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required under the EU Marine Strategy Framework Directive to prevent any harm on marine ecosystems and their biota. Sampling to evaluate effects of marine litter, including microdebris, in top predators is difficult. Usually, microdebris is examined in the gastro-intestinal tract of stranded dead animals. However, the population of the endangered Mediterranean monk seal is too small for obtaining sufficient samples. The present study implemented for the first time a non-invasive technique for collecting monk seal samples to assess the microdebris ingestion in a systematic manner. A total of 12 samples of monk seal faeces were collected from marine caves in Zakynthos Island, Greece (INTERREG MED project “Plastic Busters MPAs”). A total of 166 particles were identified; 77% of the particles were smaller than 3 mm but a piece of net larger than 5 cm was also found. The majority of particles were filaments (84%), and only one sphere was found. Faeces contained on average 14 particles per sample, half of the average in previous studies using the whole digestive tract. The use of faeces represents an effective non-invasive tool to assess the incidence of microdebris and the trophic transfer of these pollutants.

Sea turtles are considered as bio-indicators for monitoring the efficiency of restoration measures to reduce marine litter impacts on health. However, the lack of extended and standardised empirical data has prevented the accurate analysis of the factors influencing litter ingestion and the relationships with individual health. Historic data collected from 1988 and standard data collected from 2016 were harmonised to enable such analyses on necropsied loggerhead turtles (Caretta caretta) in eight Mediterranean and North-East Atlantic countries. Litter was found in 69.24 % of the 1121 individuals, mostly single-use and fishing-related plastics. Spatial location, sex and life history stage explained a minor part of litter ingestion. While no relationships with health could be detected, indicating that all individuals can be integrated as bio-indicators, the mechanistic models published in literature suggest that the high proportion of plastics in the digestive contents (38.77 % per individual) could have long-term repercussions on population dynamics. © 2022

The Eastern Mediterranean Sea (EMS) is an ultraoligotrophic semi-enclosed sea with low nutrient levels, low primary production rate, impoverished phytoplankton populations and low zooplankton standing stocks. The Cretan Passage in the western Levantine Sea is one of the least explored areas of the EMS. We measured the mesozooplankton biomass, abundance and taxon composition as well as biochemical parameters like Electron Τransport System (ETS) activity and Aminoacyl-tRNA Synthetases (AARS) activities in late spring of 2016 (28 March–17 April 2016). The sampling area is influenced by the presence of the permanent Rhodes Cyclonic Gyre east of Crete and a series of smaller permanent or recurrent gyres south of Crete which influence the pathway of the Atlantic Water from the Ionian Sea towards the Levantine Basin at surface and subsurface layers. Mesozooplankton abundance showed slightly increasing values from western to eastern stations of the Cretan Passage and although subsurface maxima were recorded in the 50–100 m layer, the abundance at all stations generally decreased with depth. The community was dominated by copepods at all stations. Juveniles of Clausocalanus followed by juveniles of Oithona and the Oncaea media group (Oncaea waldemari, Oncaea scottodicarloi, Oncaea media) were the most abundant taxa in the upper layer, whereas Mormonilla minor, Haloptilus longicornis and Subeucalanus monachus were mainly found below 100 m. ETS, AARS and biomass for carbon measurements were conducted in the same area for the 0–100 m and 0–500 m layers. ETS was highest at LV13 with 10.56 ± 1.33 μl O2 h−1 mg protein−1 in 0–500 m and carbon biomass was highest at LV18 with 4.99 mg m−3 in 0–100 m. Lowest values of 4.52 ± 1.46 μl O2 h−1 mg protein−1 at LV18 in 0–100 m were detected for ETS and 0.91 mg m−3 at LV10 in 0–500 m for carbon biomass. Specific AARS activities showed no significant differences among sites. On the basis of ETS and AARS, we calculated that the zooplankton carbon requirements follow the west to east trend of zooplankton biomass, however, the zooplankton is not well fed and the organisms are living under oligotrophic stress.

The presence of marine litter is a complex, yet persistent, threat to the health and biodiversity of the marine environment, and plastic is the most abundant, and ubiquitous type of marine litter. To monitor the level of plastic waste in an area, and the prospect of it entering the food chain, bioindicator species are used extensively throughout Northern European Seas, however due to their distribution ranges many are not applicable to the Mediterranean Sea. Guidance published for the Marine Strategy Framework Directive suggests that the contents of fish stomachs may be analyzed to determine trends of marine plastic ingestion. In order to equate transnational trends in marine plastic ingestion, the use of standardized fish species that widely occur throughout the basin is favoured, however for the Mediterranean Sea, specific species are not listed. Here we propose a methodology to assess how effective Mediterranean fish species, that are known to have ingested marine plastic, are as bioindicators. A new Bioindicator Index (BI) was established by incorporating several parameters considered important for bioindicators. These parameters included species distribution throughout the Mediterranean basin, several life history traits, the commercial value of each species, and the occurrence of marine litter in their gut contents. By collecting existing data for Mediterranean fish, ranked scores were assigned to each trait and an average value (BI value) was calculated for each species. Based on their habitat preferences, Engraulis encrasicolus (pelagic), Boops boops (benthopelagic), three species of Myctophidae (Hygophum benoiti, Myctophum punctatum and Electrona risso) (mesopelagic), Mullus barbatus barbartus (demersal) and Chelidonichthys lucerna (benthic), were identified as currently, the most suitable fish for monitoring the ingestion of marine plastics throughout the Mediterranean basin. The use of standardized indicator species will ensure coherence in the reporting of marine litter ingestion trends throughout the Mediterranean Sea.

Microplastics (MPs) are a contaminant of growing concern due to their widespread distribution and interactions with marine species, such as filter feeders. To investigate the MPs accumulation by wild and cultured mussels, a Dynamic Energy Budget (DEB) model was developed and validated with the available field data of Mytilus edulis (wild) from the North Sea and Mytilus galloprovincialis (cultured) from the North Ionian Sea. Towards a generic DEB model, the site-specific model parameter, half saturation coefficient (Xk) was applied as a power function of food density for the cultured mussel, while for the wild it was calibrated to a constant value. The DEB-accumulation model simulated the uptake and excretion rate of MPs, taking into account environmental characteristics (temperature and chlorophyll-a). An accumulation of MPs equal to 0.64 particles individual−1 (fresh tissue mass 1.9 g) and 0.91 particles individual−1 (fresh tissue mass 3.4 g) was found for the wild and cultured mussel respectively, in agreement with the field data. The inverse experiments investigating the depuration time of the wild and cultured mussel in a clean from MPs environment showed a 90 % removal of MPs load after 3 and 14 days, respectively. Furthermore, sensitivity tests on model parameters and forcing functions highlighted that besides MPs concentration, the accumulation is highly depended on temperature and chlorophyll-a of the surrounding environment. For this reason, an empirical equation was found relating directly the concentration of MPs in seawater, with MPs accumulation in mussel’s soft tissue, temperature and chlorophyll-a.

This study assesses microplastic ingestion in Boops boops at different geographical areas in the Mediterranean Sea. A total of 884 fish were caught at 20 coastal sites in Spain, France, Italy and Greece and analyzed using a common methodological protocol. Microplastics were found in 46.8% of the sampled fish, with an average number of items per individual of 1.17 ± 0.07. Filaments were the predominant shape type, while polyethylene and polypropylene were indicated by FTIR as the most common polymer types of ingested microplastics. The frequency of occurrence, as well as the abundance and proportion of types (size, shape, color and polymer) of ingested microplastics, varied among geographical areas. The spatial heterogeneity of the abundance of ingested microplastics was mainly related to the degree of coastal anthropogenic pressure at the sampling sites. Our findings further support the suitability of B. boops as bioindicator of microplastic pollution in the Mediterranean Sea.

The following protocol is intended to respond to the requirements set by the European Union’s Marine Strategy Framework Directives (MSFD) for the D10C3 Criteria reported in the Commission Decision (EU), related to the amount of litter ingested by marine animals. Standardized methodologies for extracting litter items ingested from dead sea turtles along with guidelines on data analysis are provided. The protocol starts with the collection of dead sea turtles and classification of samples according to the decomposition status. Turtle necropsy must be performed in authorized centers and the protocol described here explains the best procedure for gastrointestinal (GI) tract isolation. The three parts of the GI (esophagus, stomach, intestine) should be separated, opened lengthways and contents filtered using a 1 mm mesh sieve. The article describes the classification and quantification of ingested litter, classifying GI contents into seven different categories of marine litter and two categories of natural remains. The quantity of ingested litter should be reported as total dry mass (weight in grams, with two decimal places) and abundance (number of items). The protocol proposes two possible scenarios to achieve the Good Environmental Status (GES). First: “There should be less than X% of sea turtles having Y g or more plastic in the GI in samples of 50-100 dead turtles from each sub-region”, where Y is the average weight of plastic ingested and X% is the percentage of sea turtles with more weight (in grams) of plastic than Y. The second one, which considers the food remain versus plastic as a proxy of individual health, is: “There should be less than X% of sea turtles having more weight of plastic (in grams) than food remains in the GI in samples of 50-100 dead turtles from each sub-region”. © 2019 Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.

The environmental effects of the copper aquaculture nets were estimated by a multidisciplinary approach. A copper alloy net (brass net) cage and a conventional net cage with Cu2O antifouling paint were established at a fish farm in Bathy Limani Bay, Paleros, Aetolia Acarnania region, Greece, for a 1-year period. Copper concentrations in abiotic (water, sediment, particulate matter) and biotic (organisms) samples were measured. The biological effects of copper on organisms (mussels, farmed fish) were further assessed with a battery of biochemical biomarkers (acetylcholinesterase, catalase, glutathione-S-transferase, metallothioneins). During the first 6 months of cage installations, the copper alloy (brass) net seemed to release more Cu to the surrounding environment and affect the biota more compared to the conventional net, although non-significant correlations were found between biotic and abiotic variables. However, at the end of the experimental period, equilibrium was reached in the copper alloy (brass) net cage with similar values of dissolved Cu and Cu in organisms as the conventional net cage. An exception to these results was the sampling of August 2013, where an increased effect on water chemistry and mussels in the conventional cage was observed, which may be attributed to the use of antifoulants on the examined conventional net or on the surrounding conventional nets. No accumulation of Cu in sediment was detected. No consistent temporal or spatial trend was observed for biochemical biomarkers. In addition, for both experimental cages and in all sampling periods, dissolved Cu never reached the maximum permissible limits in the Greek legislation, and Cu concentrations in the farmed fish and mussels do not exceed established limits for edible parts set by Food and Agriculture Organization of the United Nations (FAO). Statement of relevance In this manuscript, a set of methodological tools and indicators were studied in order to assess the potential impacts of the use of copper alloy aquaculture nets on farmed fish and, eventually, the marine environment of the Mediterranean Sea. The authors believe that the aim of this work is within the journal's scope and the presented results, analysis and interpretation will be of interest to scientists involved in various aspects of aquaculture (management, environmental pollution, ecotoxicology, food safety etc.).

Plastic litter pollution is increasing in the seas and oceans worldwide, raising concern on the potential effects of plasticizer additives on marine fauna. In this study, muscle samples of 30 bogues (Boops boops; Linneaus, 1758) from the North Western Mediterranean Sea were analysed to assess the concentrations of 19 organophosphate flame retardant (OPFR) compounds and to inspect any relationship with microplastic ingestion and relative levels of anthropization. Out of the 19 OPFRs analysed, 6 compounds were detected, being tri-n-butyl phosphate (TNBP), 2-ethylhexyldiphenyl phosphate (EHDPP) and triphenylphosphine oxide (TPPO) the most abundant. As expected, OPFR concentrations were higher in samples collected off the most anthropized area of the city of Barcelona than in those from the Cap de Creus Marine Protected Area, while no significant correlation was detected between OPFR concentrations and microplastic ingestion. The results of this manuscript provide a first evidence of OPFR presence in the muscle of the bogue and identify the coastal area off Barcelona as a possible concentration area for contaminants, further supporting the use of the bogue as an indicator species of plastic pollution in the Mediterranean Sea., This study was supported by the Spanish Ministry of Science and Innovation (Project CEX2018-000794-S) and by the Generalitat de Catalunya (Consolidated Research Group Water and Soil Quality Unit 2017 SGR 1404). The authors are grateful to the Confradia de pescadors de Barcelona and the Confradia de pescadors de Roses, which provided the bogues samples. OGG’s work was funded through an FPU Ph.D. scholarship granted by the Spanish Government. Constructive feedback from two anonymous reviewers contributed to improve the manuscript.

Plastic debris has become a major threat to the marine environment and wildlife. Sea turtles are particularly vulnerable, and are known to ingest plastic debris globally; however, information from Greek waters is still absent. In this study, 36 stranded dead loggerhead turtles (Caretta caretta) were collected from the Greek coastline area, and their gastrointestinal content was analysed for ingested plastic debris. Twenty-six individuals (72%) were found to have ingested plastic, with an average of 7.94 ± 3.85 (SE) plastic items per turtle. In total, 286 plastic items were counted and categorised by size, shape, colour, and polymer type. Fourier Transform Infrared Spectrometry revealed that polypropylene and polyethylene were the dominant polymer plastic types found. Results indicated a variation in plastic ingestion amongst life stages of the loggerhead specimens. This study provides evidence of plastic ingestion by loggerhead turtles in Greek waters.

Olive oil production generates large volumes of wastewaters mostly in peri-Mediterranean countries with adverse impacts on the biota of the receiving aquatic systems. Few studies have however documented its toxicity on aquatic species, with an almost total lack of relative studies on fish. We assessed the acute and sub-chronic OMW toxicity, as well as the acute and sub-chronic behavioural, morphological and biochemical effects of OMW exposure on the mosquitofish Gambusia holbrooki. LC50 values of the acute bioassays ranged from 7.31% (24 h) to 6.38% (96 h). Behavioural symptoms of toxicity included hypoactivity and a shift away from the water surface, coupled with a range of morphological alterations, such as skin damage, excessive mucus secretion, hemorrhages, fin rot and exophhalmia, with indications also of gill swelling and anemia. Biochemical assays showed that OMW toxicity resulted in induction of catalase (CAT) and inhibition of acetylcholinesterase (AChE) activities. The implications of our results at the level of environmental policy for the sustainable management of the olive mill industry, i.e. the effective restriction of untreated OMW disposal of in adjacent waterways, as well as the implementation of new technologies that reduce their impact (detoxification and/or revalorization of its residues) are discussed.

Microplastic ingestion by marine organisms presents an emerging threat to marine ecosystems; microplastics in different marine species are currently reported worldwide. This study aims to assess microplastic ingestion in four, highly commercial, marine species from Greek waters in the Northern Ionian Sea (Mediterranean Sea). Microplastics were found in mussels (Mytilus galloprovincialis) and all three fish species (Sardina pilchardus, Pagellus erythrinus, Mullus barbatus) examined. The frequency of occurrence of ingested microplastics was 46.25% in mussels, while among fish species, S. pilchardus showed the highest frequency of microplastic ingestion (47.2%). Microplastic abundance ranged from 1.7–2 items/individual in mussels and from 1.5–1.9 items/individual in fish. The majority of ingested microplastics were fragments, while their color and size varied. Fourier Transform Infrared Spectroscopy (FT-IR) indicated polyethylene as the most common polymer type in mussels and fish. Results can be used to set baseline levels for the assessment of microplastic pollution in the Ionian Sea.

Pollution effects were assessed by means of biochemical biomarkers (catalase, glutathione S-transferase and acetylcholinesterase activities, and metallothioneins content) in five species at selected coastal sites across the Eastern Mediterranean and the Black Sea. The mussel Mytilus galloprovincialis, a well-established sentinel species, was investigated in the Adriatic Sea, Aegean Sea, and Black Sea. The mussel Brachidontes pharaonis and the striped red mullet Mullus surmuletus were used in the Levantine Sea where M. galloprovincialis is not present. The white seabream Diplodus sargus sargus and the gastropod Rapana venosa were additionally sampled in the Adriatic and the Black Sea, respectively. Mussels showed catalase, glutathione S-transferase, and acetylcholinesterase responses to pollution in most geographical areas while the response of metallothioneins was restricted to a few sites. R. venosa showed marked responses of catalase and metallothioneins whereas both fish species did not generally exhibit variations in biomarker values among sites. The approach based on the reference deviation concept using the "Integrated Biological Responses version 2" index was useful for the interpretation of overall biomarker responses.

The Mediterranean Sea has been described as one of the most affected areas by marine litter in the world. Although effects on organisms from marine plastic litter ingestion have been investigated in several oceanic areas, there is still a lack of information from the Mediterranean Sea. The main objectives of this paper are to review current knowledge on the impact of marine litter on Mediterranean biodiversity, to define selection criteria for choosing marine organisms suitable for use as bioindicator species, and to propose a methodological approach to assessing the harm related to marine litter ingestion in several Mediterranean habitats and sub-regions. A new integrated monitoring tool that would provide the information necessary to design and implement future mitigation actions in the Mediterranean basin is proposed. According to bibliographic research and statistical analysis on current knowledge of marine litter ingestion, the area of the Mediterranean most studied, in terms of number of species and papers in the Mediterranean Sea is the western sub-area as well as demersal (32.9%) and pelagic (27.7%) amongst habitats. Applying ecological and biological criteria to the most threatened species obtained by statistical analysis, bioindicator species for different habitats and monitoring scale were selected. A threefold approach, simultaneously measuring the presence and effects of plastic, can provide the actual harm and sub-lethal effects to organisms caused by marine litter ingestion. The research revealed gaps in knowledge, and this paper suggests measures to close the gap. This and the selection of appropriate bioindicator species would represent a step forward for marine litter risk assessment, and the implementation of future actions and mitigation measures for specific Mediterranean areas, habitats and species affected by marine litter ingestion Versión del editor 3,746

Microplastics have become a more and more dominant threat to marine ecosystems. The ubiquity of microplastics is one of the major problems; from the sea surface and water column to the beach and seabed sediment or even ingested by marine organisms, small plastic particles have been found. This study is focused on monitoring and assessment of microplastic pollution (plastic particles

We assessed amounts, composition and net accumulation rates every ~ 15 days of beach macro litter (≥ 2.5 cm) on 4 Mediterranean beaches, on Corfu island, N. Ionian Sea, taking into account natural and anthropogenic drivers. Average net accumulation rate on all beaches was found 142 ± 115 N/100 m/15 d. By applying a Generalized Linear Model (GzLM) it was shown that sea transport is the dominant pathway affecting the amount and variability in beach litter loadings. Principal Component Analysis (PCA) on compositional data and indicator items discerned two more pathways of beach litter, i.e. in situ litter from beach goers and wind and/or runoff transport of litter from land. By comparing the PCA results to those from a simple item to source attribution, it is shown that regardless their source litter items arrive at beaches from various pathways. Our data provide baseline knowledge for designing monitoring strategies and for setting management targets.

Microplastic pollution is a growing cause of concern for the marine environment, particularly in the Mediterranean Sea, which is considered to be one of the most polluted seas worldwide. In this study, the gastrointestinal tracts of 102 bogues (Boops boops), sampled from three areas off the Catalan coast (Spain) subject to different degrees of industrialization, were analysed to assess microplastic ingestion and thus estimate local levels of microplastic pollution. Microplastics were detected in 46% of samples analysed. As expected, the abundance and frequency of occurrence of ingested microplastics were higher off the most anthropized area of Barcelona. The majority of ingested microplastics were blue fragments ranging 0.1–0.5 mm, and the most common polymer type was polypropylene. The results of this study indicate the area off Barcelona as a possible area of concentration for microplastics, further supporting the use of B. boops as a bioindicator to assess microplastic pollution.

Research studies on the effects of microlitter on marine biota have become more and more frequent the last few years. However, there is strong evidence that scientific results based on microlitter analyses can be biased by contamination from air transported fibres. This study demonstrates a low cost and easy to apply methodology to minimize the background contamination and thus to increase results validity. The contamination during the gastrointestinal content analysis of 400 fishes was tested for several sample processing steps of high risk airborne contamination (e.g. dissection, stereomicroscopic analysis, and chemical digestion treatment for microlitter extraction). It was demonstrated that, using our methodology based on hermetic enclosure devices, isolating the working areas during the various processing steps, airborne contamination reduced by 95.3%. The simplicity and low cost of this methodology provide the benefit that it could be applied not only to laboratory but also to field or on board work.

This study aimed to assess oxidative stress and genotoxicity biomarkers in Prussian carp Carassius gibelio laboratory-exposed to water from polluted Ukrainian rivers in order to evaluate their usefulness as endpoints in a short-term bioassay for toxicity testing of freshwaters. The micronucleus (MN) test and the frequency of cells with double nuclei (DN) in erythrocytes and gill cells were used as indicators of chromosome aberrations and abnormalities in cell divisions, respectively. Cellular antioxidant defenses i.e. antioxidant enzyme activities (catalase, Se-dependent glutathione peroxidase, total glutathione peroxidase and glutathione-S-transferase) and oxidative damage, i.e. lipid peroxidation (measured as thiobarbituric acid reactive substances) in the fish liver were used as biomarkers of oxidative stress. Exposure to the polluted river water samples for 96 h resulted in significantly increased MN and DN frequencies, limited increases in antioxidant enzyme activities and no changes in lipid peroxidation. Results suggest that MN and DN frequencies in C gibelio are useful endpoints in a short-term bioassay for genotoxicity testing of environmental water samples in contrast to the oxidative stress biomarkers applied that showed low potential for assessing sublethal effects after a 96 h exposure.

Adult gilthead sea bream Sparus aurata and sea bass Dicentrarchus labrax were exposed for 24 and 48 h, respectively, to two concentrations of ammonia each (mean values of 3·34 and 13·10 mg l(-1) TA-N in S. aurata; 2·99 and 11·90 mg l(-1) TA-N in D. labrax). Light microscopy and computerized morphometry were used to evaluate ammonia-induced alterations in skin structure during exposure and following recovery in normal water. In S. aurata, ammonia exposure induced a concentration-dependent increase in the number (hyperplasia) of neutral mucous cells (mc), with peak values at 24 h recovery after exposure. An increase in the dispersion of melanosomes in skin melanocytes was also observed in the dermis and occasionally in the epidermis of S. aurata, with peak values at 24 h of ammonia exposure. Exposure of D. labrax to ammonia had, likewise, concentration-dependent effects on mucous secretion. Of the two types of mc in this species, there was an increase in the number of the neutral mc and a reduction in the much more numerous acid mc, with peak values at 24 and 48 h, respectively, of ammonia exposure. The more intense mucous secretion in D. labrax compared to S. aurata could be related to the lower tolerance to ammonia in D. labrax, as reported elsewhere. Finally, the increase in melanosome dispersion was less evident in D. labrax, due to highly variable control values. These morphological alterations to the skin could be useful indicators of non-specific stress in cultured fishes.

This study aimed to assess oxidative stress and genotoxicity biomarkers in grey mullet Mugil cephalus from a site receiving discharges from industrial and harbor activities and a reference site in Saronikos Gulf, Greece. Cellular antioxidant defenses, i.e., antioxidant enzyme activities (catalase, selenium [Se]-dependent glutathione peroxidase, glutathione-S-transferase) and oxidative damage, i.e., lipid peroxidation (measured as thiobarbituric acid reactive substances) were used as biomarkers of oxidative stress. The micronucleus test was used for evaluation of chromosome damage, and the frequency of cells with double nuclei was used as an indication of damage to cell divisions. Antioxidant enzyme activities were lower in fish from the polluted site compared with fish from the reference site, suggesting deficiency of the antioxidant system to compensate for oxidative stress. This is further supported by the higher lipid peroxidation levels in fish from the polluted site, which represent oxidative damage. Micronuclei and double-nuclei frequencies were higher in fish from the polluted site compared with fish from the reference site, indicating genotoxic effects. Correlations between biomarkers suggest that observed effects were due to contaminants exhibiting oxidative stress potential that can also induce genotoxicity. Results suggest that the selected biomarkers in M. cephalus are useful for the assessment of pollution impacts in coastal environments influenced by multiple pollution sources.

A suite of biomarkers was measured in caged mussels at areas impacted by different anthropogenic activities along the Greek coastline to assess biological effects of environmental pollution. Mussels were caged at coastal sites in the vicinity of major cities, in areas influenced by major industries, agricultural practices and in islands away from known sources of pollution. Biomarkers indicative of neurotoxicity (acetylcholinesterase, AchE), oxidative stress (catalase, CAT), phase 11 biotransformation of xenobiotics (glutathione S-transferase, GST), metal exposure (metallothioneins, MTs) and protein synthesis (RNA:DNA ratio) were measured to assess effects of various types of pollutants. AchE activity proved to be the most responsive biomarker with decreased values at sites influenced by agricultural, urban and industrial activities. Decreased CAT and GST activities and increased MTs levels were recorded at a number of anthropogenic-impacted sites. RNA:DNA ratio showed a biphasic response as both high and low values were found at impacted sites. Principal component analysis clearly distinguished sites receiving pollution inputs from non-polluted sites. The combination of the selected biomarkers used in caged mussels resulted useful in the assessment of the effects of environmental pollution.

A combination of bioenergetics and biochemical biomarkers in mussels was applied to assess possible pollution impacts in a protected semi-enclosed estuary (Amvrakikos Gulf, NW Greece) that receives pesticide discharges through riverine transport. Scope for growth, a physiological condition index representing the energy budget of the organism, was applied to detect general stress effects on the health status of mussels. The low energy budgets of mussels revealed stress conditions and provided early warning signals of possible consequences at higher levels of biological organization. Biochemical markers of exposure confirmed a risk of pesticide contamination. Decreased acetylcholinesterase activities indicated exposure to organophosphate and carbamate pesticides. Responses of the antioxidant enzyme glutathione peroxidase suggested the presence of contaminants capable of reactive oxygen species production that could be related to organochlorine pesticide contamination in the area. On the other hand, metallothionein levels implied low metal contamination.

This study examined the effects of nickel and chromium exposure on two biochemical biomarkers, glutathione peroxidase (GPX) and acetylcholinesterase (ACHE), in parallel with an indicator of overall health, Scope for Growth (SFG), in the mussel Mytilus galloprovincialis to evaluate their ecological relevance and to assess their potential use as biomarkers of Ni and Cr pollution. SFG integrates major physiological responses into an index that represents the energy status of the animals and is considered an ecologically relevant biomarker. Following exposure to a range of Ni and Cr concentrations effects of Ni and Cr were evident on GPX and SFG but not on ACHE. Accumulation in the mussels’ tissues increased according to exposure concentrations. Adverse effects on the energy status and induction of GPX were found at the higher exposure concentrations while possible hormetic effects on the energy status and moderate GPX suppression were associated with lower exposure concentrations. Nevertheless GPX was negati...

A set of biomarkers was used to assess the impact of heavy metal pollution by a ferro-nickel smelting plant in Larymna bay (North Evoikos Gulf, Greece). These included a biomarker reflecting health status of an organism (scope for growth, SFG), a cellular biomarker of heavy metal exposure (composition of metal-containing granules), and two biochemical biomarkers reflecting oxidative stress (glutathione peroxidase, GPX) and neurotoxicity (acetylcholinesterase, AChE) measured in mussels (Mytilus galloprovincialis) both native and transplanted for 1 and 6 months at the coastal area of Larymna. All biomarkers in mussels at Larymna revealed differences from mussels at a reference site, signaling effects of the increased heavy metal levels on the biota. While effects on SFG and GPX in Larymna mussels were obvious on short-term exposure and persistent during chronic exposure, only chronic exposure induced a possibly cumulative effect on AChE. To validate the causal relationship between heavy metal exposure and effects observed in Larymna, SFG, GPX, and ACHE were examined in mussels exposed to a mixture of heavy metals (Ni, Cr, and Fe) under controlled laboratory conditions. The laboratory experiment verified the causal relationship between SFG and GPX responses and heavy metals but this was not demonstrated for AChE. Results from field-collected and laboratory-exposed mussels indicated a potential of GPX as predictive biomarker of population-level effects of heavy metal exposure.

Crise, A. ... et. al.-- 12 pages, 1 figure, 5 tables, PERSEUS project aims to identify the most relevant pressures exerted on the ecosystems of the Southern European Seas (SES), highlighting knowledge and data gaps that endanger the achievement of SES Good Environmental Status (GES) as mandated by the Marine Strategy Framework Directive (MSFD). A complementary approach has been adopted, by a meta-analysis of existing literature on pressure/impact/knowledge gaps summarized in tables related to the MSFD descriptors, discriminating open waters from coastal areas. A comparative assessment of the Initial Assessments (IAs) for five SES countries has been also independently performed. The comparison between meta-analysis results and IAs shows similarities for coastal areas only. Major knowledge gaps have been detected for the biodiversity, marine food web, marine litter and underwater noise descriptors. The meta-analysis also allowed the identification of additional research themes targeting research topics that are requested to the achievement of GES. © 2015 The Authors, This work has been partially funded by the EC FP7 PERSEUS Project (Grant. Agr. 287600)

Dredged sediments derived by the low course and estuary of the metropolitan river of Athens (Kifissos River) were dumped every day for 21 months to an open-sea site in the Saronikos Gulf. The spoil-ground and surrounding area was monitored prior, during and post to dumping for 24 months, over 6-month intervals. Dumping significantly changed the granulometry of the pre-existing superficial sediments to finer-grained only in the spoil ground and increased the sediment contamination load (aliphatic, polycyclic aromatic hydrocarbons and heavy metals) throughout the study area. Microtox® SPT showed that sediment toxicity levels were high at almost all sampling stations. During dumping, burial of natural soft-bottom habitats degraded severely the communities of the spoil-ground resulting in an almost azoic state, as well as significantly declined the species number and abundance of benthic communities in locations up to 3.2 km away from the spoil-ground, due to dispersion of the spoil and smothering. Benthic indices on the surrounding sites were significantly correlated with hydrocarbon concentrations and sediment toxicity levels. Post to dumping, the macrofauna communities of the spoil-ground were still significantly degraded, but the surrounding areas showed patterns of recovery. However, the high concentrations of aliphatic, polycyclic aromatic hydrocarbons and levels of toxicity persisted in the sediments after the ceasing of dumping operations in the study area, implying the ecological hazard imposed on the area.

Progress report (T30, June 2014), Perseus Project. FP7 Grant N° 287600, Work package 2 - Pressures and impacts at coastal level, Sub-task 2.3.4. Pollution by persistent organic pollutants and toxic metals

Progress report (T30, June 2014), Perseus Project. FP7 Grant N° 287600, Work package 2 - Pressures and impacts at coastal level, Sub-task 2.3.4. Pollution by persistent organic pollutants and toxic metals

Impacts of chemical contaminants associated with dumping of dredged urban river sediments at a coastal disposal area in Saronikos Gulf (Eastern Mediterranean) were investigated through a combined approach of sediment toxicity testing and active biomonitoring with caged mussels. Chemical analyses of aliphatic hydrocarbons (AHs), polycyclic aromatic hydrocarbons (PAHs), Cu, and Zn in combination with the solid phase Microtox® test were performed on sediments. Concentrations of PAHs, AHs, Cu, and Zn as well as multiple biomarkers of contaminant exposure and/or effects were measured in caged mussels. Sediments in the disposal and neighboring area showed elevated PAHs and AHs concentrations and were characterized as toxic by the solid-phase Microtox® test during and after dumping operations. Biomarker results in the caged mussels indicated sublethal effects mainly during dumping operations, concomitantly with high concentrations of PAHs and AHs in the caged mussel tissues. Cu and Zn concentrations in sediments and caged mussels were generally not elevated except for sediments at the site in the disposal area that received the major amount of dredges. High PAHs and AHs levels as well as sublethal effects in the caged mussels were not persistent after termination of operations. The combined bioassay–biomarker approach proved useful for detecting toxicological impacts of dredged river sediment disposal in sediments and the water column. Nevertheless, further research is needed to evaluate whether sediment toxicity will have long-term effects on benthic communities of the disposal area.

The selection of the best site for the placement of dredged sedimentary material (∼7,000 m(3)) from the Aliveri coastal area in the adjacent South Euboean Gulf (Greece) was accomplished through a screening procedure. The initial stage comprised the determination of physical, chemical, and biological characteristics of the dredged sediment before the commencement of any dredging operation. Grain size measurements, geochemical analyses together with the use of pollution/toxicity indices and empirical sediment quality guidelines, and the conduct of an acute toxicity test showed that the dredged material consisted of "unpolluted to slightly polluted" silty sands and sandy silts. However, the local authorities planned to place this sediment in the neighboring open sea area, i.e., the South Euboean Gulf, due to the absence of any beneficial use or alternative dumping option (i.e., dumping on public lands). Therefore, the next stage of the screening procedure, based on criteria such as the national legislation, seabed and seawater column characteristics, influence of the water mass circulation pattern on the post-placement migration of dredged sediment, impact on living resources and human activities (i.e., aquaculture and fishing), effect on significant marine sites (i.e., sites of scientific, ecological, and historical importance, navigation routes, military zones), and seafloor engineering uses, led to the evaluation of the suitability of the South Euboean Gulf as a potential dumping area. Then, the identification of the appropriate dredged material placement sites in the South Euboean Gulf was based on a cluster analysis, which tested the physicochemical resemblance of the dredged material and the surface sediments of 19 potential placement locations in the gulf. After the statistical process, only four sites situated near the north shoreline of the South Euboean Gulf were qualified as the best dredged material placement locations.

Progress report (June 2013), Perseus Project, FP7 Grant N° 287600, Work package 2 - Pressures and impacts at coastal level, Task 2.3. Filling the knowledge gaps: dedicated field studies of the effects of pressures exerted on coastal ecosystems, Sub-task 2.3.4. Pollution by persistent organic pollutants and toxic metals.

REPORTING DECEMBER 2013, 1/ACTIVITY DETAILS, PERSEUS T2.3, Subtask 2.3.4 Pollution (persistent organic pollutants - POPs and toxic metals), Action 2: Pollutants impact on benthic communities related to sediment contamination

Metal concentrations were determined in surface sediments and transplanted mussels in a shallow semi-enclosed bay, Pagassitikos Gulf, Aegean Sea. Cu, Zn, Pb, Hg, and As levels in sediments were enriched close to point sources of pollution, i.e., Volos Port and cement plant. Fe, Mn, Cr, and Ni spatial distributions in sediments were rather uniform with increasing concentrations towards deeper sites, suggesting natural enrichment, whereas levels found in the port, although elevated, were not the highest in the study area. Spatial variations in concentrations of most metals in mussels transplanted nearshore were generally in accordance to those in sediments, with the highest Fe, Mn, Cr, Ni, and Cu levels in the port and the highest Hg levels close to the cement plant. Concentrations of metals in sediments and transplanted mussels were comparable to those reported in Mediterranean areas. Results indicate that pollution point sources contribute to anthropogenic metal enrichment in the study area.

Methods for assessing the quality of marine coastal environments are numerous and rapidly evolving. The integration of physiological parameters termed as Scope for Growth (SfG) and the luminescent bioassay Microtox® were applied in several sites of Saronikos and Amvrakikos Gulfs (Greece) to assess the environmental quality and compare the results of the two methods. Each site in the two areas undergoes different types and levels of pollution. Both methods could identify a pollution gradient reflecting the quality of each site. A very good linear correlation was noticed between the two methods (r=0.98). The stress order, which was similar in both methods, exhibited the expected pollution gradient according to the pressure that each site undergoes. Results suggest that for screening and fast isolation of polluted sites before further analysis, the rapid and simple Microtox® assay could operate as an alternative to the more complex SfG method.

Alterations in a number of biochemical parameters in marine organisms represent specific markers of exposure to particular classes of contaminants. They are used as tools for the detection and monitoring of pollution. In this study, two biochemical markers of pollution, metallothionein (MT) content and acetylcholinesterase (AChE) activity were measured in indigenous and transplanted mussels Mytilus galloprovincialis from coastal areas within the Saronikos Gulf subject to high anthropogenic activities. Biannual measurements of the two biomarkers in indigenous mussel populations for two years revealed no significant differences among stations representing a pollution gradient. Limited differences in MT levels were only found between mussel populations transplanted at lesser and more impacted stations. Both biomarkers showed a variation with respect to the season of sampling, whilst during the second year of measurements a concomitant increase in metallothionein content with a decrease in acetylcholinesterase activity was noted. Our results indicate that the applied biochemical markers in indigenous mussel populations do not reflect the type of pollution in the Saronikos Gulf to a degree that can be used for pollution monitoring in the area.

Microplastics (MPs) are a contaminant of growing concern due to their widespread distribution and interactions with marine species, such as filter feeders. To investigate the MPs accumulation in wild and cultured mussels, a dynamic energy budget (DEB) model was developed and validated with the available field data of Mytilus edulis (M. edulis, wild) from the North Sea and Mytilus galloprovincialis (M. galloprovincialis, cultured) from the northern Ionian Sea. Towards a generic DEB model, the site-specific model parameter, half-saturation coefficient (Xk), was applied as a power function of food density for the cultured mussel, while for the wild mussel it was calibrated to a constant value. The DEB-accumulation model simulated the uptake and excretion rate of MPs, taking into account environmental characteristics (temperature and chlorophyll a). An accumulation of MPs equal to 0.53 particles per individual (fresh tissue mass 1.9 g) and 0.91 particles per individual (fresh tissue mass 3.3 g) was simulated for the wild and cultured mussel after 4 and 1 years respectively, in agreement with the field data. The inverse experiments investigating the depuration time of the wild and cultured mussel in a clean-from-MPs environment showed a 90 % removal of MPs load after 2.5 and 12 d respectively. Furthermore, sensitivity tests on model parameters and forcing functions highlighted that besides MPs concentration, the accumulation is highly dependent on temperature and chlorophyll a of the surrounding environment. For this reason, an empirical equation was found, directly relating the environmental concentration of MPs, with the seawater temperature, chlorophyll a, and the mussel's soft tissue MPs load.