Ultrastructural changes, pigment responses and bioaccumulation in the microalga Phaeodactylum tricornutum Bohlin exposed to BPA analogues.

As well-known, microalgae have a pivotal role in aquatic environments, being the primary producer. In this study, we investigated the effects of Bisphenol A (BPA) analogues on cell ultrastructure, reactive oxygen species (ROS) production and photosynthetic pigment responses in the diatom Phaeodactylum tricornutum. Microalgae were exposed during both exponential and stationary growth phases to an environmental relevant concentration (300 ng/L) of three differing BPA analogues (BPAF, BPF, and BPS) and their mixture (100 ng/L of each compound). Bioaccumulation of such compounds in microalgae was also analysed. During the stationary growth phase, a significant increase in the percentage of cells with hydrogen peroxide production was recorded after exposure to both BPS and MIX. Conversely, no significant effects on total chlorophylls and carotenoids were observed. During exponential growth phase we observed that control cultures had chloroplasts with well-organized thylakoid membranes and a central pyrenoid. On the contrary, the culture cells treated with BPA analogues and MIX showed chloroplasts characterized by evident dilation of thylakoid membranes. The presence of degeneration areas in the cytoplasm was also recorded. During the stationary growth phase, control and culture cells were characterized by chloroplasts with a regular thylakoid system, whereas BPA analogues-exposed cells were characterized by a deep degradation of the cytoplasm but showed chloroplasts without evident alterations of the thylakoid system. Lipid bodies were visible in treated microalgae. Lastly, microalgae bioaccumulated mainly BPS and BPF, alone or in the MIX. Overall, results obtained revealed that BPA analogues can affect some important biochemical and ultrastructure features of microalgae, promoting ROS production. Lastly, the capability of microalgae to bioaccumulate bisphenols suggest a potential ecotoxicological risk for filter-feeders organisms.

A multibiomarker approach in clams (Ruditapes philippinarum) for a toxicological evaluation of dredged sediments.

The Lagoon of Venice is often dredged for channel maintenance. To avoid harmful consequences to the ecosystem, a proper disposal of bottom sediments requires a preliminary evaluation of its potential toxicity before excavation. Here we evaluated the effects of polluted sediments on clams (Ruditapes philippinarum) using a multibiomarker approach. Bivalves were exposed for 3 and 14 days to five sediment samples collected along a navigation canal between Venice historical centre and the industrial area of Porto Marghera. Immunological, antioxidant, detoxification, and neurotoxicity biomarkers were analysed in haemolymph, gill, and digestive gland. As a control, sediment collected far from pollution sources was used. Two experiments were performed to assess potential seasonal/gametogenic influence in clam sensitivity. A different response of clam biomarkers was observed during the two experiments and among sampling sites. Clams' digestive gland resulted to be the most sensitive tissue analysed showing significant differences among sites in all biomarkers analysed. Greater differences were present due to seasonality rather than exposure. The concentrations of metals and organic pollutants increased from the city centre to the industrial area, highlighting the influence that industrial activities had on the lagoon ecosystem. However, bioaccumulation in clams did not follow the same clear pattern, suggesting low bioavailability of compounds due to relatively high organic matter content. Biomarkers modulation was mainly driven by metals, both present in sediments and bioaccumulated. In comparison, effects of organic pollutants on the biomarkers tested were negligible. Other sources of contamination not investigated (e.g. pesticides) were suggested by neurotoxicity biomarkers alteration.

Contaminants from dredged sediments alter the transcriptome of Manila clam and induce shifts in microbiota composition.

BACKGROUND: The reuse of dredged sediments in ports and lagoons is a big issue as it should not affect the quality and the equilibrium of ecosystems. In the lagoon of Venice, sediment management is of crucial importance as sediments are often utilized to built-up structures necessary to limit erosion. However, the impact of sediment reuse on organisms inhabiting this delicate area is poorly known. The Manila clam is a filter-feeding species of high economic and ecological value for the Venice lagoon experiencing a drastic decline in the last decades. In order to define the molecular mechanisms behind sediment toxicity, we exposed clams to sediments sampled from different sites within one of the Venice lagoon navigable canals close to the industrial area. Moreover, we investigated the impacts of dredged sediments on clam's microbial communities. RESULTS: Concentrations of the trace elements and organic chemicals showed increasing concentrations from the city of Venice to sites close to the industrial area of Porto Marghera, where PCDD/Fs and PCBs concentrations were up to 120 times higher than the southern lagoon. While bioaccumulation of organic contaminants of industrial origin reflected sediments' chemical concentrations, metal bioaccumulation was not consistent with metal concentrations measured in sediments probably due to the activation of ABC transporters. At the transcriptional level, we found a persistent activation of the mTORC1 signalling pathway, which is central in the coordination of cellular responses to chemical stress. Microbiota characterization showed the over-representation of potential opportunistic pathogens following exposure to the most contaminated sediments, leading to host immune response activation. Despite the limited acquisition of new microbial species from sediments, the latter play an important role in shaping Manila clam microbial communities. CONCLUSIONS: Sediment management in the Venice lagoon will increase in the next years to maintain and create new canals as well as to allow the operation of the new mobile gates at the three Venice lagoon inlets. Our data reveal important transcriptional and microbial changes of Manila clams after exposure to sediments, therefore reuse of dredged sediments represents a potential risk for the conservation of this species and possibly for other organisms inhabiting the Venice lagoon.

Ecotoxicological effects and bioaccumulation of BPA analogues and their mixture in the clam Ruditapes philippinarum.

Bisphenol A is recognized as an endocrine disruptor that can affect several biological processes in marine species. Consequently, its use has been restricted and it has been replaced with other similar compounds named bisphenol A analogues (BPA analogues). BPA analogues are speculatively considered safer compounds than BPA and their usage is increasing with a consequent higher environmental release. In this study, specimens of the clam Ruditapes philippinarum were exposed to three main BPA analogues, namely BPAF, BPF, BPS and their mixture at an environmentally relevant concentration of 300 ng/L for 7 and 14 days. Effects on biomarkers indicative of cytotoxicity, oxidative stress and damage and neurotoxicity were evaluated. In addition, bioaccumulation of the compound tested was analysed in clam soft tissues. Results showed that BPA analogues at an environment concentration affected cellular parameters and antioxidant system causing also oxidative damage, suggesting that BPA analogues can be harmful compounds for clams.

Can BPA Analogs Affect Cellular and Biochemical Responses in the Microalga Phaeodactylum tricornutum Bohlin?

Bisphenol A analogs (BPA analogs) are emerging contaminants with a rising production caused by the replacement of BPA with these compounds. The increased production of BPA analogs is leading to their increased release into various ecosystems, including marine ones. The aim of this study was to evaluate the biological effects of BPA analogs on a primary producer, the diatom Phaeodactylum tricornutum Bohlin. Three different BPA analogs (BPAF, BPF, and BPS) and their mixture were tested at the environmental relevant concentration of 300 ng/L. Growth, cell size and several biomarkers of oxidative stress and oxidative damage were measured. Our results indicated that the tested compounds caused a reduced growth rate and induced oxidative stress, altering many antioxidant enzymes in P. tricornutum. However, no oxidative damages were observed.

Multidisciplinary long-term survey of Manila clam grown in farming sites subjected to different environmental conditions.

In recent years recurrent bivalve mass mortalities considerably increased around the world, causing the collapse of natural and farmed populations. Venice Lagoon has historically represented one of the major production areas of the Manila clam Ruditapes philippinarum in Europe. However, in the last 20 years a 75 % decrease in the annual production has been experienced. While climate change and anthropogenic interventions may have played a key role in natural and farmed stocks reductions, no studies investigated at multiple levels the environmental stressors affecting farmed Manila clam to date. In this work we carried out a long-term monitoring campaign on Manila clam reared in four farming sites located at different distances from the southern Venice Lagoon inlet, integrating (meta)genomic approaches (i.e. RNA-seq; microbiota characterization), biometric measurements and chemical-physical parameters. Our study allowed to characterize the molecular mechanisms adopted by this species to cope with the different environmental conditions characterizing farming sites and to propose hypotheses to explain mortality events observed in recent years. Among the most important findings, the disruption of clam's immune response, the spread of Vibrio spp., and the up-regulation of molecular pathways involved in xenobiotic metabolism suggested major environmental stressors affecting clams farmed in sites placed close to Chioggia's inlet, where highest mortality was also observed. Overall, our study provides knowledge-based tools for managing Manila clam farming on-growing areas. In addition, the collected data is a snapshot of the time immediately before the commissioning of MoSE, a system of mobile barriers aimed at protecting Venice from high tides, and will represent a baseline for future studies on the effects of MoSE on clams farming and more in general on the ecology of the Venice Lagoon.

A first insight into haemocytes of Pinctada imbricata radiata: A morpho-functional characterization.

The pearl oyster Pinctada imbricata radiata (Leach, 1814), from the Pacific Ocean, was one of the first species to reach via Suez the Mediterranean, colonizing the eastern basin and recently spreading to the western. The species showed to be able to adapt to a wide range of climatic, hydrological, and ecological conditions. Since 2000 it reached the Strait of Messina, where is now infesting the transitional waters of the oriented natural reserve "Laguna di Capo Peloro." Due to such resistance and adaptation ability, various assays were performed. Haemocyte morpho-functional aspects were evaluated in haemolymph samples fixed with 1% and 2% glutaraldehyde for optical and electron microscopy (TEM). The following assays were carried out: cell characterization using several dyes, detection of intra- and extracellular lipids, the capability of phagocytosis using the yeast Saccharomyces cerevisiae and to produce superoxide anion (O2- ). Detection of several enzymes, such as acid and alkaline phosphatase, arylsulfatase, chloro-acetylesterase and ß-glucuronidase was also assessed. Cell count was demonstrated to be abundant with a mean of 8.263 × 106 mm2 ± 0.935 × 106 (SD). Two main cell populations were noticed: granulocytes and hyalocytes, both competent for phagocytosis, to produce O2- , and characterized by lipids. Based on the granule analysis, enzymatic activity was also demonstrated. The observations under TEM confirmed all the results obtained. This study supports the hypothesis that P. imbricata radiata can be usefully employed as a model organism in environmental biomonitoring. Moreover, since the species represent potential threats to native species and ecosystems, further insights into its biological adaptations in invaded ecosystems are recommended.

Identification of haemocytes and histological examination of gills of the spiny oyster Spondylus gaederopus (Linnaeus, 1758).

In the framework of investigations aimed to detect new available bioindicators in marine environment, haemolymph cells and ctenidia of the Mediterranean spiny oyster, Spondylus gaederopus, have been investigated. Haemocyte count and characterisation, phagocytosis and superoxide anion production and enzyme activity assays, have been carried out. TEM observations have been performed. After gross anatomy observations, cito-histological determinations have been carried out, especially focused on ctenidia structure and function. Main results concerned the relatively low number of circulating cells, and the rich in granules granulocytes, most of which were lysosomes. Release of lysosomal enzymes was confirmed a shared trait inside bivalves. Glycogen deposits as probable result of conversion of bacteria carbohydrates, have been detected, as well as the occurrence of both acidophilic and basophilic haemocytes. Phagocytosis, both in granulocytes and agranulocytes, has been recorded, together with the production of superoxide anion. Haemocytes were found positive to acid phosphatase, alkaline phosphatase, ß-glucuronidase, chloroacetylesterase and arylsulphatase. Ctenidia showed a complex organization, including two demibranch to each ctenidium, two different kinds of lamellae filament and specialized structures as ciliated disks connecting filaments in "eutherorhabdic ctenidia". The occurrence of three different types of mucous cells in the same region of ordinary filaments has been underlined. Such features, suggesting high resistance to environmental stress and disease, allow to consider spiny oysters as promising bioindicators, although deserving of further investigations to evaluate the physiological responses to stress in controlled conditions. Present data, moreover, providing basic information on the biology of S. gaederopus, notably implement the present knowledge on the Mediterranean spiny oysters, whose under-evaluated ecological role should be carefully considered.

Zinc oxide, titanium dioxide and C60 fullerene nanoparticles, alone and in mixture, differently affect biomarker responses and proteome in the clam Ruditapes philippinarum.

Continuous release of nanoparticles (NPs) into marine coastal environments results in an increased risk of exposure to complex NP mixtures for marine organisms. However, to date, the information on the effects at molecular and biochemical levels induced by the exposure to NPs, singly and as a mixture, is still scant. The present work aimed at exploring the independent and combined effects and the mechanism(s) of action induced by 7-days exposure to 1 µg/L nZnO, 1 µg/L nTiO2 and 1 µg/L FC60 fullerene in the Manila clam Ruditapes philippinarum, using a battery of immunological and oxidative stress biomarkers in haemolymph, gills and digestive gland. In addition, proteomics analyses were performed in gills and the digestive gland, where NP bioaccumulation was also assessed. Increased bioaccumulation of single NPs and the mixture was linked with increased oxidative stress and higher damage to proteins, lipids and DNA in all tissues analysed. The proteomics approach highlighted protein modulation in terms of abundance and damage (higher redox-thiol and carbonylated groups content). In particular, the modulated proteins (16 in gills and 18 in digestive gland) were mostly related to cytoskeleton and energetic metabolism. The digestive gland was the tissue more affected. For all biomarkers measured, increased detrimental effects were observed in the mixture compared to single NP exposures.

Dinitroaniline herbicide pendimethalin affects development and induces biochemical and histological alterations in zebrafish early-life stages.

Pendimethalin (PND) is a dinitroaniline preemergent herbicide widely used to control grasses and weeds. The present study aimed to evaluate the PND potential effects on the development of zebrafish early-life stages. The research focuses first on acute toxicity, followed by the integration of toxicity results through histopathology, oxidative status, and neurotoxicity evaluation of sublethal and environmentally relevant concentrations. Zebrafish larvae exposed to PND showed mortality and developed sublethal alterations including impaired fin development, lordosis, scoliosis, blood congestion, impaired blood flow, and reduced heartbeat. PND exposure (0.5 mg/L) affects musculoskeletal development leading to delayed and reduced ossification of the vertebral centra in the developing vertebral column and disruption of muscle morphology. Herbicide exposure (0.5 mg/L and 0.05 mg/L) led also to biochemical changes of antioxidant enzymes, increasing the activity of CAT, GR, and GPx, while no effects were observed on the activity of SOD and GST in zebrafish larvae. Lastly, AChE activity, a biochemical marker of neurotoxicity, was also increased in zebrafish larvae exposed to 0.5 mg/L of PND. These results confirm the developmental toxicity of PND in zebrafish early-life stages, pointing out the potential role of oxidative stress in the onset of sublethal alterations.

Effects of long-term exposure of Mytilus galloprovincialis to thiacloprid: A multibiomarker approach.

Thiacloprid is a neonicotinoid insecticide widely exploited in agriculture and easily mobilized towards aquatic environments by atmospheric agents. However, little information about its toxicological effects on aquatic invertebrate bioindicators is available. In this study, specimens of the mussel Mytilus galloprovincialis were exposed to thiacloprid at environmental (4.5 µg L-1) and 100 times higher than environmental (450 µg L-1) concentrations for 20 days. Thiacloprid affected haemolymph biochemical parameters, cell viability in the digestive gland, antioxidant biomarkers and lipid peroxidation in the digestive gland and gills at environmentally relevant concentrations (4.5 µg L-1). In addition, thiacloprid exposure caused histological damage to the digestive gland and gills. Interestingly, the pesticide was detected at levels equal to 0.14 ng g-1 in the soft tissues of sentinels exposed for 20 days to 450 µg L-1 thiacloprid in seawaterµ. Due to its harmful potential and cumulative effects after long-term exposure of M. galloprovincialis, thiacloprid may pose a potential risk to nontarget aquatic organisms, as well as to human health. This aspect requires further in-depth investigation.

First Evidence of In Vitro Effects of C6O4-A Substitute of PFOA-On Haemocytes of the Clam Ruditapes philippinarum.

Alternative chemicals to per- and poly-fluoroalkyl substances have recently been introduced in various industrial processes. C6O4 (difluoro{[2,2,4,5-tetrafluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}acetic acid) is a new surfactant and emulsifier used as a replacement for perfluorooctanoic acid (PFOA). From an ecotoxicological point of view, in vitro assays are useful tools for assessing the negative effects and understanding the mechanisms of action of chemicals at the cellular level. Here, we present the results of an in vitro study in which the effects of C6O4 were evaluated-for the first time-on haemocytes of the clam Ruditapes philippinarum. Cells were exposed to three concentrations of C6O4 (0.05, 0.5, 5 µg/mL) and the effects on haemocyte viability, haemocyte morphology, differential haemocyte count, lysosomal membrane stability, superoxide anion production, acid phosphatase, and ß-glucuronidase activities, as well as on the percentage of micronuclei and chromosomal aberrations were evaluated. The results demonstrated that C6O4 significantly affected haemocyte morphology, lysosomal membrane stability, hydrolytic enzyme activity, and superoxide anion production, and promoted chromosomal aberrations. To the best of our knowledge, this is the first study revealing the in vitro effects of C6O4, a substitute for PFOA, on haemocytes from a bivalve species.

New compounds, old problems. The case of C6O4 - a substitute of PFOA - and its effects to the clam Ruditapes philippinarum.

C6O4 (difluoro{[2,2,4,5-tetrafluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}acetic acid) is a new surfactant and emulsifier used as a substitute of perfluorooctanoic acid (PFOA). Recently, C6O4 has been detected in aquatic environments, but, at present, no information concerning the effects of C6O4 on aquatic species, such as bivalves, are available in the literature. Therefore, in this study we evaluated for the first time the effects of C6O4 (0.1 and 1 µg/L) and PFOA (1 µg/L) to the clam Ruditapes philippinarum. Short-term (7 days) and long-term (21 days) exposures of clams to the two compounds were carried out and numerous biomarkers were measured in haemocytes/haemolymph, as well as in gills and digestive gland. The MANOVA analysis demonstrated statistically significant effects of the independent variables "treatment", "time" and "treatment-time interaction" on the whole dataset of biomarker responses. The two-way ANOVA analysis performed for each biomarker response indicated that the two compounds affected most of the cellular and tissue parameters measured. Despite preliminary, the results obtained suggested that C6O4 - similarly to PFOA - can affect both cellular and biochemical parameters of clams.

The new PFAS C6O4 and its effects on marine invertebrates: First evidence of transcriptional and microbiota changes in the Manila clam Ruditapes philippinarum.

There is growing concern for the wide use ofperfluorooctanoic acid (PFOA) because of its toxic effects on the environment and on human health. A new compound - the so called C6O4 (perfluoro ([5-methoxy-1,3-dioxolan-4-yl]oxy) acetic acid) - was recently introduced as one of the alternative to traditional PFOA, however this was done without any scientific evidence of the effects of C6O4 when dispersed into the environment. Recently, the Regional Agency for the Protection of the Environment of Veneto (Italy) detected high levels of C6O4 in groundwater and in the Po river, increasing the alarm for the potential effects of this chemical into the natural environment. The present study investigates for the first time the effects of C6O4 on the Manila clam Ruditapes philippinarum exposed to environmental realistic concentrations of C6O4 (0.1 µg/L and 1 µg/L) for 7 and 21 days. Furthermore, in order to better understand if C6O4 is a valid and less hazardous alternative to its substitute, microbial and transcriptomic alterations were also investigated in clams exposed to 1 µg/L ofPFOA. Results indicate that C6O4 may cause significant perturbations to the digestive gland microbiota, likely determining the impairment of host physiological homeostasis. Despite chemical analyses suggest a 5 times lower accumulation potential of C604 as compared to PFOA in clam soft tissues, transcriptional analyses reveal several alterations of gene expression profile. A large part of the altered pathways, including immune response, apoptosis regulation, nervous system development, lipid metabolism and cell membrane is the same in C6O4 and PFOA exposed clams. In addition, clams exposed to C6O4 showed dose-dependent responses as well as possible narcotic or neurotoxic effects and reduced activation of genes involved in xenobiotic metabolism. Overall, the present study suggests that the potential risks for marine organism following environmental contamination are not reduced by replacing PFOA with C6O4. In addition, the detection of both C6O4 and PFOA into tissues of clams inhabiting the Lagoon of Venice - where there are no point sources of either compounds - recommends a similar capacity to spread throughout the environment. These results prompt the urgent need to re-evaluate the use of C6O4 as it may represent not only an environmental hazard but also a potential risk for human health.

Effects of a mixture of glyphosate, 17α-ethynylestradiol and amyl salicylate on cellular and biochemical parameters of the mussel Mytilus galloprovincialis.

In this study the effects of a mixture of glyphosate (herbicide), 17a-ethinylestradiol (synthetic estrogen) and amyl salicylate (fragrance) to the mussel Mytilus galloprovincialis were evaluated. Mussels were exposed for 7 days to two realistic concentrations of the mixture (10 and 100 ng/L) and the effects on total haemocyte counts, haemocyte diameter and volume, haemocyte proliferation, haemolymph lactate dehydrogenase activity and haemocyte lysate lysozyme activity were measured. In addition, superoxide dismutase, catalase, acetylcholinesterase, glutathione-S-transferase and glutathione reductase activities were measured in gills and digestive gland. The survival-in-air test was also performed. Results demonstrated that the mixture affected both cellular and biochemical biomarkers, but not tolerance to aerial exposure of M. galloprovincialis. The negative effects recorded in this study suggested that more efforts should be done to assess the ecotoxicological risks posed by contaminant mixture to aquatic invertebrates.

Toxicological effects and bioaccumulation of fullerene C60 (FC60) in the marine bivalve Ruditapes philippinarum.

Fullerene C60 (FC60), with its unique physical properties, has been used in many applications in recent decades. The increased likelihood of direct release into the environment has raised interest in understanding the biological effects of FC60 to aquatic organisms. Nowadays, only few studies have analysed FC60 effects and bioaccumulation in marine organisms following in vivo exposure. To provide new data about FC60 toxicity, Ruditapes philippinarum was selected as target species to assess potential adverse effects of the contaminant. Clams were exposed for 1, 3 and 7 days to predicted environmental concentrations of FC60 (1 and 10 µg/L) and cellular and biochemical responses were evaluated in clams' gills, digestive gland and haemolymph. The FC60 content in gills and digestive gland was determined in all experimental conditions after 7 days of exposure. Results showed an increase in oxidative stress. In particular, a significant modulation in antioxidant enzyme activities, and changes in glutathione S-transferase activity were observed in gills. Moreover, damage to lipids and proteins was detected in FC60-treated (10 µg/L) clams. In digestive gland, slighter variations in antioxidant enzyme activities and damage to molecules were detected. CAT activity was significantly affected throughout the exposure, whereas damage to lipids was evident only at the end of exposure. FC60 accumulation was revealed in both gills and digestive gland, with values up to twelve-fold higher in the latter. Interestingly, haemolymph parameters were slightly affected by FC60 compared to the other tissues investigated. Indeed, only Single Cell Gel Electrophoresis and Neutral Red uptake assays showed increased values in FC60-exposed clams. Moreover, volume and diameter of haemocytes, haemocyte proliferation, and micronucleus assay highlighted significant variations in treated clams, but only in the first phases of exposure, and no changes were detected after 7 days. Our results suggested clam gills as the target tissue for FC60 toxicity under the exposure conditions tested: the high damage detected to lipids and proteins could contribute to long-term problems for the organism.

Acute effects of neonicotinoid insecticides on Mytilus galloprovincialis: A case study with the active compound thiacloprid and the commercial formulation calypso 480 SC.

Pesticides can enter aquatic environments potentially affecting non-target organisms. Unfortunately, the effects of such substances are still poorly understood. This study investigated the effects of the active neonicotinoid substance thiacloprid (TH) and the commercial product Calypso 480 SC (CA) (active compound 40.4% TH) on Mytilus galloprovincialis after short-term exposure to sublethal concentrations. Mussels were tested for seven days to 0, 1, 5 and 10 mg L-1 TH and 0, 10, 50 and 100 mg L-1 CA. For this purpose, several parameters, such as cell viability of haemocytes and digestive cells, biochemical haemolymph features, superoxide dismutase (SOD) and catalase (CAT) enzymatic activity of gills and digestive gland, as well as histology of such tissues were analysed. The sublethal concentrations of both substances lead to abatement or completely stopping the byssal fibres creation. Biochemical analysis of haemolymph showed significant changes (P < 0.01) in electrolytes ions (Cl-, K+, Na+, Ca2+, S-phosphor), lactate dehydrogenase (LDH) enzyme activity and glucose concentration following exposure to both substances. The TH-exposed mussels showed significant imbalance (P < 0.05) in CAT activity in digestive gland and gills. CA caused significant decrease (P < 0.05) in SOD activity in gills and in CAT activity in both tissues. Results of histological analyses showed severe damage in both digestive gland and gills in a time- and concentration-dependent manner. This study provides useful information about the acute toxicity of a neonicotinoid compound and a commercial insecticide on mussels. Nevertheless, considering that neonicotinoids are still widely used and that mussels are very important species for marine environment and human consumption, further researches are needed to better comprehend the potential risk posed by such compounds to aquatic non-target species.

Do males and females respond differently to ocean acidification? An experimental study with the sea urchin Paracentrotus lividus.

Seawater pH lowering, known as ocean acidification, is considered among the major threats to marine environment. In this study, post-spawning adults of the sea urchin Paracentrotus lividus were maintained at three pH values (8.0, 7.7, 7.4) for 60 days. Physiological, biochemical, cellular, behavioural and reproductive responses were evaluated in males and females. Significant differences between sexes were observed, with higher ammonia excretion and lower catalase activity in males. Respiration rate (after 21 days), catalase activity in gonads and total coelomocyte count showed the same increasing trend in males and females under low pH. Ammonia excretion, gonadosomatic index and lysozyme activity exhibited opposite responses to low pH, with an increasing trend in males and decreasing in females. Results demonstrated that exposure to low pH could result in different response strategies of male and female sea urchins at a physiological, biochemical and immunological level. Reduced female gonadosomatic index under low pH suggested decreased energy investment in reproduction.

Molecular and biochemical responses of vitellogenin in the mussel Mytilus galloprovincialis exposed to the glyphosate-based herbicide Roundup® Power 2.0.

Glyphosate-based herbicides (GBHs) occur in aquatic ecosystems at concentrations of hundreds of micrograms per liter. As formulation adjuvants are suspected to be endocrine-disrupting chemicals, we assessed the effects of the recent GBH formulation Roundup® Power 2.0 on vitellogenin (VTG) in Mytilus galloprovincialis. Mussels were exposed for 7, 14, and 21 days to two concentrations of the commercial formulation, corresponding to 100 and 1000 µg/L of glyphosate. The expression of the vtg gene in gonads of females and males, as well as the levels of alkali labile phosphates (ALP) in gonads and non-gonadal tissues from the two sexes were measured. No significant alterations were observed in vtg expression values during the exposure. Conversely, a significant reduction in gonadal ALP levels was observed in females exposed for 21 days and in males exposed for 7 days. In addition, ALP levels increased significantly in gonads from males exposed for 21 days to the two concentrations of Roundup®. As for non-gonadal tissues, ALP levels did not change significantly in females, whereas ALP levels decreased significantly in non-gonadal tissues from males exposed for 21 days to the lowest concentration tested. An overall statistically significant difference in ALP levels was found between females and males. Although preliminary, our study suggests that GBH can affect reproduction-related parameters in mussels.

Assessing the effects of neonicotinoid insecticide on the bivalve mollusc Mytilus galloprovincialis.

In the present work, the marine invertebrate Mytilus galloprovincialis was used as model organism to evaluate the toxic effects of the neonicotinoid Calypso 480 SC (CAL) following 20 days of exposure to sub-lethal concentrations of 7.77 mg L-1 (0.1% 96 h-LC50) and 77.70 mg L-1 (1% 96 h-LC50), and a recovery period of 10 days in uncontaminated seawater. Results revealed that exposure to both concentrations of CAL increased significantly mortality rate in the cells of haemolymph and digestive gland, while digestive gland cells were no longer able to regulate cell volume. Exposure significantly reduced haemolymph parameters (Cl-, Na+), affected the enzymatic activities of superoxide dismutase of digestive gland and catalase of gill, and caused also histopathological alterations in digestive gland and gills. Main histological damages detected in mussels were lipofuscin accumulation, focal points of necrosis, mucous overproduction and infiltrative inflammations. Interestingly, alterations persisted after the recovery period in CAL-free water, especially for haemocyte parameters (K+, Na+, Ca2+, lactate dehydrogenase, glucose). A slight recovery of histological conditions was detected. These findings suggested that sub-chronic exposure to the neonicotinoid insecticide caused significant alterations in both cell and tissue parameters of M. galloprovincialis. Considering the ecologically and commercially important role of mussels in coastal waters, a potential risk posed by neonicotinoids to this essential aquatic resource can be highlighted.