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.     

Spatial and temporal variation of biomarkers in mussels (Mytilus galloprovincialis) from the Lagoon of Venice, Italy

A multiple biomarker approach was adopted in a seasonal study carried out in the Lagoon of Venice, with the double aim of evaluating the natural and anthropic stresses influencing the biological responses of mussel (Mytilus galloprovincialis) and of assessing the effects due to spatial rather than temporal variations.Biochemical (aldehyde dehydrogenase and catalase activities), cellular (neutral red retention time) and physiological (survival in air and condition index) biomarkers were determined in mussels collected in four differently impacted lagoon areas.Multivariate analysis showed that samples were distributed mainly according to temperature and four seasonal groups were identified. The combination of spatial and temporal information enabled us to distinguish physiological variations due to natural causes from those due to anthropic stress, and to identify the sampling period when several biomarkers are less influenced by both natural and endogenous factors.     

Polycyclic aromatic hydrocarbons in surface seawater and in indigenous mussels (Mytilus galloprovincialis) from coastal areas of the Saronikos Gulf (Greece)

Polycyclic aromatic hydrocarbons (PAHs) were identified and measured in surface seawater and in the tissues (gills and mantle) of indigenous black mussels, Mytilus galloprovincialis, collected from three coastal sites of Saronikos Gulf (Greece), a gulf that exhibits high levels of pollution. The total PAHs measured by spectrofluorometry in the surface seawater were found in the range of 425–459 ng L⁻¹ at the most polluted sites 1 and 2 (Elefsis Bay–Salamis Island) and in the range of 103–124 ng L⁻¹ at site 3 (Aegina Island). PAHs' sources in seawater were identified by application of specific PAH ratios, such as phenanthrene/anthracene and fluoranthene/pyrene. Levels of PAHs in soft tissues (gills and mantle) of indigenous mussels were much higher than those reported for seawater. Total PAH concentrations in mantle tissues were in the range of 1300–1800 ng g⁻¹ dry weight (dw) tissue at sites 1 and 2 and approximately 380 ng g⁻¹ dw at site 3. In gill tissues total PAH concentrations were in the range of 1480–2400 ng g⁻¹ dw at sites 1 and 2 and approximately 430 ng g⁻¹ dw at site 3. PAHs composition was dominated by two-, three- and four-ring compounds in seawater, where 17 different PAH compounds were identified and measured in mussel tissues. Mussels can be used as sentinel organisms to monitoring PAHs' contamination, since they concentrate PAHs from the surrounding water media and therefore making the chemical analysis simpler and less prone to error than that for water. In surface seawater possible weathering and photodegradation due to hot climates contribute to reduced PAHs concentrations.     

Effects of 17β-estradiol exposure in the mussel Mytilus galloprovincialis

Mussels Mytilus galloprovincialis were exposed to different concentrations of estradiol (20, 200, and 2000 ng/l) in a semi-static regime (1-day dosing intervals) for up to 7 days in an attempt to see how mussels dealt with exogenous estrogenic compounds. Sex hormone levels were determined in whole tissue. Free-estradiol was only significantly elevated at the highest exposure dose (up to 10-fold). Most of the estradiol was in the tissues as fatty acid esters (>78%), which sharply increased in a dose-dependent manner (from 4 ng/g in controls to 258 ng/g at the high exposure group). In contrast, neither free nor esterified testosterone levels showed significant differences between control and exposure groups. The results suggest the existence of mechanisms that allow mussels to maintain their hormonal status, and the important role that fatty acid esterification may play within those mechanisms. Synthesis and conjugation rates of estradiol were further investigated by measuring the activity of P450 aromatase, and palmitoyl-CoA:estradiol acyltransferase, in digestive gland microsomal fractions. Overall, the study contributes to the better knowledge of molluscan endocrinology, and defines new mechanisms of regulation of free steroid-levels in mussels.     

Polycyclic aromatic hydrocarbon (PAH) burden of mussels (Mytilus sp.) in different marine environments in relation with sediment PAH contamination, and bioavailability

Sediments and mussels (Mytilus edulis, Mytilus galloprovincialis) were sampled in different European coastal environments (Germany, France, Spain) and analysed for their polycyclic aromatic hydrocarbon (PAH) content by Gas Chromatography/Mass Spectrometry (GC-MS). Bioaccumulation factors of individual compounds from the sediment were calculated and discussed according to the compound solubility. The mussels showed different accumulation patterns according to the pollution source they were exposed to (dissolved fraction of PAHs, particulate fraction, petroleum present in the water column). The exposure source also depends on the geographical location of the mussels. In the Mediterranean Sea, the bivalves were mainly exposed to the dissolved fraction of PAHs, while in the Baltic Sea and in the Atlantic Ocean, the PAHs associated to the particles were significant sources.     

Implications from a field study regarding the relationship between polycyclic aromatic hydrocarbons and glutathione S-transferase activity in mussels

An aluminium smelter on the west coast of Scotland discharges an aqueous effluent containing polycyclic aromatic hydrocarbons (PAHs) at the head of Loch Leven. The loch also supports two mussel (Mytilus edulis) farms. Data are presented on burdens of PAHs in the soft tissues of mussels and the effect of these contaminants on glutathione S-transferase (GST) activity in mussel hepatopancreas. GST activity is shown to be correlated with total PAH burden and also with the concentrations of certain individual PAHs. These field data show that high molecular weight PAHs are closely correlated to GST activity, whereas low molecular weight PAHs are not. This suggests that 5- and 6-ring PAHs have a more pronounced role than 2- to 4-ring compounds in inducing GST activity in mussels from Loch Leven. It is proposed that it may be more appropriate to link GST activity with 5- and 6-ring compounds only, rather than with the total PAH burden.     

Concentrations of unsubstituted polynuclear aromatic hydrocarbons in bay mussels (Mytilus edulis) from Oregon, USA

Concentrations of fifteen unsubstituted polynuclear aromatic hydrocarbons (PNAH) were measured in Mytilus edulis from two sites in Yaquina Bay, Oregon, USA, during 1979–1980. There were significant differences in PNAH levels between the two populations. The average total concentration in mussels inhabiting the more industrialized bayfront was 986·2 μg/kg compared with 273·9 μg/kg in mussels from a more remote site across the bay. Substantial differences were found in the concentrations of different PNAH in M. edulis examined during this study. The smaller more water soluble, compounds were concentrated to one or two orders of magnitude above the larger, less soluble PNAH.     

福建台山列岛厚壳贻贝资源调查与评估

为了解海域厚壳贻贝栖息和资源补充情况,探索资源持续利用,作者对福建台山列岛周边海域厚壳贻贝(Mytilus coruscus)资源进行潜水调查及样方采样分析,研究了该海域厚壳贻贝的分布密度、生物量、生物学特征;结合旁侧声纳和测深仪对海底面状地貌与海底地形进行扫测,并对海底表层沉积物进行采样分析,研究了厚壳贻贝生境底质特点、栖息覆盖率等。台山列岛海域调查结果显示岩礁面积共2.189822km2,厚壳贻贝平均密度151.33个/m~2、平均生物量6.89 kg/m~2,平均质量45.55 g/个。计算显示当其栖息覆盖率达到10%、20%、30%和40%时,该海域厚壳贻贝资源增长量分别为520、1 039、1 559和2 079 t/a。目前该海域年均采挖量约为762.8~1 125 t/a。据此,只有当厚壳贻贝覆盖面积达到该岩礁区域面积的20%~30%时,其资源量才能得到有效补充。剖面调查结果表明目前该海域厚壳贻贝覆盖率仅为12%~24%,由此可见,该海域厚壳贻贝资源将面临资源枯竭风险,为更有效地保护和持续利用厚壳贻贝资源,必须采取有效的管理对策措施。     

Mortality and quantitative aspects of storage cell utilization in mussels, Mytilus edulis, following exposure to diesel oil hydrocarbons

These investigations assess the consequences of a resistance strategy for the nutrient storage matrix in mussels involving the reallocation of reserves previously directed towards gamete production, on exposure to low-level, chronic diesel oil hydrocarbons using stereological techniques. In addition, the extent to which survival is dependent upon the levels of these reserves at the moment of hydrocarbon insult is also investigated. The results indicate that hydrocarbon exposure leads to a reduction in the levels of storage reserves, an increase in gamete atresia (degeneration) and resorption but, as a result of this, the storage pool is partially replenished and, in the short term, the animals are better able to tolerate the hydrocarbon insult.     

Development of two novel CYP-antibodies and their use in a PCB exposure experiment with Mytilus edulis

In an attempt to learn more about the cytochrome P450 (CYP) system of mussels, we used protein databases and alignment software to extract highly conserved CYP sequences. From these alignments synthetic peptides were produced and used for rabbit immunisation, which yielded polyclonal antibodies against the CYP families 2 and 4. The antibodies were evaluated with Western Blot and ELISA assays, using digestive gland microsomal samples from the mussel Mytilus edulis. Western Blots revealed immunoreactions for both antibodies. The anti-CYP2 sequence rendered one major immunopositive protein of ≈49 kDa size, and weak signals for proteins of ≈41 and 56 kDa size. The anti-CYP4 sequence rendered two major bands of ≈56 and 59 kDa size, and also a weak immunoreaction with a protein of ≈43 kDa size. ELISA rendered only weak signals even with a 1:50 dilution of IgG-purified serum. A 10-day exposure to Aroclor 1254 did not appear to affect any of the immunopositive proteins, while total PCBs in soft bodies increased from 14–40 ng/g DW in controls to 373–638 ng/g DW in exposed mussels.     

A 15-month study of zooplankton ingestion by farmed mussels (Mytilus edulis) in Bantry Bay, Southwest Ireland

There is a growing body of evidence to suggest that bivalve molluscs routinely ingest zooplankton. To elucidate further these observations, a 15-month study of zooplankton ingestion by farmed mussels was conducted using mussel long-lines in Bantry Bay, Ireland. Stomach content analysis of the mussels showed that there was evidence of zooplankton ingestion throughout the sampling period, but that highest mean numbers of zooplankters were ingested by mussels in the spring and summer months. Various zooplankton species were present in mussel stomachs. Harpacticoid copepods were found more often in stomach contents than calanoid copepods, probably due to their proximity to the bivalves' inhalent siphons. Barnacle cyprids featured in large numbers in stomach contents, but only for a period of 3 months which broadly corresponded with their pelagic phase. Sizes of ingested zooplankton ranged from 126 μm to 6 mm, but more of the smaller zooplankters (e.g. crustacean nauplii) were ingested. When lengths of ingested copepods were compared with those found in plankton net samples, it was found that the net-sampled copepods were significantly larger than those found in mussel stomachs, suggesting that mussels select for smaller categories within the zooplankton available to them. Soft bodied zooplankton was rarely found in mussel stomachs but their absence may be due to rapid digestion or they may have been destroyed in the preservation process. Ingestion of zooplankton by bivalves is discussed in the context of the impacts mussel farms have on resident zooplankton populations.     

CYP1A- and CYP3A-immunopositive protein levels in digestive gland of the mussel Mytilus galloprovincialis from the Mediterranean Sea

CYP1A-immunopositive protein can be elevated in response to planar PAHs and PCBs in Mytilus sp. digestive gland whilst CYP3A-immunopositive protein has been associated with testosterone 6β-hydroxylation in fish. Levels of CYP1A- and CYP3A-immunopositive protein were determined in Mytilus galloprovincialis digestive gland microsomes collected from 12 sites in the Mediterranean Sea during May and September 2001. CYP1A-immunopositive protein was significantly highest at contaminated sites whilst CYP3A-immunopositive protein was significantly lowest. A weak negative correlation (r²=0.21) was seen between CYP1A- and CYP3A-immunopositive protein. Little evidence of differences at the different sampling times was observed. These results confirm previous work indicating elevation of CYP1A-immunopositive protein in Mytilus sp. digestive gland at contaminated sites. Further study is required to characterise CYP3A-like expression in Mytilus and to elucidate the consequences of possible CYP3A-like down-regulation at contaminated sites.     

花生四烯酸调控网络在贻贝麻痹性贝毒代谢中的作用基于转录组和代谢组联合分析

紫贻贝（Mytilus galloprovincialis）是富集麻痹性贝类毒素（Paralytic Shellfish Toxins, PSTs）能力很强的代表性双壳贝类,但研究表明紫贻贝暴露于PSTs也会引起机体炎症反应,其作用机理及对毒素代谢影响尚不清楚。本研究采用转录组学与代谢组学联合分析技术,比较了产PSTs的链状亚历山大藻（Alexandrium catenella）暴发的不同时期,紫贻贝机体的基因表达量与代谢物含量差异,以解析PSTs胁迫下紫贻贝机体的炎症反应机制。结果表明,紫贻贝暴露于PSTs后表达的基因和代谢物均发生显著变化,其中差异表达基因17232个,差异代谢物341个。基于联合分析,差异表达基因与差异代谢物显著富集在花生四烯酸和谷胱甘肽代谢通路中,基因PLA2G2F、ALOX5、TBXAS1以及代谢物ARA、PGH₂、TXA₂、LTA₄、5-HETE对贻贝机体的促炎反应发挥重要作用;而基因GPX4、CYP2J2和代谢物15-HETE、GSH则调节机体炎症的消退。本研究揭示花生四烯酸相关通路在贻贝机体炎症反应过程中具有重要的调控作用,为下一步深度揭示PSTs胁迫下贻贝机体炎症网络化响应机制提供了研究基础。     

Variation in levels of cytochrome P4501A, 2B, 2E, 3A and 4A-immunopositive proteins in digestive gland of indigenous and transplanted mussel Mytilus galloprovincialis in Venice Lagoon, Italy

Mytilus galloprovincialis digestive gland microsomes were prepared from (i) indigenous populations sampled from a clean reference (Lido) and an urban-contaminated site (Salute) and (ii) mussels transplanted for up to 3 weeks from Lido to an industrial-contaminated site (CVE) in Venice Lagoon, Italy. Western blot analysis was performed using antibodies to five mammalian or fish CYP forms (1A, 2B, 2E, 3A, 4A). Simultaneously run M. edulis digestive gland partially purified CYP aided identification of immunopositive bands. Levels of CYP1A, CYP2E, and CYP4A-immunopositive proteins were 50 to 300% higher in indigenous M. galloprovincialis from Salute compared to Lido (p < 0.05). Three weeks after transplantation to CVE, levels of only the CYP1A-immunopositive protein were determined to be higher (63%) than levels for Lido (p < 0.05), indicating that anti-CYP1A shows greater specificity for a contaminant-inducible CYP form than the other antibodies.     

Quantitative assessment of intertidal blue mussel (Mytilus edulis L.) stocks: combined methods of remote sensing, field investigation and sampling

The aim of this investigation was to develop appropriate methods for quantitative assessment of blue mussel beds. Combined methods of remote sensing, ground truth investigation and sampling in the field were applied to mature and young intertidal mussel beds. Three variables were measured to obtain reliable quantitative estimates of biomass and abundance: the total area covered by a bed; the cover, i.e. the area of mussel-covered patches in relation to the total mussel bed area; and the proportion, i.e. the area covered by mussels within the patches in relation to the total area of mussel patches. Cover and proportion of intertidal blue mussel beds were measured in the field by the transection method. Aerial photographs enabled the total area of the beds to be determined and large-scale surveys of mussel beds to be carried out synchronously. By examining large-scale aerial photographs it was possible also to determine the cover by remote sensing. The proportion could not be derived from aerial photographs, even from pictures at the largest scale tested during this investigation (scale 1:2500). By statistical methods it was estimated that 12 sub-samples (each covering 177 cm²) from mature beds, and 12 sub-samples (each covering 38 cm²) from young beds are needed to obtain reliable data on biomass and abundance.     

三种典型POPs对紫贻贝不同组织DNA损伤的比较研究

采用彗星电泳技术,比较研究了Aroclor 1254、BaP、DDT三种典型POPs对紫贻贝(Mytilus edulis)鳃、性腺、消化腺细胞的DNA损伤效应的差异,以期为进一步评价这三种典型POPs遗传毒理机制及其预警监测提供科学依据.结果表明,Tail DNA%是评价DNA损伤的理想指标,另外,在急性毒性试验中,...     

条斑星鲽幼鱼变态期间核酸及蛋白的变化

本试验测定了条斑星鲽(Verasper moseri)幼鱼变态过程中DNA、RNA、蛋白及其比值的变化,作为评价幼鱼变态阶段生长潜能的指标。结果表明,试验期间培育水温为16.5～18.0°C,DNA、RNA及蛋白的变化都带有发育阶段特异性。DNA含量在34～42日龄增加缓慢,然后快速增加直至44日龄,在45日龄和46日龄分别保持下降和上升趋势。蛋白含量和RNA含量的变化趋势相似,在试验期间保持快速增长的趋势。RNA/DNA比值从试验开始至35日龄呈上升状态,36日龄时下降,38日龄时达到最高值5.01,然后显著下降(P0.05),在试验结束时达到最低值1.76。蛋白/DNA在40日龄时达到最高值58.64,在46日龄时达到最低值21.28。RNA、DNA及蛋白含量跟全长和体质量有明显的线性关系(P0.05)。RNA/DNA与全长和体质量的关系比蛋白/DNA与全长和体质量的关系密切。蛋白/DNA比值的变化趋势跟RNA/DNA比值类似,但前者的变化滞后于后者,表明RNA/DNA比值是评价条斑星鲽生理状况的更有效的生理指标。     

Biochemical and histological responses in mussel (Mytilus edulis) exposed to North Sea oil and to a mixture of North Sea oil and alkylphenols

Several environmental chemicals are suspected to be responsible for adverse health effects on the reproductive system in various organisms. During this work, environmentally relevant concentrations of North Sea oil were used alone or in combination with alkylphenols and additional PAH to study the effect on vitellogenin-like protein expression and gonadal development in mussels. North Sea oil (0.5 ppm) induced the expression of phospho-proteins in both sexes indicating that some compounds are oestrogen-mimics. This induction was not seen in samples dosed with the mixture but signs of toxic effects were observed in the gonads. Indeed, numerous degenerating ovarian follicles in females and foci, similar to vertebrate melanomacrophage centres, were observed in testes.     

The blooms of a cyanobacterium, Microcystis cf. aeruginosa in a severely polluted estuary, the Golden Horn, Turkey

The distribution of toxic cyanobacterium Microcystis cf. aeruginosa in the severely polluted Golden Horn Estuary was studied from 1998 to 2000. Microcystis persisted at the upper estuary where the water circulation was poor and values ranged between 2.9 × 10⁴ and 2.7 × 10⁶ cells ml⁻¹ throughout the study. Simultaneously measured physical (salinity, temperature, rainfall and secchi disc) and chemical parameters (nutrients and dissolved oxygen) were evaluated together with Microcystis data. Although the Microcystis blooms generally occur in summer due to the increase in temperature, the blooms were recorded in winter in the present study. The abundance of Microcystis depended on the variations in salinity and both blooms were recorded below S = 2. A moderate partial correlation between Microcystis abundance and salinity was detected in the presence of temperature, dissolved oxygen and precipitation data (r = −0.561, p = 0.002). The M. cf. aeruginosa abundance was low in the summer when the salinity was higher than winter. A remarkable increase in the eukaryotic phytoplankton abundance following the improvements in the water quality of the estuary occurred, whilst the Microcystis abundance remained below bloom level.