Pharmaceutical drugs in the aquatic medium may pose significant risk to non-target organisms. In this study, the potential toxicity of a mixture of three compounds commonly detected in marine waters ...(ibuprofen, ciprofloxacin and flumequine) was assessed, by studying bioaccumulation, oxidative stress and neurotoxicity parameters (catalase CAT, superoxide dismutase SOD, glutathione reductase GR, glutathione S-transferase GST, lipid peroxidation LPO, glutathione peroxidase GPX, metallothionein MT and acetylcholinesterase AChE) in the clam Scrobicularia plana. Temporal evolution of selected endpoints was evaluated throughout an exposure period (1, 7 and 21 days) followed by a depuration phase. The accumulation of all drugs was fast, however clams showed the ability to control the internal content of drugs, keeping their concentration constant throughout the exposure and reducing their content after 7 days of depuration. The induction of biochemical alterations (SOD, CAT, LPO, MT, AChE) was observed in gills and digestive gland probably related to an imbalance in the redox state of clams as a consequence of the exposure to the drug mixture. These alterations were also maintained at the end of the depuration week when the high levels of SOD, CAT, GST and LPO indicated the persistence of oxidative stress and damage to lipids despite the fact that clams were no longer exposed to the mixture.
•Clams quickly accumulate all selected drugs and regulate the internal level over time.•Pharmaceuticals showed low bioconcentration potential for S. plana.•Drug mixture induces oxidative stress both in digestive gland and gills.•Antioxidant defense mechanisms were different depending on tissue and exposure dose.•Oxidative stress persists after one depuration week.
Linear alkylbenzene sulfonate (LAS) is a group of anionic surfactants employed in the formulation of laundry and cleaning products, with a global production rate of 4 million metric tons. Sediments ...from the Polish coast of the southern Baltic Sea were collected at ten stations. Total LAS concentrations, measured by high-performance liquid chromatography, were between 0.04 and 0.72 mg LAS·kg
−1
dry weight. Highest LAS concentrations were found in suspended matter collected from the Vistula River, sediment collected close to the Vistula River mouth and from the Gdańsk Deep, known as the depositional area. With the obtained environmental LAS concentrations, a risk assessment for this surfactant has been carried out, based on publicly available acute and chronic toxicity data in target organisms. The results indicated that LAS could pose a low risk for the existing benthic community applying worst case scenario assessment. This is the first time that levels of LAS have been measured in environmental samples of the southern Baltic Sea.
•Effects of CBZ, DCF and IBU exposure are assessed in the clam, Ruditapes philippinarum.•Antioxidant system is induced by drugs at early exposure period.•No oxidative stress is reported at the end of ...the exposure period.•Changes in biochemical responses can be related to a dual role of pharmaceuticals.
The presence of pharmaceuticals in the aquatic ecosystem has become a topic of growing interest in recent years. In this study, the marine clam Ruditapes philippinarum was exposed during 14 days to concentrations close to those found in the environment: (15 μg L−1) of carbamazepine (CBZ), diclofenac (DCF) and ibuprofen (IBU), three pharmaceuticals widely used worldwide and commonly found within the aquatic environment. Additionally, exposure was followed by a depuration phase (7 days). A battery of biomarkers (superoxide dismutase SOD, catalase CAT, glutathione reductase GR, total glutathione peroxidase T-GPx, glutathione transferase GST, lipid peroxidation LPO, acetylcholinesterase AChE and metallothionein MT) was evaluated throughout the exposure and depuration. The Integrated Biomarker Response index was calculated with all selected biomarkers and used as a complementary tool in the evaluation of the organisms’ health status. Exposure induced changes in the clams’ biochemical responses that led to the hypothesis of the harmful role of the pharmaceuticals resulting in negative effects (changes in enzyme activities, LPO and MT levels, related to ROS production) particularly after short-term exposure. However, the clams showed the ability to cope with these imbalances by recovering their general oxidative status by the end of exposure.
•RT-qPCR normalization in bivalve ecotoxicology is largely performed using single reference genes.•Less than 40% of reviewed studies experimentally validated reference gene stability.•The use of ...un-validated genes can introduce artefactual variance.•The use of more than one established validation algorithm is recommended.•Combining the ranks from multiple algorithms is not a suitable validation process.
The appropriate selection of reference genes for the normalization of non-biological variance in reverse transcription real-time quantitative PCR (RT-qPCR) is essential for the accurate interpretation of the collected data. The use of multiple validated reference genes has been shown to substantially increase the robustness of the normalization. It is therefore considered good practice to validate putative genes under specific conditions, determine the optimal number of genes to be employed, and report the method or methods used.
Under this premise, we assessed the current state of reference gene based normalization in RT-qPCR bivalve ecotoxicology studies (post 2011), employing a systematic quantitative literature review. A total of 52 papers met our criteria and were analysed for genes used, the use of multiple reference genes, as well as the validation method employed. We further critically discuss methods for reference gene validation based on a case study using copper exposed primary hemocytes from the marine bivalve Ruditapes philippinarum; including the established algorithms geNorm, NormFinder and BestKeeper, as well as the popular online tool RefFinder.
We identified that RT-qPCR normalization is largely performed using single reference genes, while less than 40% of the studies attempted to experimentally validate the expression stability of the genes used. 18s rRNA and β-Actin were the most popular genes, yet their un-validated use did introduce artefactual variance that altered the interpretation of the resulting data. Our findings further suggest that combining the results from multiple individual algorithms and calculating the overall best-ranked gene, as computed by the RefFinder tool, does not by default lead to the identification of the most suitable reference genes.
The individual and combined toxicities of acetaminophen, carbamazepine, diclofenac and ibuprofen have been examined in neonate nauplii (<24 h-old) of the harpacticoid copepod Tisbe battagliai. Based ...on acute toxicity data (LC50) obtained, diclofenac was the most toxic compound with an LC50 value of 9.5 mg·L−1; this is between 5 and 7 times lower than the LC50 value for acetaminophen, carbamazepine and ibuprofen (67.8 mg·L−1, 59 mg·L−1 and 49.7 mg·L−1 respectively). The environmental risk posed by the selected pharmaceuticals was assessed by calculating risk quotients (RQs) based on MEC (the highest exposure concentration of the compound in the medium)/PNEC (predicted no effect concentration) ratios. Results suggest that, at environmental concentrations, none of the compounds is harmful for the aquatic environment (low or no risk). Toxicity data obtained for mixtures were compared with predictions derived from three different models: Concentration Addition (CA), Independent Action (IA) and Combination Index (CI). The classical modeling approaches CA and IA failed to predict the observed mixture toxicity, thus indicating that single compound toxicity data are not sufficient to predict toxicity of drug mixtures on Tisbe species. However, the use of the CI seems to provide better predictions of pharmaceutical toxicity.
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•Nauplii (<24 h) of Tisbe battagliai were used to test acute toxicity of pharmaceuticals individually and in mixtures.•The ranking of acute toxicity to Tisbe battagliai of the four pharmaceutical compounds was DCF > IBU > CBZ > APAP.•The pharmaceuticals were classified as toxic (DCF) or harmful (APAP, CBZ and IBU) based on obtained LC50 values.•Risk quotients (RQ) indicate that, at environmental concentrations, there is no risk for Tisbe battagliai populations•CA and IA prediction models cannot well predict joint behavior of mixture of pharmaceuticals. However CI model provides better predictions
The knowledge about the effects of pharmaceuticals on aquatic organisms has been increasing in the last decade. However, due to the variety of compounds presents in the aquatic medium, exposure ...scenarios and exposed organisms, there are still many gaps in the knowledge on how mixtures of such bioactive compounds affect exposed non target organisms. The crayfish Procambarus clarkii was used to analyze the toxicity effects of mixtures of ciprofloxacin, flumequine and ibuprofen at low and high concentrations (10 and 100 μg/L) over 21 days of exposure and to assess the recovery capacity of the organism after a depuration phase following exposure during additional 7 days in clean water. The crayfish accumulated the three compounds throughout the entire exposure in the hepatopancreas. The exposure to the mixture altered the abundance of proteins associated with different cells functions such as biotransformation and detoxification processes (i.e. catalase and glutathione transferase), carbohydrate metabolism and immune responses. Additionally changes in expression of genes encoding antioxidant enzymes and in activity of the corresponding enzymes (i.e. superoxide dismutase, glutathione peroxidase and glutathione transferase) were reported. Alterations at different levels of biological organization did not run in parallel under all circumstances and can be related to changes in the redox status of the target tissue. No differences were observed between control and exposed organisms for most of selected endpoints after a week of depuration, indicating that exposure to the drug mixture did not produce permanent damage in the hepatopancreas of P. clarkii.
Antibiotics, frequently detected in aquatic ecosystems, can negatively impact the health of resident organisms. Although the study on the possible effects of antibiotics on these organisms has been ...increasing, there is still little information available on the molecular effects on exposed non-target organisms. In our study we used a label free proteomic approach and sea bream, Sparus aurata, to evaluate the effects of exposure to environmentally relevant concentrations of the antibiotic compounds ciprofloxacin (CIP), sulfadiazine (SULF) and trimethoprim (TRIM) produced at the protein level. Individuals of sea bream were exposed to single compounds at 5.2 ± 2.1 μg L−1 of CIP, 3.8 ± 2.7 μg L−1 of SULF and 25.7 ± 10.8 μg L−1 of TRIM for 21 days. After exposure, the number of differentially expressed proteins in the liver was 39, 73 and 4 for CIP, SULF and TRIM respectively. In the brain, there was no alteration of proteins after CIP and TRIM treatment, while 9 proteins were impacted after SULF treatment. The differentially expressed proteins were involved in cellular biological, metabolic, developmental, growth and biological regulatory processes. Overall, our study evidences the vulnerability of Sparus aurata, after exposure to environmentally relevant concentrations of the major antibiotics CIP, SULF and TRIM and that their chronic exposure could lead to a stress situation, altering the proteomic profile of key organs such as brain and liver.
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•Most molecular alterations occurred in the liver of sea bream.•CIP and SULF mainly affected carbohydrate metabolism and protein synthesis.•Protein expression was not significantly modulated after TRIM treatment.•SULF was the compound causing the most alterations in protein expression.
Ultraviolet filters (UV-filters) are compounds extensively used in personal care products. These compounds are produced at increasing rates and discharged into marine ecosystems in unknown quantities ...and with no regulation, making them emerging contaminants. Among those, the UV-filter 4-Methylbenzylidene camphor (4-MBC) is used in a variety of personal care products such as sunscreens, soaps, or lipsticks. This high consumption has resulted in its presence in various environmental matrices at in concentrations ranging from ng to μg L−1. Very little is known, however, about the possible adverse effects in exposed non-target organisms. Our study presents novel data on the bioconcentration, toxicokinetics, and molecular effects of 4-MBC in a marine bivalve species of commercial interest, Ruditapes philippinarum (Manila clam). Organisms were exposed at two different concentrations (1.34 and 10.79 μg L−1) of 4-MBC for 7 days, followed by a 3-day depuration period (clean sea waters). Bioconcentration factors (BCF) were 3562 and 2229 L kg−1 for the low and high exposure concentrations, respectively, making this pollutant bioaccumulative according to REACH criteria. Up to six 4-MBC biotransformation products (BTPs)were identified, 2 of them for the first time. Transcriptomic analysis revealed between 658 and 1310 differently expressed genes (DEGs) after 4-MBC exposure. Functional and enrichment analysis of the DEGs showed the activation of the detoxification pathway to metabolize and excrete the bioconcentrated 4-MBC, which also involved energy depletion and caused an impact on the metabolism of carbohydrates and lipids and in the oxidative phosphorylation pathways. Oxidative stress and immune response were also evidenced through the activation of cathepsins and the complement system. Such elucidation of the mode of action of a ubiquitous pollutant such as 4-MBC at the molecular level is valuable both from an environmental point of view and for the sustainable production of Manila clam, one of the most cultivated mollusk species worldwide.
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•4-MBC has a bioconcentration fold >2000 in Manila clam.•Integration of bioconcentration and toxicokinetics with transcriptomic data was applied.•Inflammatory-immune response evidenced by impacted cathepsin-encoding genes.•Detoxification of 4-MBC caused energy depletion and oxidative stress in Manila clam.
Large ships, particularly container ships, tankers, bulk carriers and cruise ships are significant individual contributors to air pollution. The European Environment Agency recognizes that air ...pollution in Europe is a local, regional and transborder problem caused by the emission of specific pollutants, which either directly or through chemical reactions lead to negative impacts, such as damage to human health and ecosystems. In the Marine Strategy Framework Directive 2008/56/EC of the European Parliament emissions from ships are mentioned explicitly in the list of pressures and impacts that should be reduced or minimized to maintain or obtain a good ecological status. While SOx and NOx contribute mainly to ocean and soil acidification and climate change, PM (particularly ultrafine particles in the range of nanoparticles) has the potential to act more directly on human and ecosystem health. Thus, in terms of risk assessment, one of the most dangerous atmospheric aerosols for environmental and human health is in the size range of nanoparticles. To our knowledge, no study has been carried out on the effects of the fraction that ends up in the water column and to which aquatic and sediment-dwelling organisms are exposed. Therefore, an integrated environmental risk assessment of the effects of emissions from oceangoing ships including the aquatic compartment is necessary. Research should focus on the quantitative and qualitative determination of pollutant emissions from ships and their distribution and fate. This will include the in situ measurement of emissions in ships in order to derive realistic emission factors, and the application of atmospheric and oceanographic transportation and chemistry models.
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•Large ships are significant contributors to air pollution.•Ultrafine particles have the potential to act on human and ecosystem health.•Ship emission inventories are subjected to high uncertainties.•Integrated risk assessment of the emissions from oceangoing ships should be carried out.
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