Effects of contaminant exposure and food restriction on hepatic autophagic lysosomal parameters in Herring Gull (Larus argentatus) chicks.

Lysosomal autophagic responses, such as lysosomal membrane stability, neutral lipids (NL), lipofuscin (LF), and malondialdehyde (MDA) levels, are valuable measures of cellular early-onset effects induced by environmental stress factors, such as contaminant exposure and fasting. In this study, these parameters were analysed and related to levels of halogenated organic contaminants (HOCs) in 40 Herring Gull (Larus argentatus) chicks. Chicks were experimentally exposed to HOCs through diet and went through a period of nutrient deprivation at the end of the experiment. HOC exposure and fasting were conducted separately and in combination. NL storages were depleted, and lysosomal membranes were destabilised after HOC exposure and nutrient deprivation. These responses were not related specifically to one type of stress or the extent of the treatment. No synergistic or additive effects from the combination of HOC exposure and fasting were observed. LF accumulated, and MDA levels increased as a result of fasting, but were unaffected by HOC exposure. LF accumulation was strongly associated with the percent weight change in the chicks. Large weight loss was associated with high LF levels, and slight weight gain was associated with low LF levels. Hence, food deprivation affected all the measured parameters, and HOC exposure decreased NL levels and lysosomal membrane stability in HG chick liver. Furthermore, autophagic lysosomal parameters have frequently been applied as biomarkers of cellular health status in previous studies of marine and terrestrial invertebrates, and this study suggests that these parameters may be good candidates for biomarkers of cellular health status in seabirds as well.

Lysosomal and lipid-associated parameters in the livers of three species of arctic seabird chicks: species differences and relationships with contaminant levels.

Lysosomal membrane stability, lipofuscin (LF), malondialdehyde (MDA), neutral lipid (NL) levels, as well as halogenated organic compounds (HOCs), Cr, Cd, Pb and Fe concentrations were analyzed in liver of black-legged kittiwake (BK), herring gull (HG), and northern fulmar (NF) chicks. There were significant species differences in the levels of NL, LF and lysosomal membrane stability. These parameters were not associated with the respective HOC concentrations. LF accumulation was associated with increasing Cr, Cd and Pb concentrations. HG presented the lowest lysosomal membrane stability and the highest. LF and NL levels, which indicated impaired lysosomes in HG compared to NF and BK. Lipid peroxidation was associated with HOC and Fe2+ levels. Specific HOCs showed positive and significant correlations with MDA levels in HG. The study indicates that contaminant exposure can affect lysosomal and lipid associated parameters in seabird chicks even at low exposure levels. These parameters may be suitable markers of contaminant induced stress in arctic seabirds.

Hepatic antioxidant responses related to levels of PCBs and metals in chicks of three Arctic seabird species.

The efficiency of antioxidant defenses and relationship with body burden of metal and organic contaminants has not been previously investigated in arctic seabirds, neither in chicks nor in adults. The objective of this study was to compare such defenses in chicks from three species, Black-legged kittiwake (Rissa tridactyla), Northern fulmar (Fulmarus glacialis), and Herring gull (Larus argentatus), and the relationship with tissue concentrations of essential metals such as selenium and iron and halogenated organic compounds, represented by polychlorinated biphenyl (PCB). The results showed significant species-specific differences in the antioxidant responses which also corresponded with metal and PCB levels in different ways. The capability to neutralize hydroxyl radicals (TOSC-HO•) and the activities of catalase and Se-dependent glutathione peroxidases (GPX) clearly increased in species with the higher levels of metals and PCBs, while the opposite trend was observed for Se-independent GPX, TOSC against peroxyl radicals (ROO•) and peroxynitrite (ONOOH). Less clear relationships were obtained for glutathione levels, GSH/GSSG ratio, glutathione reductase and superoxide dismutase. The results showed differences in antioxidant efficiency between the species, and some of these defenses exhibited dose-response-like relationships with measured levels of selenium, iron and ΣPCBs. PCBs, selenium and iron levels were positively related to the responses of antioxidants with potential to reduce HO•/H2O2 (Se-dependent GPX, CAT and TOSC against HO•). However, direct causal relationships between antioxidant responses and contaminant concentrations could not be shown on individual level. Varying levels of metals and contaminants due to different diet and age were probably the main explanations for the species differences in antioxidant defense.

Contaminant accumulation and biomarker responses in caged mussels, Mytilus galloprovincialis, to evaluate bioavailability and toxicological effects of remobilized chemicals during dredging and disposal operations in harbour areas.

Remobilization of chemicals from contaminated sediments is a major risk associated with dredging and disposal operations in harbour areas. In this work caged mussels, Mytilus galloprovincialis, were chosen as bioindicator organisms to reveal the impact and recovery of organisms from these activities in the harbour of Piombino (Tuscany, Italy) where approximately 100,000 m(3) of sediments were removed and disposed in a local confined disposal facility (CDF). Organisms were deployed before, during and after the end of operations, selecting sites differently impacted by these activities. Temporal changes in environmental bioavailability and biological effects of pollutants were assessed by integrating analyses of trace metals and polycyclic aromatic hydrocarbons (PAHs) accumulated in tissues of caged mussels with a wide array of biomarkers reflecting exposure to specific classes of pollutants and different levels of cellular unbalance or toxicity. Such biological responses included levels of metallothioneins, activity of acyl CoA oxidase (AOX) as a marker of peroxisome proliferation, oxidative stress biomarkers (content of glutathione, enzymatic activities of catalase, glutathione S-transferases, glutathione reductase, glutathione peroxidases), total oxyradical scavenging capacity (TOSC) toward peroxyl and hydroxyl radicals, lysosomal membrane stability and genotoxic effects measured as DNA strand breaks and frequency of micronuclei. Obtained results indicated that a general disturbance was already present in the whole harbour area and especially in the inner site before the beginning of operations, when caged mussels exhibited a significant accumulation of PAHs and Pb, lower TOSC values and higher levels of both lysosomal and genotoxic damages. Bioavailability of trace metals and PAHs markedly increased during dredging activities with values up to 40 microg/g for Pb and up to 2200 ng/g for PAHs in tissues of caged mussels, a significant inhibition of antioxidant efficiency and increase of oxidative damages. While bioavailability of trace metals returned to the pre-dredging values after the end of operations, the accumulation of PAHs, oxidative effects and genotoxic damages remained elevated in mussels caged in the inner area and in front of CDF. Overall this study confirmed the utility of caged mussels to assess the remobilization of chemicals from dredged sediments and the onset of potentially harmful biological effects.

Seasonal variations of exposure biomarkers, oxidative stress responses and cell damage in the clams, Tapes philippinarum, and mussels, Mytilus galloprovincialis, from Adriatic sea.

This work investigated the natural variability of several biomarkers in Tapes philippinarum and Mytilus galloprovincialis, sampled from Northern Adriatic where these organisms are important sentinel species for future environmental impact assessment. Levels of metallothioneins, peroxisomal enzymes and acetylcholinesterase, showed a significant seasonality and marked differences between clams and mussels. Among antioxidant enzymes, catalase and GST decreased during the warmer period, the latter enzyme activity resulting particularly high in clams. The total oxyradical scavenging capacity toward peroxyl radicals decreased in mussels from winter to summer, indicating a prooxidant challenge due to higher seawater temperature and intensity of light irradiance. Lysosomal membrane stability did not exhibit significant seasonal variations, while some variations were observed for DNA damages. Overall results indicated a significant influence of seasonal variability on several biomarkers and species-specific differences which should be considered to discriminate the appearance of anthropogenic disturbance.

Seasonal variability of oxidative biomarkers, lysosomal parameters, metallothioneins and peroxisomal enzymes in the Mediterranean mussel Mytilus galloprovincialis from Adriatic Sea.

The Mediterranean mussel, Mytilus galloprovincialis, is a classical sentinel organism for monitoring the biological effects of contaminants through the use of molecular and cellular biomarkers. These biological responses can be modulated also by seasonal changes of both environmental and biological factors, potentially influencing responsiveness and sensitivity to pollutants. The aim of this study was to characterize in a reference mussel population from the Adriatic Sea, the natural fluctuations of several oxidative stress biomarkers widely used in ecotoxicological applications. Analyses of individual antioxidant defenses (catalase, glutathione S-transferases, glutathione reductase, glutathione peroxidases, levels of glutathione) were integrated with those of the total oxyradical scavenging capacity (TOSC-assay), which quantify the overall capability of a tissue to neutralize different forms of oxyradicals. Due to the close relationship between antioxidant efficiency and onset of various cellular alteration, the seasonal characterization was carried out also on lysosomal membrane stability, accumulation of malondialdehyde, neutral lipids and lipofuscin, levels of metallothioneins and activity of peroxisomal enzymes (acyl-CoA oxidase and d-aminoacid oxidase). Results indicated a significant seasonality for several biological responses; major variations occurred especially in spring and summer months suggesting the influence of natural factors, such as temperature, reproductive cycle and food availability. The observed seasonal oscillations revealed both similarities and differences with those reported for other Mediterranean mussel populations suggesting that opposite trends can occur when the same environmental prooxidant factors have a different regional influence.

Use of the land snail Helix aspersa as sentinel organism for monitoring ecotoxicologic effects of urban pollution: an integrated approach.

Atmospheric pollution from vehicular traffic is a matter of growing interest, often leading to temporary restrictions in urban areas. Although guidelines indicate limits for several parameters, the real toxicologic impacts remain largely unexplored in field conditions. In this study our aim was to validate an ecotoxicologic approach to evaluate both bioaccumulation and toxicologic effects caused by airborne pollutants. Specimens of the land snail Helix aspersa were caged in five sites in the urban area of Ancona, Italy. After 4 weeks, trace metals (cadmium, chromium, copper, iron, manganese, nickel, lead, and zinc) and polycyclic aromatic hydrocarbons (PAHs) were measured and these data integrated with the analyses of molecular and biochemical responses. Such biomarkers reflected the induction of detoxification pathways or the onset of cellular toxicity caused by pollutants. Biomarkers that correlated with contaminant accumulation included levels of metallothioneins, activity of biotransformation enzymes (ethoxyresorufin O-deethylase, ethoxycoumarin O-deethylase), and peroxisomal proliferation. More general responses were investigated as oxidative stress variations, including efficiency of antioxidant defenses (catalase, glutathione reductase, glutathione S-transferases, glutathione peroxidases, and total glutathione) and total oxyradical scavenging capacity toward peroxyl and hydroxyl radicals, onset of cellular damages (i.e., lysosomal destabilization), and loss of DNA integrity. Results revealed a marked accumulation of metals and PAHs in digestive tissues of organisms maintained in more traffic-congested sites. The contemporary appearance of several alterations confirmed the cellular reactivity of these chemicals with toxicologic effects of potential concern for human health. The overall results of this exploratory study suggest the utility of H. aspersa as a sentinel organism for biomonitoring the biologic impact of atmospheric pollution in urban areas. Key words: atmospheric pollutants, bioindicators, biomarkers, DNA integrity, lysosomes, metallothioneins, oxidative stress, peroxisomes, polycyclic aromatic hydrocarbons, trace metals.

Pro-oxidant effects of extremely low frequency electromagnetic fields in the land snail Helix aspersa.

Pro-oxidant effects of extremely low frequency (ELF) 50-Hz magnetic fields were investigated in the land snail Helix aspersa exposed both in short-term laboratory treatments and under field conditions by maintaining the organisms in the proximity of a power line for up to 2 months. Oxidative perturbations were investigated as individual antioxidants (catalase, glutathione reductase, glutathione S-transferases, and total glutathione) and total scavenging capacity toward peroxyl radicals and hydroxyl radicals. Accumulation of lipid peroxidation products, destabilization of lysosomal membranes, and loss of DNA integrity were also evaluated as markers of cell damage. The overall results indicated an oxidative challenge caused by ELF magnetic fields with particularly prompt and sensitive responses for catalase, glutathione reductase, and the overall capability to neutralize peroxyl radicals. Cell injuries occurred to different extents according to duration and intensity of electromagnetic exposure and confirmed complex cause-effect relationships between pro-oxidant factors, efficiency of antioxidant defenses, and the onset of oxidative toxicity. This study highlights the importance of a multimarker approach for detecting a wide panel of biological responses, the necessity of investigating the long-term effects of early oxidative responses, and the role of ELF in enhancing susceptibility to other forms of pathologies or diseases.

Total content and chemical speciation of arsenic in the polychaete Sabella spallanzanii.

An unusual feature has been observed in polychaetes regarding their capacity to accumulate high levels of relatively toxic forms of arsenic in specific tissues. Basal levels of arsenic and distribution of its compounds were investigated in tissues of the Mediterranean polychaete Sabella spallanzanii. Particularly high concentrations were measured in the branchial crown (1036+/-136 microg/g d.w.) and chemical speciation revealed that the predominant form was the relatively toxic dimethylarsinate (DMA). These data suggest a potential role of As as anti-predatory strategy in polychaetes.

Time-course evaluation of ROS-mediated toxicity in mussels, Mytilus galloprovincialis, during a field translocation experiment.

Harbours can be considered as model environments for developing and validating field monitoring procedures and to investigate mechanistic relationships between different biological responses. In this study several biomarkers were investigated in marine mussels caged for 4 weeks (June-July 2001) into an industrialized harbour of NW Italy. Organisms were collected at different time-intervals to better characterize the sensitivity, temporal variations and interactions of analysed responses. Besides single antioxidants the total oxyradical scavenging capacity (TOSC) assay was used to analyse the capability of the whole antioxidant system to neutralize specific forms of radicals: these data were further integrated by measurement of DNA integrity, oxidized bases and the impairment of lysosomal membrane stability in haemocytes. Results showed a biphasic trend for single antioxidants and TOSC, with an increase during the first 2 weeks of exposure to the polluted site followed by a progressive decrease up to a severe depletion in the final part of the experiment.

Time-course variations of oxyradical metabolism, DNA integrity and lysosomal stability in mussels, Mytilus galloprovincialis, during a field translocation experiment.

Harbours can be considered as model environments for developing and validating field monitoring procedures and to investigate mechanistic relationships between different biological responses. In this study, several biomarkers were investigated in marine mussels caged for 4 weeks into an industrialised harbour of north-west Italy. Organisms were collected at different time intervals to better characterise the sensitivity, temporal variations and interactions of analysed responses. Besides single antioxidants (catalase, glutathione S-transferases, glutathione reductase, total glutathione), the total oxyradical scavenging capacity (TOSC) assay was used to analyse the capability of the whole antioxidant system to neutralise specific forms of radicals: these data were further integrated by measurement of DNA integrity, oxidised bases and the impairment of lysosomal membrane stability in haemocytes. Results showed a biphasic trend for single antioxidants and TOSC, with no variation or increase during the first 2 weeks of exposure to the polluted site followed by a progressive decrease up to a severe depletion in the final part of the experiment. These findings suggest an initial counteractive response of mussels toward the enhanced prooxidant challenge, while antioxidants appeared overwhelmed at longer exposure periods. The hypothesis of reactive oxygen species (ROS) mediated toxicity is supported by the appearance of cell damages (DNA integrity and lysosome membrane stability), which exhibited a progressive enhancement during the course of the experiment with a maximum impairment after 30 days of exposure.