This investigation using a molluscan animal model tested the hypothesis that experimentally induced lysosomal autophagy protects against oxidative cell injury. Induction of augmented lysosomal autophagy has previously been implicated in this protective process. Four treatment groups of blue mussels (Mytilus galloprovincialis) were used: Group 1 (fed - control), Group 2 (fasted), Group 3 (copper + fed) and Group 4 (copper + fasted). Groups 2 and 4 were fasted in order to trigger autophagy; and samples of hepatopancreas (liver analogue or digestive gland) from all 4 groups were taken at 3, 6 and 15 days. Treatment with copper provided a positive reference for oxidative stress: Groups 3 and 4 were treated with copper (10 μg Cu2+/animal/day) for three days only. Oxidative damage and cellular injury in hepatopancreatic digestive cells was found to decrease in Group 2 (fasted) compared to Group 1 (fed - control). Group 3 (fed + copper) showed clear evidence of oxidative stress and cell injury, as well as induction of antioxidant activities. Group 4 (copper + fasted) had a reduced uptake of copper and toxicity of copper was also reduced, compared with Group 3. It was concluded that augmented autophagy had a hormetic cytoprotective anti-oxidant effect.

Autophagy is a highly conserved evolutionary survival or defence process that enables cells and organisms to survive periods of environmental stress by breaking down cellular organelles and macromolecules in autolysosomes to provide a supply of nutrients for cell maintenance. However, autophagy is also a part of normal cellular physiology that facilitates the turnover of cellular constituents under normal conditions: it can be readily augmented by mild environmental stress; but becomes dysfunctional with severe oxidative stress leading to cellular pathology. The molluscan hepatopancreas or digestive gland provides a versatile and environmentally relevant model to investigate lysosomal autophagy and stress-induced dysfunctional autophagy. This latter process has been implicated in many animal and human disease conditions, including degenerative and neurodegenerative diseases, as well as obesity related conditions. Many environmental pollutants have also been found to induce dysfunctional autophagy in molluscan hepatopancreatic digestive cells, and in this study, the marine blue mussel Mytilus galloprovincialis was exposed for 7 days to: 0.1 μM, 1 μM and 10 μM concentrations of fluoranthene and phenanthrene (PAHs); chlorpyrifos and malathion (organophosphorus compounds); atrazine (triazine herbicide); copper (transition metal) and dodecylbenzene sulphonic acid (LAS, surfactant). The marine snail or periwinkle, Littorina littorea, was also exposed to phenanthrene, chlorpyrifos and copper. Indices of oxidative stress, cell injury and dysfunctional autophagy were measured (i.e., lysosomal membrane stability, protein carbonyls, lipofuscin, and lysosomal accumulation of lipid or lipidosis). Evidence of oxidative stress, based on the elevation of lipofuscin and protein carbonyls, was found for all compounds tested; with chlorpyrifos being the most toxic to both species. Dysfunctional autophagy was induced by all of the compounds tested in both species, except for atrazine in mussels. This failure of normal autophagy was consistently associated with oxidative stress. Autophagic dysfunction is an important emerging feature in the aetiology of many disease conditions in animals and humans; and an explanatory conceptual mechanistic model has been developed for dysregulation of autophagy in response to oxidative stress.

A mesocosm experiment was conducted to quantify the relationships between the presence and body size of two burrowing heart urchins (Brissopsis lyrifera and Echinocardium cordatum) and rates of sediment nutrient flux. Furthermore, the impact of seawater acidification on these relationships was determined during this 40-day exposure experiment. Using carbon dioxide (CO2) gas, seawater was acidified to pHNBS 7.6, 7.2 or 6.8. Control treatments were maintained in natural seawater (pH ≈ 8.0). Under normocapnic conditions, burrowing urchins were seen to reduce the sediment uptake of nitrite or nitrate whilst enhancing the release of silicate and phosphate. In acidified (hypercapnic) treatments, the biological control of biogeochemical cycles by urchins was significantly affected, probably through the combined impacts of high CO2 on nitrifying bacteria, benthic algae and urchin behaviour. This study highlights the importance of considering biological interactions when predicting the consequences of seawater acidification on ecosystem function.

The aim of this study was to examine whether a combination of biochemical, histopathological and toxicogenomic data could be used as a valuable tool for the assessment of biological risk associated with pollutants within the Tamar River and Estuary, S.W. England, U.K. Accordingly, biochemical and histopathological biomarkers (protein carbonyls, lipofuscin, neutral lipids, lysosomal stability [N-acetyl-β-hexosaminidase and neutral red], lysosomal volume, ferric reducing antioxidant power [FRAP] and malonaldehyde [MDA]) and gene expression profiles were assessed in 5 sites from the Tamar River and Estuary (Neal Point, Town Quay, Wilcove, Cremyll Ferry and Whitsand; and a reference site, Trebarwith Strand, N. Cornwall). PAHs were measured in mussel tissue and sediment and metals were measured in mussel tissue only. Data from the biomarkers was integrated into a Mussel Expert System (MES) model to produce a simple assessment of mussel stress. Clear gradients of mussel toxicity were identified by the biomarkers (with the exception of neutral lipids) with the highest impacted animals found furthest up the Tamar, whilst the MES was unable to identify a gradient of effect. Gene expression profiles also indicated a gradient of stress with the greatest number of significantly up- or down- regulated genes found at the uppermost 2 sites. The MES did, however, determine that mussels from all sites, except the reference site, were highly stressed; a conclusion that could not be inferred from the biomarker data alone. It is concluded that the MES is a valuable tool that permits integration and interpretation of complex sets of biomarker data by identifying the biological meaning of biomarker changes.

In case of an avian-influenza-derived human flu pandemic, an inordinately high use of medicines over several weeks is predicted, in particular for the recommended influenza antiviral oseltamivir (Tamiflu). While the risk of oseltamivir to sewage works and freshwater bodies has already been assessed, the fact that a large percentage of the human population worldwide lives relatively close to the sea raises concern for its environmental compatibility in coastal marine waters. The potential risk of high oseltamivir use to the marine compartment is assessed in this publication, based on the 2003 European Community Technical Guidance Document (TGD) for risk assessment. Subchronic embryo–larval ecotoxicity tests with three marine invertebrates (Pomatoceros triqueter, Annelida; Mytilus edulis, Mollusca; Paracentrotus lividus, Echinodermata) and chronic growth inhibition tests with two different groups of marine microalgae (Isochrysis galbana, Haptophyta; Skeletonema costatum, Heterokontophyta) were performed with the active substance oseltamivir carboxylic acid to derive a dependable marine predicted no-effect concentration (PNEC). This was compared to a predicted environmental concentration (PEC) for oseltamivir in coastal waters, based on the worst-case freshwater PEC. The PEC/PNEC risk characterisation ratio for the marine compartment is well below 1, which in the terminology of the TGD signifies no immediate concern. Further, while oseltamivir may be persistent (P), it is not bioaccumulative (B) nor highly ecotoxic (T) and therefore not a PBT substance. In conclusion, even a high pandemic use of oseltamivir would not lead to a significant risk for the marine compartment, in confirmation of the risk assessment for sewage works and freshwaters.

A mesocosm experiment was conducted to quantify the effects of short- (2 wk) and long-term (20 wk) exposure to acidified seawater on the structure and diversity of macrofaunal and nematode assemblages in 2 different sediment. types. The impact of acidified seawater on sediment nutrient fluxes was also determined. Using carbon dioxide (CO2) gas, seawater was acidified to pH 7.3 (mimicking ocean acidification), 6.5 or 5.6 (mimicking leakage from a sub-seabed CO2 store site). Control treatments were maintained in natural seawater [pH approximate to 8.0). Exposure to acidified seawater significantly altered community structure and reduced diversity for both macrofaunal and nematode assemblages, However, the impact on nematodes was less severe than that on macrofauna. While the communities in both sediment types were significantly affected by changes in seawater pH, impacts on sandy sediment fauna were greater than those on muddy sediment fauna. Sandy sediments also showed the greatest effects with respect to nutrient fluxes. In sand, the efflux of nitrite, nitrate and silicate decreased in response to increased acidification while the efflux of ammonium increased, In mud, acidification increased the efflux of ammonium but had no effect on the other nutrients. We conclude that both leakage from carbon storage and ocean acidification could cause significant changes in the structure and diversity of coastal sediment communities. Lowered seawater pH could also affect nutrient cycling directly by altering bacterial communities and indirectly through impacts on the abundance and activity of key bioturbators.

The toxicity and ecotoxicity of nanomaterials is poorly understood and the risks of environmental exposure are largely unknown. In vitro cellular toxicity of C60-fullerene and carbon nanotubes was investigated by measuring the retention of a cationic probe (neutral red) within the lysosomal compartment of phagocytic blood cells (hemocytes) from marine mussels (Mytilus galloprovincialis). Intra-lysosomal retention of neutral red, over time, is a measure of the stability of the lysosomal membrane and the health of the cell. Aqueous suspensions of carbon nanoparticles (C60-fullerene – 1.0 and 10 µg.ml−1) induced cytotoxicity in circulating phagocytic hemocytes, which are a key component of the molluscan innate immune system. Hemocytes exposed to the same concentration range of carbon nanotubes were unaffected. These findings are consistent with the hypothesis that C60-fullerene is cytotoxic on uptake into the endocytic-lysosomal system and that cell injury may be mediated by generation of reactive oxygen species (ROS).

The impact of CO2-acidified seawater (pH 7.8, 7.6, or 6.5, control = pH 8) on the health of Mytilus edulis was investigated during a 60 d mesocosm experiment. Mussel health was deter- mined using the neutral red retention (NRR) assay for lysosomal membrane stability and from histopathological analysis of reproductive, digestive and respiratory tissues. Seawater acidification was shown to significantly reduce mussel health as measured by the NRR assay, and it is suggested that this impact is due to elevated levels of calcium ions (Ca 2+ ) in the haemolymph, generated by the dissolution of the mussels' calcium carbonate shells. No impact on tissue structures was observed, and it is concluded that M. edulis possess strong physiological mechanisms by which they are able to protect body tissues against short-term exposure to highly acidified seawater. However, these mech- anisms come at an energetic cost, which can result in reduced growth during long-term exposures. Consequently, the predicted long-term changes to seawater chemistry associated with ocean acidifi- cation are likely to have a more significant effect on the health and survival of M. edulis populations than the short-lived effects envisaged from CO2 leakage from sub-seabed storage. Ocean acidifica- tion could reduce the general health status of this commercially and ecologically important marine species.

Larval stages of two economically important flatfish, the sole (Solea solea) and turbot (Scophthalmus maximus) were exposed to ambient and elevated levels of UV-B. Sole larvae, which naturally occur in the plankton in early spring, demonstrated skin lesions at elevated levels of UV-B. Histopathology of the sole revealed cellular changes in the integument, characteristic of sunburn damage, with a reduction in the size of mucus-secreting cells and an increased epidermal thickening, especially at the highest doses of UV-B (2.15 KJ bio eff/m2). Pigmentation in the sole is restricted to a few isolated melanocytes. The integrity of the heavily pigmented skin of turbot appeared to be unaffected by comparable doses of UV-B. Both species have protective mechanisms, which minimize the effects of naturally-occurring levels of UV-B. However, sole appear to be poorly adapted to accommodate any further increase in solar radiation.

Indigenous and deployed mussels, Mytilus edulis, were examined at sites on the Tees estuary, an industrialized region of North-east England. Four sites were sampled for water and biota, two of which had been recently exposed to an acute effluent discharge. Mussels were deployed for 24 h before return to the laboratory. Tests for lysosomal stability were conducted on the haemolymph. Heart rate activity was measured. Bioassays were conducted on larvae from indigenous and deployed mussels. Clean site (Whitsand Bay, South-west England) larvae were exposed to the sampled water. Tissue was preserved for histopathology. Lysosomal stability, heart rate activity, larval bioassays and histopathology showed significant differences between sites. Correlations between these techniques were observed. There is a need for biomarkers that can rapidly provide sensitive environmental data on marine organisms in a cost-effective manner. These results are encouraging for the continued application of these biomarkers as initial indicators of environmental quality.

Indigenous and deployed mussels, Mytilus edulis, were examined at four sites on the Tees Estuary, in northeast England. Two of the sites had recently been exposed to an acute effluent discharge. Mussels (obtained from Whitsand Bay, a reference site in southwest England) were deployed at the sites for 24 h; water samples and indigenous mussels were also collected. Tests for haemocyte lysosomal stability and heart rate activity were conducted on adult mussels. Adult mussels were sacrificed and preserved for histopathology. Indigenous and deployed mussels were induced to spawn and the resultant larvae examined for abnormalities. Larvae (from Whitsand Bay adult mussels) were also exposed to the sampled water. The lysosomal stability, heart rate activity, histopathology, and larval bioassays showed significant differences between sites. A relationship between these techniques was observed. These results are encouraging for the continued application of these biomarkers as rapid indicators of environmental quality.