Metal exposure and biological responses in resident and transplanted blue mussels (Mytilus edulis) from the Scheldt estuary

The Western Scheldt river and estuary is known to be highly polluted as it receives industrial, agricultural and domestic effluents from one of the most populated and industrialised zones in Europe. Aquatic organisms are exposed to pollutants, specifically metals that are present in different environmental phases, e.g. dissolved, suspended material or sediment phases. The objective of this study was to study the relationship that exists between environmental metal levels, the degree of metal uptake by aquatic organisms with the concomitant biological responses. For this purpose the bivalve mollusk, Mytilus edulis, was selected as bioaccumulation indicator organism. Environmental (water and sediment) and mussel samples were collected during the late winter (March 2000) from sampling sites in the Scheldt estuary. Sites were selected to represent a salinity gradient from the mouth of the estuary to the furthest distribution area of mussels upstream in the system. Together with standard water quality parameters (e.g. salinity, dissolved oxygen, dissolved organic carbon, etc.) concentrations of twelve metals were analysed in the water (dissolved and suspended matter) and sediments. Levels of these metals were also measured in the soft tissue of M. edulis, together with concomitant biomarker responses in resident mussel populations at each site. The biomarkers that were included in this study were condition index, scope for growth, survival in air, cell membrane stability, DNA damage, and metallothioneins. Data were subjected to multivariate statistical analysis. The physico-chemical parameters and metals in the environmental samples clustered the sites to reflect the distribution based on the salinity gradient. Bioaccumulation results revealed increased metal uptake along a pollution gradient with the highest metal bioaccumulation occurring at the upstream most sites and therefore closest to the in the industrial activities. However, the biomarker responses clustered the sites in a manner that reflected the influence of combination of internal exposure (bioaccumulation) and external exposure (physico-chemical conditions). These differences in biomarker responses clearly demonstrated were attributed to abiotic factors other than metal pollution alone e.g. localized short-term increases in increased suspended sediment concentrations and decreased dissolved oxygen concentrations.     

Metal exposure and biological responses in resident and transplanted blue mussels (Mytilus edulis) from the Scheldt estuary

The Western Scheldt river and estuary is known to be highly polluted as it receives industrial, agricultural and domestic effluents from one of the most populated and industrialised zones in Europe. Aquatic organisms are exposed to pollutants, specifically metals that are present in different environmental phases, e.g. dissolved, suspended material or sediment phases. The objective of this study was to study the relationship that exists between environmental metal levels, the degree of metal uptake by aquatic organisms with the concomitant biological responses. For this purpose the bivalve mollusk, Mytilus edulis, was selected as bioaccumulation indicator organism. Environmental (water and sediment) and mussel samples were collected during the late winter (March 2000) from sampling sites in the Scheldt estuary. Sites were selected to represent a salinity gradient from the mouth of the estuary to the furthest distribution area of mussels upstream in the system. Together with standard water quality parameters (e.g. salinity, dissolved oxygen, dissolved organic carbon, etc.) concentrations of twelve metals were analysed in the water (dissolved and suspended matter) and sediments. Levels of these metals were also measured in the soft tissue of M. edulis, together with concomitant biomarker responses in resident mussel populations at each site. The biomarkers that were included in this study were condition index, scope for growth, survival in air, cell membrane stability, DNA damage, and metallothioneins. Data were subjected to multivariate statistical analysis. The physico-chemical parameters and metals in the environmental samples clustered the sites to reflect the distribution based on the salinity gradient. Bioaccumulation results revealed increased metal uptake along a pollution gradient with the highest metal bioaccumulation occurring at the upstream most sites and therefore closest to the in the industrial activities. However, the biomarker responses clustered the sites in a manner that reflected the influence of combination of internal exposure (bioaccumulation) and external exposure (physico-chemical conditions). These differences in biomarker responses clearly demonstrated were attributed to abiotic factors other than metal pollution alone e.g. localized short-term increases in increased suspended sediment concentrations and decreased dissolved oxygen concentrations.     

Cellular energy allocation in mussels (Mytilus galloprovincialis) from the stratified estuary as a physiological biomarker

This study demonstrates the usefulness of cellular energy allocation (CEA) evaluations as a physiological biomarker to infer the occurrence of natural stress in native populations of mussels inhabiting the stratified estuary (Krka River estuary, Croatia). Sampling sites were selected based on their differences in the salinity and temperature. The CEA value was calculated as a ratio between available energy (E_a) and energy consumption (E_c). Generally, higher values of E_a were recorded in June than in November, which were especially evident in the storage components (carbohydrates and lipids), while the constitutive component (proteins) remained relatively constant. The highest E_c was recorded in mussels at estuarine site compared to coastal site, which may be caused by the energetically costly maintenance of osmotic balance. Decrease in CEA recorded in estuarine mussels may ultimately result in a lower amount of energy available for growth, reproduction, or defence against other stresses (e.g. pollution).     

Effects of chronic exposure to dispersed oil on selected reproductive processes in adult blue mussels (Mytilus edulis) and the consequences for the early life stages of their larvae

Mussels (Mytilus edulis) were continuously exposed to dispersed crude oil (0.015-0.25 mg/l) for 7 months covering the whole gamete development cycle. After 1 month exposure to 0.25 mg oil/l, the level of alkali-labile phosphates (ALP) and the volume density of atretic oocytes in female gonads were higher than those in the gonads of control females, indicating that oil affected the level of vitellogenin-like proteins and gamete development. Spawning of mussels was induced after 7 months oil exposure. Parental oil exposure did not affect subsequent fertilization success in clean seawater but this was reduced in 0.25 mg oil/l. Parental exposure to 0.25 mg oil/l caused both slow development and a higher percentage of abnormalities in D-shell larvae 2 days post-fertilization; reduced growth 7 days post-fertilization. These effects were greatly enhanced when larval stages were maintained at 0.25 mg oil/l. Similar studies are warranted for risk assessment prognosis.     

Biomarker responses as indication of contaminant effects in blue mussel (Mytilus edulis) and female eelpout (Zoarces viviparus) from the southwestern Baltic Sea

During a field study performed in spring and autumn 2001 and 2002, blue mussels (Mytilus edulis) and female eelpout (Zoarces viviparus) were collected at three locations in the Wismar Bay (Baltic Sea), and several biomarkers of contaminant effects were analysed. Besides seasonal and inter-annual variations, biomarker signals were most pronounced at the location closest to Wismar Harbour (Wendorf) in both species. Lysosomal membrane stability (LMS) was lowest and acetylcholinesterase activity (AChE) was significantly reduced. Frequency of micronuclei (MN) was significantly higher (in blue mussels), indicating mutagenic effects. In eelpout elevated levels of DNA adducts, EROD induction and PAH-metabolites were measured. Metallothionein (MT), biomarker for trace metal exposure, showed a gradient only in spring. Organochlorine contaminant analyses (PCBs, DDTs) corresponded to the observed biomarker levels. The results obtained clearly demonstrate pollution effects in the southwestern Baltic Sea. Moreover, they show that a multibiomarker approach is also applicable in a brackish water environment.     

PAH body burden and biomarker responses in mussels (Mytilus edulis) exposed to produced water from a North Sea oil field: laboratory and field assessments

In order to study the impact of produced water (PW) from a North Sea oil field on blue mussels (Mytilus edulis), chemical and biological markers were selected. A laboratory exposure (0.125%, 0.25% and 0.5% of PW) and a field study (6 stations 0.2-2 km from a PW discharge point) were conducted. In the laboratory study, PAH bioaccumulation increased in mussel soft tissue even at the lowest exposure dose. Micronuclei frequency demonstrated a dose-response pattern, whereas lysosomal membrane stability showed tendency towards a dose-response pattern. The same markers were assessed in the field study, biomarker analyses were consistent with the contamination level, as evaluated by mussel polycyclic aromatic hydrocarbons body burden. Overall, obtained results confirmed the value of an ecotoxicological approach for a scientifically sound characterisation of biological effects induced by offshore oilfield operational discharges.     

Dose-dependent response of a benthic system to biodeposition from suspended blue mussel (Mytilus edulis) culture

This study reports the results of a field experiment using benthic mesocosms that examined dose-dependent effects of mussel biodeposition on the benthic environment. Mesocosms were placed in the natural sea bottom and subjected to one of eight levels of biodeposition (from 0 to 1400 mussels m^?2). Most analyses indicated non-linear (i.e., threshold) effects. Sediment characteristics changed significantly between 200 and 400 mussels m^?2 as did multivariate community structure. Community structure effects were characterised by changes in abundances of species that are very sensitive or tolerant to organic loading. The multivariate AZTI Marine Biotic Index (M-AMBI) indicated that the benthic status changed from High to Good in all mesocosms receiving biodeposits. Sediments acted as a sink for oxygen (O₂), but results suggest O₂ sediment demand was not sensitive enough to evaluate organic loading impacts. Results from this and improved experiments can be used to determine the environmental carrying capacity of sites for bivalve culture.     

Disseminated neoplasia in blue mussels, Mytilus galloprovincialis, from the Black Sea, Romania

Disseminated neoplasia, also called leukemia or hemic neoplasia, has been detected in 15 species of marine bivalve mollusks worldwide. The disease is characterized by the presence of single anaplastic cells with enlarged nuclei and sometimes frequent mitosis, in hemolymph vessels and sinuses. The neoplastic cells gradually replace normal hemocytes leading to the increased mortality of animals. The neoplasia reaches epizootic prevalences in blue mussels, Mytilus trossulus, in some areas, whereas prevalences in Mytilus edulis are generally very low. Mytilus galloprovincialis was suggested to be resistant to the disease although very low prevalences were documented from Spain in the Atlantic Ocean and Italy in the Mediterranean Sea. A case of disseminated neoplasia was discovered in M. galloprovincialis from among 200 specimens studied from the coast of the Romanian Black Sea. Histological preparation revealed the presence of large anaplastic cells with lobed nuclei. This observation extends the geographic range of marine bivalve mollusks with disseminated neoplasia to include the Black Sea.     

Spatial and temporal distribution of heavy metal concentrations in mussels (Mytilus edulis) from the Baie des Chaleurs, New Brunswick, Canada

Previous studies on heavy metal contamination of the Baie des Chaleurs focus only on industrial centers and overlooked the ecosystem as a whole. To fill this gap, the objective of this study is to establish a baseline of the spatio-temporal distribution of heavy metals in mussels from the Baie des Chaleurs based on the ecosystem approach. Our results show, for the first time, a cadmium contamination in mussels across the south coast of the Baie des Chaleurs and not only in industrial centers. Our results also confirm previous studies showing heavy metal contamination of the Belledune area. This study demonstrates that the use of the ecosystemic approach is essential to obtain a comprehensive picture of environmental contamination in marine ecosystems.     

Comparative petroleum hydrocarbons levels and biochemical responses in mussels from hydrothermal vents (Bathymodiolus azoricus) and coastal environments (Mytilus galloprovincialis)

Aliphatic hydrocarbons and PAHs in the whole soft tissues of Bathymodiolus azoricus from three Mid-Atlantic Ridge hydrothermal vents (Menez-Gwen, Lucky Strike and Rainbow), and Mytilus galloprovincialis from three contaminated coastal sites in South Portugal were analysed, and its effects on the digestive gland microsomes mixed-function oxygenase system (MFO) were assessed. Aliphatic hydrocarbons levels were present in the same magnitude in both coastal and hydrothermal environments, while the UCM (unresolved complex mixture) for coastal mussels were higher than in vent mussels. In general, significantly higher PAHs concentrations were found in coastal mussels, compared to B. azoricus where low molecular weight PAHs (2-3 rings) represented the majority of PAHs contrarily to what was observed in M. galloprovincialis. The MFO components were present in both mussel species, and were detected in vent mussels for the first time. However this system seems to have different roles in species from these contrasting environments. In coastal mussels MFO responded to hydrocarbon contamination while response in hydrothermal organisms appeared to be related mainly to endogenous factors.     

Comparative petroleum hydrocarbons levels and biochemical responses in mussels from hydrothermal vents (Bathymodiolus azoricus) and coastal environments (Mytilus galloprovincialis)

Aliphatic hydrocarbons and PAHs in the whole soft tissues of Bathymodiolus azoricus from three Mid-Atlantic Ridge hydrothermal vents (Menez-Gwen, Lucky Strike and Rainbow), and Mytilus galloprovincialis from three contaminated coastal sites in South Portugal were analysed, and its effects on the digestive gland microsomes mixed-function oxygenase system (MFO) were assessed.Aliphatic hydrocarbons levels were present in the same magnitude in both coastal and hydrothermal environments, while the UCM (unresolved complex mixture) for coastal mussels were higher than in vent mussels. In general, significantly higher PAHs concentrations were found in coastal mussels, compared to B. azoricus where low molecular weight PAHs (2–3 rings) represented the majority of PAHs contrarily to what was observed in M. galloprovincialis. The MFO components were present in both mussel species, and were detected in vent mussels for the first time. However this system seems to have different roles in species from these contrasting environments. In coastal mussels MFO responded to hydrocarbon contamination while response in hydrothermal organisms appeared to be related mainly to endogenous factors.     

Heavy metal concentrations and biomarkers of oxidative stress in native mussels (Mytilus edulis chilensis) from Beagle Channel coast (Tierra del Fuego, Argentina)

The aim of this study was to evaluate the usefulness of oxidative stress biomarkers of pollution in native mussels Mytilus edulis chilensis from the Beagle Channel. Spatial and seasonal variations of catalase, glutathione-S-transferase and lipid peroxidation in gills and digestive gland were analyzed in relation to environmental parameters, heavy metals in sediment and in tissue. Four sites with anthropogenic impact and a control site were selected and monitored during the four seasons of 2007. We found significant differences among sites in concentrations of dissolved nutrients and heavy metals in sediments, with the highest values recorded at sites with anthropogenic pressure. Different patterns were observed between concentrations of metals in tissues and in sediments suggesting differences in bioavailability. There were also significant differences in biomarker responses among sites, despite the strong seasonal variability. Our results showed relatively moderate levels of pollution in the study area as a result of urban influences.     

Molecular and biochemical biomarkers responses in the mussel Mytilus edulis collected from Southern Brazil coast

Marine ecosystems are typically subjected to a variety of stressors containing complex xenobiotics mixtures. This study aims to evaluate the responses of molecular and biochemical biomarkers in the mussel Mytilus edulis providing data for environmental monitoring programs development in Southern Brazil. Mussels were collected at a polluted site, near Patos Lagoon outfall, and at a control site. Gills, muscle and mantle samples were used for biomarker determinations. Mussels collected at the polluted site significantly increased superoxide dismutase (SOD) and glutathione S-transferase (GST) activities and decreased catalase (CAT) activity. Moreover, an increase in sod1, gstπ and hsp70 mRNA expression was observed. Overall, biochemical and molecular biomarkers responses were observed, but these responses varied depending on the analyzed tissue. These results indicate possible contaminants effects on organisms and the need for effective environmental monitoring programs in this ecosystem.     

Uptake and loss of zinc and lead by mussels (Mytilus edulis) and relationships with body weight and reproductive cycle

The uptake and loss of zinc and lead from the whole soft parts of the mussel Mytilus edulis were investigated under natural conditions in north-east England. Mussels in which high concentrations of these metals were found were transferred to an area where low concentrations were found, and vice versa. Mussels from both areas were also kept in tanks and periodic measurements of metal contents were made. Indications of uptake and loss were greatly affected by changing body weights and the work suggests that phase of reproductive cycle (for animals in the environment) and condition (for animals in tanks) have to be closely considered when reporting on levels of metals in mussels, especially in terms of concentration.     

Total oxyradical scavenging capacity responses in Mytilus galloprovincialis transplanted into the Venice lagoon (Italy) to measure the biological impact of anthropogenic activities

Oxidative stress related investigations to monitor the impact of the pollutant discharges into the Venice lagoon (Italy) originating from anthropogenic activities (raw sewage water, agricultural and industrial effluents, oil tanker traffic), on marine organisms have classically been carried out by analyzing specific, single antioxidants (i.e. catalase, superoxide dismutase). In this paper, two studies are reported where the total oxyradical scavenging capacity assay (TOSC) was selected and measured toward peroxyl, and hydroxyl free radicals, and peroxynitrite in mussels (Mytilus galloprovincialis) transplanted into the Venice city and throughout the lagoon to measure the biological effects of anthropogenic activities. In the first experiment, mussels from a clean site (farm) were transplanted to the urban area of Venice for 0, 1, 2 and 4 weeks; cytosolic TOSC toward peroxyl and hydroxyl free radicals, and peroxynitrite revealed that the transplantation process caused a stress (handling stress, anoxia, oxidative burst) resulting in a reduction of TOSC in both control and urban sites, therefore, preventing clear interpretation of the data after one week. At week 2, a significant TOSC reduction (P < 0.05) toward peroxyl and hydroxyl radicals in the urban site revealed that mussels experienced oxidative pressure exerted by pollutants. Most TOSC values returned to initial levels at week 4; however, TOSC induction was noticed in the control group toward peroxyl and hydroxyl radicals while in the exposed group it was not indicating an inhibition of the oxidative metabolism. In the second experiment, mussels were deployed at seven different sites throughout the lagoon. After five weeks of exposure, significant TOSC reduction was measured (P < 0.05) toward peroxyl for Palude della Rosa, Chioggia and Valle Millecampi, toward hydroxyl radicals for Valle Millecampi and Campalto and toward peroxynitrite for Valle Millecampi. Although these data indicate a depletion of the low molecular weight scavengers, additional biomarkers are needed to draw a conclusion on the health of the mussels. TOSC was proved to be an interesting health index parameter to measure pollution impact in a transplantation study provided that the mussels are exposed for two weeks and a control is run in parallel.     

Oil change in excretory products of mussels (Mytilus galloprovincialis)

Oil ingested by mussels appears in the faeces and pseudofaeces but differs in composition from the oil to which the animals are exposed. It contains more heavy fractions and larger aromatics, and there is a shift of aliphatic hydrocarbons towards heavier alkanes and isoprenoids. There is evidence that these changes are the result of metabolic processes in the mussels.     

Effects of 4-nonylphenol exposure in mussels (Mytilus galloprovincialis) and crabs (Carcinus aestuarii) with particular emphasis on vitellogenin induction

Since it is often difficult to estimate possible adverse effects due to contamination in selected ecosystems, multi-species biomonitoring may provide more information, taking into account different routes of exposure, ecological roles and metabolic capabilities of animals. In this context, we exposed for 7 days the mussel Mytilus galloprovincialis and the crab Carcinus aestuarii to 4-nonylphenol (NP), a well-known xenoestrogen. In mussels (0–0.2 mg NP l⁻¹), we measured NP bioaccumulation in soft tissues and vitellogenin (Vg)-like protein levels in digestive glands from both males and females by the alkali-labile phosphate assay (ALP). As no reference data were available for crab exposure, the NP 96-h LC₅₀ value was previously determined. Then, in sublethally exposed (0–1.0 mg NP l⁻¹) male crabs, NP bioaccumulation and Vg levels were measured in hemolymph, gonads and digestive gland. Bioaccumulation of NP increased from 43 to 371 μg g⁻¹ d.w. in mussels, and from 3.6 to 37 μg g⁻¹ d.w. in crabs, depending on the NP concentration in water. Dose-dependent Vg-like protein induction was observed in both species, appearing to be related to NP bioaccumulation, although a partial decrease was recorded at the highest concentration tested. A similar trend was observed in both digestive gland and gonad of exposed crabs; Vg increased to a lesser extent, although significantly, in hemolymph. Results demonstrated that NP induces Vg synthesis both in male and female mussels, as well as in male crabs. On the basis of the responsiveness of both species investigated, a multi-species approach is indicated in biomonitoring programmes.     

Effects of 4-nonylphenol exposure in mussels (Mytilus galloprovincialis) and crabs (Carcinus aestuarii) with particular emphasis on vitellogenin induction

Since it is often difficult to estimate possible adverse effects due to contamination in selected ecosystems, multi-species biomonitoring may provide more information, taking into account different routes of exposure, ecological roles and metabolic capabilities of animals. In this context, we exposed for 7 days the mussel Mytilus galloprovincialis and the crab Carcinus aestuarii to 4-nonylphenol (NP), a well-known xenoestrogen. In mussels (0-0.2 mg NP l(-1)), we measured NP bioaccumulation in soft tissues and vitellogenin (Vg)-like protein levels in digestive glands from both males and females by the alkali-labile phosphate assay (ALP). As no reference data were available for crab exposure, the NP 96-h LC(50) value was previously determined. Then, in sublethally exposed (0-1.0 mg NP l(-1)) male crabs, NP bioaccumulation and Vg levels were measured in hemolymph, gonads and digestive gland. Bioaccumulation of NP increased from 43 to 371 microg g(-1) d.w. in mussels, and from 3.6 to 37 microg g(-1) d.w. in crabs, depending on the NP concentration in water. Dose-dependent Vg-like protein induction was observed in both species, appearing to be related to NP bioaccumulation, although a partial decrease was recorded at the highest concentration tested. A similar trend was observed in both digestive gland and gonad of exposed crabs; Vg increased to a lesser extent, although significantly, in hemolymph. Results demonstrated that NP induces Vg synthesis both in male and female mussels, as well as in male crabs. On the basis of the responsiveness of both species investigated, a multi-species approach is indicated in biomonitoring programmes.     

Susceptibility to oxidative stress of mussels (Mytilus galloprovincialis) in the Venice Lagoon (Italy)

The aim of this study was to evaluate the susceptibility to pollutant mediated oxidative stress of the Mediterranean mussel Mytilus galloprovincialis in the Venice lagoon (Italy).

In June 2003, mussels from a farm were transplanted to eight sites in the lagoon for five weeks. Oxidative stress responses were measured by: (i) total oxyradical scavenging capacity (TOSC) assay, for an overall evaluation of the oxidative stress response capability; (ii) catalase (CAT), as a key enzyme involved in the antioxidant defence system; (iii) malondialdehyde (MDA), as an indicator of lipid peroxidation, to evaluate an oxidative damage; (iv) metallothioneins (MTs), as they play a role in the antioxidant defence.

The TOSC analysis revealed a reduced capability to eliminate: (i) peroxyl radical in mussels transplanted at Palude della Rosa, Valle Millecampi and Chioggia; (ii) hydroxyl radical at Campalto and Valle Millecampi; (iii) peroxynitrite at Valle Millecampi.

Inhibition in CAT activity, observed in all the monitored sites, confirms the presence of an oxidative pressure in transplanted mussels.

In addition, Pearson correlation analysis was performed in order to observe possible links between the various parameters. The PCA was a powerful tool to discriminate impacted sites, suggesting that the mussels transplanted throughout the Venice lagoon were subjected to different levels of oxidative pressure. Furthermore, it provided an easy and useful tool to summarize the obtained results.