Elutriate preparation affects embryo development test with Paracentrotus lividus: An in-depth study on the differences between two protocols and three different sediment/water mixing times.

Coastal areas are under continuous and increasing pressure from different human activities. A mixture of contaminants (e.g. hydrocarbons, pesticides, persistent organic pollutants (POPs), emerging contaminants, and others), originating mainly from populated, industrialised and agricultural areas, can reach the marine environment through different means such as wastewater discharge, soil runoffs, leaching from agriculture, and volatilisation/deposition. In this context, marine sediments have increasingly been considered repositories for a variety of pollutants that can accumulate and be stored for long periods, acting as a secondary source of contaminants during subsequent dredging operation or vessel manoeuvring. Chemical and ecotoxicological analyses of sediments are routinely conducted to evaluate the potential hazard/risk to the environment, either on bulk sediment or elutriate. In general, sediment elutriates are commonly prepared according to ASTM Guide even if alternative protocols are proposed by USACE for the various condition that they have to represent. The goal of the present study was to determine if the toxicological properties of ASTMprepared elutriates are comparable to those obtained from the USACE protocol. Sediment coming from 3 harbours (Olbia, Cagliari, and Toulon), as part of the "Se.D.Ri.Port" Interreg Project, were processed to obtain elutriates according to ASTM Guide and USACE Dredging Elutriate protocol and tested with the sea urchin Paracentrotus lividus embryo development test. Moreover, the significance of different stirring times of water/sediment mixture (1 h, 3 h, and 24 h) was tested with both the ASTM and USACE protocol. In addition to the biological analysis, for each sediment sample, heavy metals concentration, granulometry, and organic matter were determined. Even if for the ports of Toulon and Cagliari, the ASTM and USACE elutriates showed comparable results with P. lividus bioassay, for the port of Olbia the two protocols showed different criticalities. Preliminary results show that for the site Olbia elutriates prepared with the USACE protocol resulted in higher toxicity than elutriates obtained with ASTM (p < 0.001). In conclusion, differences in preparation protocols appear to be significant and can lead to different results in biological testing. To overcome this problem and to obtain more reliable evaluations of risk to the environment, standardisation and regulation must be the next goals in sediment management procedure.

Development and validation of new analytical methods using sea urchin embryo bioassay to evaluate dredged marine sediments.

Management of dredged materials disposal is regulated by several environmental normative requirements, and it is often supported by the integration of chemical data with ecotoxicological characterization. The reliability of a bioassay to assess the potential toxicity of dredged sediments requires the selection of quality criteria that should be based on simple analytical methods and easily understandable hazard for politicians and environmental managers. The sea urchin embryo-toxicity bioassay is considered an essential component for evaluating the quality of sediments in harbour areas but its use, when based exclusively on the observation of normal vs. abnormal embryos, may alter the interpretation of the results, overestimating the risk assessment. To improve the reliability of this assay in establishing a causative relationship between quality of sediments and sea urchin embryonic development, here we developed and validated three Integrative Toxicity Indexes (ITI 2.0, ITI 3.0, ITI 4.0), modifying the already-known ITI (here ITI 1.0). Based on this aim, we used Taranto harbour as a model pilot-study to compare results to those obtained from standard criteria. Among the tested indexes, the ITI 4.0, discriminating strictly developmental delay and morphological defects from fertilized egg to gastrula stage, resulted in the most promising.

Larval development ratio test with the calanoid copepod Acartia tonsa as a new bioassay to assess marine sediment quality.

The copepod Acartia tonsa was used as a model species to assess marine sediment quality. Acute and chronic bioassays, such as larval development ratio (LDR) and different end-points were evaluated. As a pelagic species, A. tonsa is mainly exposed to water-soluble toxicants and bioassays are commonly performed in seawater. However, an interaction among A. tonsa eggs and the first larval stages with marine sediments might occur in shallow water environments. Here we tested two different LDR protocols by incubating A. tonsa eggs in elutriates and sediments coming from two areas located in Tuscany Region (Central Italy): Livorno harbour and Viareggio coast. The end-points analyzed were larval mortality (LM) and development inhibition (DI) expressed as the percentage of copepods that completed the metamorphosis from nauplius to copepodite. Aims of this study were: i) to verify the suitability of A. tonsa copepod for the bioassay with sediment and ii) to compare the sensitivity of A. tonsa exposed to different matrices, such as water and sediment. A preliminary acute test was also performed. Acute tests showed the highest toxicity of Livorno's samples (two out of three) compared to Viareggio samples, for which no effect was observed. On the contrary, LDR tests with sediments and elutriates revealed some toxic effects also for Viareggio's samples. Results were discussed with regards to the chemical characterization of the samples. Our results indicated that different end-points were affected in A. tonsa, depending on the matrices to which the copepods were exposed and on the test used. Bioassays with elutriates and sediments are suggested and LDR test could help decision-makers to identify a more appropriate management of dredging materials.

The impact of benzodiazepine use in patients enrolled in opioid agonist therapy in Northern and rural Ontario.

BACKGROUND: Benzodiazepine use is common among patients in opioid agonist therapy; this puts patients at an increased risk of overdose and death. In this study, we examine the impact of baseline and ongoing benzodiazepine use, and whether patients are more likely to terminate treatment with increasing proportion of benzodiazepine positive urine samples. We also study whether benzodiazepine use differs by geographic location. METHODS: We conducted a retrospective cohort study using anonymized electronic medical records from 58 clinics offering opioid agonist therapy in Ontario. One-year treatment retention was the primary outcome of interest and was measured for patients who did and did not have a benzodiazepine positive urine sample in their first month of treatment, and as a function of the proportion of benzodiazepine-positive urine samples throughout treatment. Cox proportional hazard model was used to characterize one-year retention. RESULTS: Our cohort consisted of 3850 patients, with the average retention rate of 43.4%. Baseline benzodiazepine users had a retention rate of 39.9% and non-users had a retention rate of 44%. Patients who were benzodiazepine negative on admission benefited from an increased median days retained of 265 vs. 215 days. Patients with more than 75% of urines positive for benzodiazepines were 175% more likely to drop out of treatment than those patients with little or no benzodiazepine use. CONCLUSIONS: Baseline benzodiazepine use is predictive of decreased retention. Patients who have a higher proportion of benzodiazepine-positive urine samples are more likely to drop out of treatment compared to those who have little or no benzodiazepine detection in their urine.

Toxicological effects of CdSe/ZnS quantum dots on marine planktonic organisms.

Quantum dot nanoparticles (QDs) are proposed as novel materials for photovoltaic technologies, light emitting devices, and biomedical applications. In this study we investigated the effect of CdSe/ZnS QDs on the growth rate of four microalgae: the diatom Phaeodactylum tricornutum, the cryptophyte Rhinomonas reticulata, the prymnesiophyte Isochrysis galbana and the green alga Dunaliella tertiolecta. In addition we analyzed the effect of QDs on the copepod Acartia tonsa. A classical acute test (48-h) with embryos was carried out to evaluate naupliar survival. Moreover, a 4-day chronic test with adult copepods was conducted to evaluate their fecundity (embryos f(-1)day(-1)) and egg hatching success. QDs in the range from 1 to 4nM gradually inhibited the growth rate of P. tricornutum, I. galbana, R. reticulata and D. tertiolecta with an EC50 of 1.5, 2.4, 2.5 and 4.2nM, respectively. Acute tests with A. tonsa (QD concentration tested from 0.15 to 1.5nM) showed an increased naupliar mortality in response to QD treatment, exhibiting an EC50 of 0.7nM. Chronic test showed no negative effect on egg production, except on the last two days at the highest QD concentration (2.5nM). No significant reduction of the percentage of egg hatching success was recorded during the exposure. Toxicity assessment of QDs was also investigated at the molecular level, studying heat shock protein 70 gene expression (hsp 70). Our results indicate that hsp70 was upregulated in adults exposed 3 days to 0.5nM QDs. Overall, these results suggest that species unable to swim along the water column, like P. tricornutum and early hatched copepods, could be more exposed to toxic effects of QDs which tend to aggregate and settle in seawater.

Detection of malformations in sea urchin plutei exposed to mercuric chloride using different fluorescent techniques.

Embryos of Mediterranean sea urchin Paracentrotus lividus and subtropical Echinometra mathaei were exposed to 5,10, 15 and 20µgL(-1), and to 1, 2, 3 and 4µgL(-1) mercuric chloride (HgCl2), respectively. The effective concentration (EC50) inducing malformation in 50% of 4-arm pluteus stage (P4) was 16.14µgL(-1) for P. lividus and 2.41µgL(-1) for E. mathaei. Two-photon (TP), second (SHG) and third harmonic generation (THG) microscopy techniques, TUNEL staining, propidium iodide (PI) and Hoechst 33342 probes were used to detect light signals or to stain apoptotic and necrotic cells in fixed and alive plutei. Signals were detected differently in the two species: TP fluorescence, commonly associated with apoptotic cells, did not increase with increasing HgCl2 concentrations in P. lividus and in fact, the TUNEL did not reveal induction of apoptosis. PI fluorescence increased in P. lividus in a dose-dependent manner, suggesting a loss of cell permeability. In E. mathaei plutei TP fluorescence increased at increasing HgCl2 concentrations. THG microscopy revealed skeletal rods in both species. Different fluorescent techniques, used in this study, are proposed as early-warning systems to visualize malformations and physiological responses in sea urchin plutei.

Bioconcentration and arsenic speciation analysis in ragworm, Hediste diversicolor (Muller 1776).

This study focused on bioconcentrations of arsenic in Hediste diversicolor (Müller 1776) after exposure to three different molecule solutions: arsenate, dimethyl-arsinate and arsenobetaine. Speciation analysis was carried out after exposing the organisms to these solutions in order to investigate their arsenic biotransformation capacity. Arsenic reached to the maximum level in these tissues after 15 days' exposure to a solution of 100 µg L(-1) of arsenobetaine, although a significant increase was obtained in worms exposed to arsenate. Speciation analysis shows that trimethyl-arsine oxide is the slowest detoxification phase recorded in experiment.

Arsenic exposure affects embryo development of sea urchin, Paracentrotus lividus (Lamarck, 1816).

Toxicity tests were performed with embryos of Paracentrotus lividus to investigate the toxicological effect of two arsenic species: arsenate (As(V)), expected to be more toxic, and dimethyl-arsinate (DMA) expected to be less toxic. Exposures to toxicants were performed at different developmental stages in order to identify the most sensitive phase of embryological development. Statistical analysis revealed a high significance of each factor (Molecule, Concentration and Time of exposure) and their interaction for the dependent variable "Percentage of normal-shaped plutei". In particular, the 8 cell stage was the most sensitive to arsenic; at a concentration of 50 µg L(-1) DMA proved to be more toxic than As(V), resulting in nearly 50 % of normal-shaped plutei against the 74 % recorded for As(V). Starting the administration of arsenic at the morula stage, arsenate proved to be significantly more toxic when compared to DMA.

Echinoderm larvae as bioindicators for the assessment of marine pollution: Sea urchin and sea cucumber responsiveness and future perspectives.

Echinoderms play a crucial role in the functioning of marine ecosystems and due to their extensive distribution, rapid response, and the high sensitivity of their planktonic larvae to a large range of stressors, some species are widely used as biological indicators. In addition to sea urchins, sea cucumbers have recently been implemented in embryotoxicity bioassays showing high potential in ecotoxicological studies. However, the use of this species is still hindered by a lack of knowledge regarding their comparative responsiveness. The present study aimed to investigate the responsiveness of different echinoderm species to environmental pollution in order to develop their integration in batteries of ecotoxicological bioassays. To this end, the embryos of two sea urchins (Paracentrotus lividus and Arbacia lixula) and two sea cucumbers (Holothuria polii and Holothuria tubulosa) were incubated with inorganic and organic toxicants (cadmium, copper, mercury, lead, sodium dodecyl sulphate and 4-n-Nonhyphenol) and elutriates from contaminated marine sediments, chosen as a case study model. The results obtained, expressed through the percentage of abnormal embryos and Integrative Toxicity Indices (ITI), indicated species-specific sensitivities to pollutants, with comparable and correlated responsiveness between sea urchins and sea cucumbers. More specifically, sea cucumber larvae exposed to elutriates appear to be more sensitive than sea urchins, especially when incubated with samples containing trace metals, PCB and TBT. These results indicate that toxic responses in embryos exposed to environmental matrices are probably modulated by interactions between different variables, including additive, synergistic and antagonistic effects. These findings suggest that performing a larval test using different echinoderm classes can integrate the interactive effects of bioavailable fraction of contaminants on various levels, providing sensitive, representative and all year-round batteries of bioassays to apply in ecotoxicological studies.

Eco-Friendly Engineered Nanomaterials Coupled with Filtering Fine-Mesh Net as a Promising Tool to Remediate Contaminated Freshwater Sludges: An Ecotoxicity Investigation.

The use of eco-friendly engineered nanomaterials represents a recent solution for an effective and safe treatment of contaminated dredging sludge. In this study, an eco-designed engineered material based on cross-linked nanocellulose (CNS) was applied for the first time to decontaminate a real matrix from heavy metals (namely Zn, Ni, Cu, and Fe) and other undesired elements (mainly Ba and As) in a lab-scale study, with the aim to design a safe solution for the remediation of contaminated matrices. Contaminated freshwater sludge was treated with CNS coupled with a filtering fine-mesh net, and the obtained waters were tested for acute and sublethal toxicity. In order to check the safety of the proposed treatment system, toxicity tests were conducted by exposing the bacterium Aliivibrio fischeri and the crustacean Heterocypris incongruens, while subtoxicity biomarkers such as lysosomal membrane stability, genetic, and chromosomal damage assessment were performed on the freshwater bivalve Dreissena polymorpha. Dredging sludge was found to be genotoxic, and such genotoxicity was mitigated by the combined use of CNS and a filtering fine-mesh net. Chemical analyses confirmed the results by highlighting the abetment of target contaminants, indicating the present model as a promising tool in freshwater sludge nanoremediation.

Impact of sheep wool residues as soil amendments on olive beneficial symbionts and bacterial diversity.

In recent years the use of organic matter soil amendments, such as agricultural by-products, has been implemented with the aim of increasing soil fertility, while minimizing the environmental impact of agriculture. Sheep wool residues (SWR) have shown beneficial effects on plant nutrition and soil properties, while only few works assessed their impact on soil microbial communities. The main aim of this work was to investigate the possible valorization of two SWR types (scoured residues, white wool, WW, and carbonized scoured residues, black wool, BW) as organic soil amendments, in pot-grown olive trees, by evaluating their impact on soil bacterial communities and mycorrhizal symbionts. The two SWR types did not negatively impact on the diversity and composition of soil bacterial communities, as revealed by PCR-denaturating gradient gel electrophoresis (PCR-DGGE) of partial 16S rRNA gene, and on the activity of native arbuscular mycorrhizal fungi (AMF), while positively affecting plant growth. Only the highest doses of one SWR type (2% BW) caused a decrease in bacterial diversity and native AMF ability to colonize olive roots. DGGE bands sequencing allowed the identification of the major bacterial taxa. Sequences corresponding to Ohtaekwangia spp., Beta proteobacterium, Blastocatella sp., Ramlibacter monticola and Massilia frigida/rubra, Dongia sp. and Chloroflexi were mainly represented in SWR-amended soils, while those represented by Chryseolinea soli and Acidobacteria were abundant in control soil. Overall, this work showed that SWR may be valorized as organic soil amendments, as soil bacteria and AMF, representing key factors of biological soil fertility, were not negatively affected, while the activity of bacterial genera and species known for their ability to decompose complex compounds was boosted. Further studies will investigate the biodegradation efficiency of the diverse bacterial taxa developing in SWR-amended soils.

Primary Diffuse Large B-Cell Lymphoma of the Urinary Bladder: Update on a Rare Disease and Potential Diagnostic Pitfalls.

Diffuse large B-cell lymphoma (DLBCL) represents the most frequent type of non-Hodgkin lymphoma. Globally, DLBCL is an aggressive disease, requiring an accurate diagnosis and prompt treatment. The diagnosis is often made on biopsy samples of a nodal mass, however, approximately 40% of DLBCL cases arise at extranodal sites. The most common extranodal site is the gastrointestinal tract, however any extranodal area may be primarily involved. Primary urinary bladder lymphoma represents only 0.2% of extranodal non-Hodgkin lymphomas, whereas secondary involvement of the urinary bladder by a systemic lymphoma is a more common event. Despite being rare, DLBCL is considered to represent the predominant primary urinary bladder lymphoma. The majority of cases reported in the bladder belong to the DLBCL, NOS group, and there are only rare cases of EBV-positive DLBCL, NOS. In this review, we summarize the current knowledge on DLBCL primarily occurring in the urinary bladder, with the aim of increasing clinician and pathologist awareness on this aggressive lymphoma rarely arising in the urinary bladder. Additionally, we focus on those entities which should be taken into consideration in the differential diagnosis, highlighting potential diagnostic pitfalls.

Historical control data in ecotoxicology: Eight years of tests with the copepod Acartia tonsa.

The calanoid copepod Acartia tonsa is one of the most frequently used organisms in acute, short-term bioassays to assess effects induced by marine matrices or chemicals on different life stages. Physiological responses in such tests can be highly variable and historical control data (HCD), values recorded from previous studies performed under similar conditions, can be useful to recognise the average responses over time. Here, we analysed egg hatching success and larval (naupliar) immobilisation/mortality of A. tonsa Mediterranean strain, cultured in laboratory conditions since 2008 and used as model organisms in ecotoxicology tests. Our aims were to evaluate the physiological response and sensitivity of A. tonsa over eight years of bioassays, and to compare our HCD with reference values, in order to assess the suitability of such a long-term culture for ecotoxicology studies. Acartia tonsa eggs were exposed for 48 h to the reference toxicant nickel chloride (NiCl2) and the % of egg hatching success and naupliar viability were compared to controls. A total of 59 acute tests, displayed in Shewhart-like control charts, showed a high mean percentage of egg hatching success (85.60% ± 5.90 SD) recorded for the whole period, and a low mean percentage of naupliar immobilisation/mortality (6.73% ± 6.38 SD) in controls. Effective concentration (EC50) for NiCl2 registered a stable mean of 0.14 mg Ni/L (± 0.047 SD) over time. Overall, our long-term dataset confirms the suitability of this copepod species for ecotoxicology studies even after years of culturing in laboratory conditions. It is advisable that other laboratories with long-term datasets made their own control charts, to allow data comparison and to improve test protocols. Considering our HCD, we suggest an EC50 of NiCl2 of 0.14 ± 0.09 mg Ni/L for acute tests with the Mediterranean strain of A. tonsa.

Comparison of different ecotoxicological batteries with WOE approach for the environmental quality evaluation of harbour sediments.

This study was conducted under the Italian Ministerial Decree D.M. 173/2016 which regulates the assessment of the Sediment Class Quality in Italy using ecotoxicological bioassay and chemical analysis (Weight-Of-Evidence model). The aim of this work was to evaluate the real classification obtained by the theoretically equivalent responses of nine different combinations of batteries based on six different species: Aliivibrio fischeri (inhibition of bioluminescence), Phaeodactylum tricornutum, Skeletonema costatum, Dunaliella tertiolecta (inhibition of algal growth), Paracentrotus lividus and Crassostrea gigas (embryotoxicity). Bioassays, in many cases, showed a non-bioavailability effect of the pollutants; these one highly revealed by the chemical analyses. Algal species showed responses very similar from each other. Otherwise, species used for embryotoxicity produced wide responses, consequently modifying the quality class of sediments and the handling management (i.e. landfill confinement or beach nourishment) allowed by the Law.

Cellular Responses Induced by Zinc in Zebra Mussel Haemocytes. Loss of DNA Integrity as a Cellular Mechanism to Evaluate the Suitability of Nanocellulose-Based Materials in Nanoremediation.

Zinc environmental levels are increasing due to human activities, posing a threat to ecosystems and human health. Therefore, new tools able to remediate Zn contamination in freshwater are highly recommended. Specimens of Dreissena polymorpha (zebra mussel) were exposed for 48 h and 7 days to a wide range of ZnCl2 nominal concentrations (1-10-50-100 mg/L), including those environmentally relevant. Cellulose-based nanosponges (CNS) were also tested to assess their safety and suitability for Zn removal from freshwater. Zebra mussels were exposed to 50 mg/L ZnCl2 alone or incubated with 1.25 g/L of CNS (2 h) and then removed by filtration. The effect of Zn decontamination induced by CNS has been verified by the acute toxicity bioassay Microtox®. DNA primary damage was investigated by the Comet assay; micronuclei frequency and nuclear morphological alterations were assessed by Cytome assay in mussels' haemocytes. The results confirmed the genotoxic effect of ZnCl2 in zebra mussel haemocytes at 48 h and 7-day exposure time. Zinc concentrations were measured in CNS, suggesting that cellulose-based nanosponges were able to remove Zn(II) by reducing its levels in exposure waters and soft tissues of D. polymorpha in agreement with the observed restoration of genetic damage exerted by zinc exposure alone.

EBV-Driven Lymphoproliferative Disorders and Lymphomas of the Gastrointestinal Tract: A Spectrum of Entities with a Common Denominator (Part 3).

EBV is the first known oncogenic virus involved in the development of several tumors. The majority of the global population are infected with the virus early in life and the virus persists throughout life, in a latent stage, and usually within B lymphocytes. Despite the worldwide diffusion of EBV infection, EBV-associated diseases develop in only in a small subset of individuals often when conditions of immunosuppression disrupt the balance between the infection and host immune system. EBV-driven lymphoid proliferations are either of B-cell or T/NK-cell origin, and range from disorders with an indolent behavior to aggressive lymphomas. In this review, which is divided in three parts, we provide an update of EBV-associated lymphoid disorders developing in the gastrointestinal tract, often representing a challenging diagnostic and therapeutic issue. Our aim is to provide a practical diagnostic approach to clinicians and pathologists who face this complex spectrum of disorders in their daily practice. In this part of the review, the chronic active EBV infection of T-cell and NK-cell type, its systemic form; extranodal NK/T-cell lymphoma, nasal type and post-transplant lymphoproliferative disorders are discussed.

Sea cucumber Holothuria polii (Delle Chiaje, 1823) as new model for embryo bioassays in ecotoxicological studies.

The sea cucumbers are common members of marine benthic communities, widespread distributed, easily available and handled. Nevertheless, no data are available on embryo toxicity assays using sea cucumbers, despite some of these species could fully meet the requirements for model test organisms. Holothuria polii is a key species in soft sediments and seagrass meadows; the aim of the present study was the standardization of a new embryo bioassay with this species, as an ecologically relevant test to evaluate the effects of environmental stressors. Sequential experiments were carried out, allowing to define the test acceptability, and a minimum sample size of 240 embryos. Temperature of 26 °C, salinity at 36‰ and a density of 60 eggs/ml were identified as optimum experimental conditions for performing the bioassay. The EC50 calculated for Cd2+ and Cu2+ in dose-response experiments indicated a good sensitivity of H. polii, with comparable values with those obtained in embryo toxicity bioassays of other marine invertebrates. An Integrative Toxicity Index (ITI) was calculated by integrating the frequency of abnormal embryos with the severity of observed abnormalities. The index allows to better discriminate different levels of toxicity, appearing particularly relevant for validating the usefulness of H. polii in embryo assays and ecotoxicological studies on environmental quality.

Integrated characterization and risk management of marine sediments: The case study of the industrialized Bagnoli area (Naples, Italy).

The aim of the present work is to demonstrate the practical importance of a multidisciplinary approach and weighted criteria to synthesize and integrate different typologies of data (or lines of evidence, LOEs), including chemical levels in marine sediments, their bioavailability to specific indicator species, ecotoxicological effects measured through subcellular biomarkers and batteries of bioassays, and potential impacts of pollution on local benthic communities. The area of Bagnoli (Gulf of Naples, Southern Italy) was selected as a model case-study, as it is a coastal area chronically impacted by massive industrial contamination (trace metals and hydrocarbons), and dismissed decades ago without any subsequent remediation or habitat restoration. The results of each LOE were elaborated to provide specific hazard indices before their overall integration in a weight of evidence (WOE) evaluation. Levels of some trace metals and PAHs revealed a severe contamination in the entire study area. Bioavailability of hydrocarbons was evident particularly for high molecular weight PAHs, which also caused significant variations of cellular biomarkers, such as cytochrome P450 metabolization in fish, lysosomal membrane destabilization in mussels, genotoxic effects both in fish and molluscs. The results of a battery of bioassays indicated less marked responses compared to those obtained from chemical and biomarkers analyses, with acute toxicity still present in sediments close to the source of contamination. The analysis of benthic assemblages showed limited evidence of impact in the whole area, indicating a good functioning of local ecosystems at chronic contamination. Overall, the results of this study confirm the need of combining chemical and biological data, the quantitative characterization of various typologies of hazard and the importance of assessing an integrated environmental WOE risk, to orientate specific and scientifically-supported management options in industrialized areas.

In situ embryo toxicity test with sea urchin: Development of exposure chamber for test execution.

In situ toxicity tests represent a technique rarely performed owing to the lack of standard methodologies or to technical or economic problems. Nevertheless, its application would allow for a more realistic interpretation of pollution effects than those obtained by standard laboratory tests. The goal of this study is to develop and validate a specific exposure chamber for in situ exposition of Paracentrotus lividus embryos to obtain a defined methodology to perform the sea urchin embryo toxicity test in field conditions. After a first part of the study to verify the feasibility of the test chamber and the methodology, this approach was used as a tool to investigate whether the cruise ship "Costa Concordia" shipwrecked on Giglio Island (Tuscany, Italy), could have acted as a source of pollution. The results obtained for in situ tests showed, on average, percentages of normal embryos lower than those obtained in laboratory conditions and a greater sensitivity than for those obtained in the laboratory owing to the time-integration of results. Thus the exposure chamber and the in situ methodology so far developed would appear to be suitable tools for future application in the environmental quality evaluation of marine waters.