, however, that our knowledge is still limited to a few model species, while there are many groups of organisms completely unexplored regarding the effects of drugs. Therefore, the main aim of this paper was to summarise the current state of knowledge on the toxicity of NSAIDs in aquatic animals, also identifying important gaps and major issues requiring further analysis.

Mytilidae as model organisms in the marine ecotoxicology of pharmaceuticals - A review.

Growing production and consumption of pharmaceuticals is a global problem. Due to insufficient data on the concentration and distribution of pharmaceuticals in the marine environment, there are no appropriate legal regulations concerning their emission. In order to understand all aspects of the fate of pharmaceuticals in the marine environment and their effect on marine biota, it is necessary to find the most appropriate model organism for this purpose. This paper presents an overview of the ecotoxicological studies of pharmaceuticals, regarding the assessment of Mytilidae as suitable organisms for biomonitoring programs and toxicity tests. The use of mussels in the monitoring of pharmaceuticals allows the observation of changes in the concentration and distribution of these compounds. This in turn gives valuable information on the amount of pharmaceutical pollutants released into the environment in different areas. In this context, information necessary for the assessment of risks related to pharmaceuticals in the marine environment are provided based on what effective management procedures can be developed. However, the accumulation capacity of individual Mytilidae species, the bioavailability of pharmaceuticals and their biological effects should be further scrutinized.

Aromatisation of steroids in the bivalve Mytilus trossulus.

In this study, we demonstrated the presence of the enzymatic complex able to perform aromatization (estrogen synthesis) in both, the microsomal and mitochondrial fractions of gills and gonads from Mytilus trossulus. Based on in vitro experiments, we highlighted the importance of temperature as the limiting factor of aromatisation efficiency (AE) in mussels. After testing range of temperatures (4-23 °C), the highest AE was found during incubation at 8 °C and pH 7.6 (41.66 pmol/h/mg protein in gills and 58.37 pmol/h/mg protein in gonads). The results were confirmed during field studies where the most efficient aromatisation occurred in bivalves collected in spring while the least effective in those collected in winter. During in vitro studies, AE turned out to be more intensive in female gonads than in male gonads. The process was also more intensive in mitochondrial fraction than in microsomal one (62.97 pmol/h/mg protein in male gills and 73.94 pmol/h/mg protein in female gonads). Enzymatic complex (aromatase-like enzyme) catalysing aromatisation in mussels was found to be insensitive to inhibitory effect of selective inhibitors of mammalian aromatase such as letrozole and anastrazole, suggesting its different structure from vertebrate aromatase. Further in vivo studies using 13C-labeled steroids at 8 °C temperature window confirmed that bivalves are able to uptake testosterone and androstenedione from the ambient environment and metabolise them to estrone and 17ß-estradiol thus confirming endogenous estrogen' synthesis.

Multi biomarker analysis of pollution effect on resident populations of blue mussels from the Baltic Sea.

Anthropogenic pollution including metals, petroleum, toxins, nutrients and many others is a growing problem in the marine environment. These are important factors altering the environment and by that the fate of many local populations of marine organisms. The aim of this study was to assess the impact of selected point pollution sources on resident populations of the blue mussel (Mytilus edulis trossulus) in the Baltic Sea using multiple biomarker approach. The study used a nested sampling scheme in which sites from reference (REF) habitats are geographically paired with selected sites from sewage treatment plants (STP) and harbors (HAR). The results showed that mussels from harbors had a higher frequency of histological abnormalities in the digestive gland compared to mussels from sewage effluent affected areas and reference sites. However these mussels together with mussels from STPs had higher lipid content, body mass index (BMI) and gonado-somatic index (GSI) compared to mussels from reference sites. A marked spatial variability was found with a stronger toxicity of ambient environment affecting resident mussel populations in the Gulf of Gdansk area, while an opposite pattern was found in Tvärminne area. Yet the blue mussels sampled in the Gulf of Gdansk were characterized by the highest GSI and BMI values compared to Askö and Tvärminne populations. No differences in analyzed biomarker response related to species identity, measured by a species-specific genetic marker, were found indicative of strong genetic introgression in the Baltic Proper.

Changes in steroid profiles of the blue mussel Mytilus trossulus as a function of season, stage of gametogenesis, sex, tissue and mussel bed depth.

This paper describes changes in the content of free steroid hormones e.g. testosterone (T), estradiol-17ß (E2), estrone (E1) and estriol (E3) of Mytilus trossulus from the southern Baltic Sea as a function of season, stage of gametogenesis, sex, tissue (gonadal and somatic) and depth. The highest levels of T, E2, E1 and E3 were found in mussels sampled in spring and summer while the lowest levels were found in winter. This pattern was stable and was seen in both sexes and tissues in mussels from both mussel beds. The spring and summer peaks in steroid levels (SL) coincided with advanced levels of gametogenesis (the highest gonadal index, GI) of our model species. But, the lowest GI (autumn) and the lowest steroids content (winter) did not overlap. Instead, water temperature increase was followed by increase of SL and vice versa. This suggests that steroids may not be actively involved in the early stages of gamete development and does not preclude them from potentially being involved as endogenous modulators in the final stages of reproduction (e.g. spawning). Hence, observed fluctuations in SL in our model species are unlikely to be caused by reproductive cycle but are rather of unknown nature, likely linked with environmental conditions. Sex-related differences in steroid content included estrogen domination in females and androgen domination in males. A trend towards higher level of steroids in gills than in gonads was found, supporting the hypothesis about an exogenous origin of steroids in bivalves. However, based on the present results, we cannot exclude the possibility that these steroids have both an endogenous and exogenous origin.

Elevated gonadal atresia as biomarker of endocrine disruptors: Field and experimental studies using Mytilus trossulus (L.) and 17-alpha ethinylestradiol (EE2).

In the present work we compared the type and frequency of gonadal abnormalities among Mytilus trossulus populations from the Gulf of Gdansk, Baltic Sea. Observed histopathologies were grouped as regressive changes (RC, gonadal atresia (GA) and regression (GR)), progressive changes (PC, gonadal tumors) and intersex. Sex-based and spatial differences in frequency of RC were found, with the highest frequency of RC and PC found in mussels from polluted station B followed by mussels from station A located near a purification plant outlet. Bivalves from the reference area had the lowest frequency of RC. In order to confirm biomarker applicability of RC, an exposure experiment with model xenoestrogen 17α-ethinylestradiol (EE2) was performed. The exposure of M. trossulus to 50 and 500ngdL-1 of EE2 resulted in an increased frequency of gonadal regression and atresia, including melanized hemocytes infiltration in seminiferous tubules. We thus suggest that these changes can serve as biomarkers of endocrine disrupting compounds in biomonitoring studies.

Sewage treatment plant associated genetic differentiation in the blue mussel from the Baltic Sea and Swedish west coast.

Human-derived environmental pollutants and nutrients that reach the aquatic environment through sewage effluents, agricultural and industrial processes are constantly contributing to environmental changes that serve as drivers for adaptive responses and evolutionary changes in many taxa. In this study, we examined how two types of point sources of aquatic environmental pollution, harbors and sewage treatment plants, affect gene diversity and genetic differentiation in the blue mussel in the Baltic Sea area and off the Swedish west coast (Skagerrak). Reference sites (REF) were geographically paired with sites from sewage treatments plant (STP) and harbors (HAR) with a nested sampling scheme, and genetic differentiation was evaluated using a high-resolution marker amplified fragment length polymorphism (AFLP). This study showed that genetic composition in the Baltic Sea blue mussel was associated with exposure to sewage treatment plant effluents. In addition, mussel populations from harbors were genetically divergent, in contrast to the sewage treatment plant populations, suggesting that there is an effect of pollution from harbors but that the direction is divergent and site specific, while the pollution effect from sewage treatment plants on the genetic composition of blue mussel populations acts in the same direction in the investigated sites.