Food sources used by sediment meiofauna in an intertidal Zostera noltii seagrass bed: a seasonal stable isotope study

In an intertidal Zostera noltii Hornem seagrass bed, food sources used by sediment meiofauna were determined seasonally by comparing stable isotope signatures (δ¹³C, δ¹⁵N) of sources with those of nematodes and copepods. Proportions of different carbon sources used by consumers were estimated using the SIAR mixing model on δ¹³C values. Contrary to δ¹⁵N values, food source mean δ¹³C values encompassed a large range, from −22.1 ‰ (suspended particulate organic matter) to −10.0 ‰ (Z. noltii roots). δ¹³C values of copepods (from −22.3 to −12.3 ‰) showed that they use many food sources (benthic and phytoplanktonic microalgae, Z. noltii matter). Nematode δ¹³C values ranged from −14.6 to −11.4 ‰, indicating a strong role of microphytobenthos and/or Z. noltii matter as carbon sources. The difference of food source uses between copepods and nematodes is discussed in light of source accessibility and availability.     

Quantitative alterations in the structure of the digestive cell of Littorina littorea on exposure to cadmium

The effects of sublethal concentrations of cadmium (Cd) on the morphology of the digestive gland of the periwinkle Littorina littorea have been investigated using a planimetric procedure. Five planimetric parameters were recorded (mean epithelial thickness: MET; mean diverticular radius: MDR; mean luminal radius: MLR; MLR/MET and MET/MDR) and tested by diverse statistical methods. The results indicate that MET is depressed after long exposures at high Cd concentrations whilst MLR increases. Variations in MDR have been attributed to different conditions in the tissues surrounding digestive tubules, and are not-significant in relation to the treatment. However, certain increases might be significant due to the presence of parasites or hemocyte proliferation. The ratios MLR/MET and MET/MDR are highly sensitive, and can be expressed as statistical functions of metal concentration and exposure time. Combination of these five planimetric parameters is reliably indicative of the structural changes occurring in the digestive gland of molluscs under conditions of environmental stress.     

Application of a battery of biomarkers in mussel digestive gland to assess long-term effects of the Prestige oil spill in Galicia and Bay of Biscay: tissue-level biomarkers and histopathology

In order to assess the biological effects of the Prestige oil spill (POS), mussels, Mytilus galloprovincialis, were collected in 22 localities along the North coast of the Iberian Peninsula over 3 years (April 2003-April 2006). Different tissue-level biomarkers including cell type composition (volume density of basophilic cells, Vv(BAS)) in digestive gland epithelium, structural changes of digestive alveoli (mean luminal radius to mean epithelial thickness, MLR/MET) and histopathological alterations (prevalence and intensity) of the digestive gland were analysed. Severe alterations in the general condition of the digestive gland tissue were observed all over the study area up to 2004-2005. High Vv(BAS) values were recorded mainly in Galicia but also to a lesser extent in the Bay of Biscay in 2003-2004. Atrophy of the digestive alveoli, measured in terms of MLR/MET, was detected all along the studied area up to 2006. Inflammatory responses cannot be related to pollution due to the POS: (a) prevalence and intensity of focal hemocytic infiltration were higher in the Bay of Biscay than in Galicia but they did not show a clear temporal trend; (b) high intensities of brown cell aggregates were only sporadically recorded; and (c) granulocytomas were more frequently recorded in the Bay of Biscay than in Galicia and especially in localities (i.e. Arrigunaga) subjected to chronic pollution. Likewise, Marteilia, trematodes, intracellular ciliates, unidentified eosinophilic bodies, R/CLO and Mytilicola did not follow any recognisable pattern that could be associated to the POS. In contrast, high Nematopsis intensities recorded in several localities in 2003 might suggest some response of local interest after the POS (i.e., in combination with particular factors/conditions). More data at a regional scale are needed before histopathology may provide a reliable ecosystem health assessment but the present results suggest that the approach is worthwhile. Overall, although Vv(BAS) returned to reference values by 2004-2005, MLR/MET values indicated that the mussel health condition was affected during the whole study period up to April 2006.     

Digestive cell turnover in digestive gland epithelium of slugs experimentally exposed to a mixture of cadmium and kerosene

Slugs, Arion ater (L), have been proposed as sentinel organisms to assess soil health. In slugs under the influence of pollutants, digestive cell loss and the concomitant increase of excretory cells of the digestive gland have been described. The aim of the present work was to determine up to what extent digestive cell loss affects biomarkers and whether the affectation is reversible after exposure to a mixture of metal and organic pollutants. Slugs were dosed with a mixture of cadmium and kerosene in the food for 27 days. Apart from chemical analyses, the volume density of black silver deposits (Vv(BSD)) after autometallography, and acyl-CoA oxidase (AOX) activity were used as biomarkers of exposure to metals and organic compounds, respectively. As effect biomarkers, changes in the volume density of the cell types that constitute the digestive gland epithelium were calculated. Proliferating cells were identified by means of bromodeoxyuridine (BrdU) immunohistochemistry. Results revealed that the mixture of pollutants provoked an increase in Vv(BSD) and AOX activity and a decrease in the number of digestive cells. These changes had no effect in the digestive gland accumulation capacity or in the effect and exposure biomarkers employed. BrdU-labelling showed that exposure to pollutants provoked an enhanced digestive cell proliferation.     

Application of a battery of biomarkers in mussel digestive gland to assess long-term effects of the Prestige oil spill in Galicia and the Bay of Biscay: lysosomal responses

In order to assess the long-term lysosomal responses to the Prestige oil spill (POS), mussels, Mytilus galloprovincialis, were collected in 22 localities from Galicia and the Bay of Biscay (North Iberian peninsula) in July, and September 2003, April, July, and October 2004-2005 and April 2006. Lysosomal membrane stability (labilisation period, LP) and lysosomal structural changes (lysosomal volume density, Vv(L) and lysosomal surface-to-volume ratio, S/V(L)) were measured as general stress biomarkers. The most remarkable long-term effects after the POS were drastic changes in lysosomal size (lysosomal enlargement) and membrane stability (extremely low LP values) up to April-04. Later on, a recovery trend was envisaged all along the studied area after July-04, albeit membrane stability continued to be below 20 min throughout the studied period up to April-06, which indicates a "distress-to-moderate-stress" condition. Lysosomal Response Index (LRI) revealed that environmental stress was more marked in Galicia than in the Bay of Biscay, mainly in the first sampling year, although a "moderate-to-high-stress" condition persisted until July-05. Overall, although lysosomal size returned to reference values, membrane stability was not fully recovered indicating a stress situation throughout the studied period.     

Bioavailability and cellular effects of metals on Lumbricus terrestris inhabiting volcanic soils

Whether the radial thickness (RT) of the chloragogenous tissue and intestinal epithelium of earthworms (Lumbricus terrestris) reflects the bioavailability of metals in soils was investigated in two areas, one with active volcanism (Furnas) and another with no volcanic activity since 3 million years ago (Santa Maria), in the Azores. Metal contents in soil samples and earthworms from the two areas were analyzed. Autometallography and measurements of the RT were performed in the chloragogenous tissue and intestinal epithelium. Earthworms from the active volcanic area demonstrated lower RT of chloragogenous tissue and intestinal epithelium as well as higher levels of bioavailable metals, especially Zn and Cd. Comparison of bioavailable metal contents between both areas suggests a higher risk for uptake of potentially toxic metals in the active volcanic area than in the non-active volcanic area, which is reflected by the lower RT of the chloragogenous tissue and intestinal epithelium in the former.     

Effects of the fuel oil spilled by the Prestige tanker on reproduction parameters of wild mussel populations

The aim of this work was to assess possible effects of the Prestige oil spill on reproduction parameters of mussels along the Galician and Bay of Biscay coast. Studied endpoints included sex ratio, gonad histology and vitellogenin-like proteins using the alkali-labile phosphate (ALP) method. A high prevalence of haemocytic infiltration of follicles and severe oocyte atresia was found in most localities in April 2003. Spawning gonads were observed in most impacted populations in the same sampling. In April 2004 mature small sized follicles were observed. No histopathological changes were observed in April 2005 and 2006, except a high prevalence of necrotic gametes in 6 out of 22 localities in April 2006. Female ALP levels showed high interindividual variability in April 2004, which was reduced in April 2005 and 2006. No xenoestrogenic effects were observed in male mussels. Overall, gamete alterations were detected during 2003-2004 and a recovery trend was observed afterwards.     

Application of a battery of biomarkers in mussel digestive gland to assess long-term effects of the Prestige oil spill in Galicia and Bay of Biscay: correlation and multivariate analysis

According to published reports, selected cell and tissue-level biomarkers revealed alterations in the health status of mussels collected after the Prestige oil spill (POS) in Galicia and the Bay of Biscay (2003-2006). Presently, univariate and multivariate analysis provides a comprehensive view of the whole data set. Correlation and principal component analysis (PCA) were performed using biomarkers corresponding to or derived from those data, say: induction, AOX(exp), and inhibition, AOX(eff), of Acyl-CoA oxidase enzyme activity; membrane labilisation period, LP, volume density, Vv(LYS), and surface-to-volume ratio, S/V(LYS), of digestive cell lysosomes; volume density of neutral lipids, Vv(NL); volume density of basophilic cells, Vv(BAS), mean luminal radius to mean epithelial thickness in digestive gland epithelium, MLR/MET; gonad index, GI; cumulative intensity of inflammatory responses, CI(IR) and parasitic infestation, CI(PI). The PCA results were used to classify sampling times and localities by means of hierarchical cluster analysis. Biomarkers were more frequently correlated with naphthalene than with total PAH concentration in mussel tissues. Many biological parameters were correlated with each other in agreement with their involvement in toxic processes or their role in the mussels' response against environmental insult. Naphthalene appears to be the most effective toxic herein, which supports that the reported effects may be mainly due to POS. According to the PCA, 4 groups of biomarkers explained 67.37% variability and the factors that most strongly contributed were S/V(LYS), CI(IR), Vv(BAS) and MLR/MET. These factors were used in the hierarchical cluster analyses carried out. The cluster analysis performed to classify sampling times discriminated 3 significantly different periods after POS: (a) a high affection period (until April-04); (b) the beginning of the recovery; and (c) an advanced recovery status in which geographical variability increased (from April-05 onwards). As regards the clustering of localities, major geographical differences were not detected but three outliers were identified: (a) early and highly impacted localities in Galicia and Bay of Biscay; (b) Agui?o that was characterised by extremely high Vv(BAS) during 2003; and (c) localities that were subjected to persistent chronic pollution.     

Assessment of biological effects of environmental pollution along the NW Mediterranean Sea using mussels as sentinel organisms

With the aim of assessing the biological effects of pollution along three gradients of pollution in the NW Mediterranean Sea, a biomonitoring survey was implemented using a battery of biomarkers (lysosomal membrane stability, lysosomal structural changes, metallothionein (MT) induction and peroxisome proliferation) in mussels over a period of two years as part of the EU-funded BEEP project. Mussels from the most impacted zones (Fos, Genova and Barcelona harbours) showed enlarged lysosomes accompanied by reduced labilisation period of lysosomal membranes, indicating disturbed health. MT levels did not reveal significant differences between stations and were significantly correlated with gonad index, suggesting that they were influenced by gamete development. Peroxisomal acyl-CoA oxidase (AOX) activity was significantly inhibited in polluted stations possibly due to interactions among mixtures of pollutants. In conclusion, the application of a battery of effect and exposure biomarkers provided relevant data for the assessment of biological effects of environmental pollution along the NW Mediterranean Sea.     

Toxicity assessment through multiple endpoint bioassays in soils posing environmental risk according to regulatory screening values

Toxicity profiles of two soils (a brownfield in Legazpi and an abandoned iron mine in Zugaztieta; Basque Country) contaminated with several metals (As, Zn, Pb and Cu in Legazpi; Zn, Pb, Cd and Cu in Zugaztieta) and petroleum hydrocarbons (in Legazpi) were determined using a multi-endpoint bioassay approach. Investigated soils exceeded screening values (SVs) of regulatory policies in force (Basque Country; Europe). Acute and chronic toxicity bioassays were conducted with a selected set of test species (Vibrio fischeri, Dictyostelium discoideum, Lactuca sativa, Raphanus sativus and Eisenia fetida) in combination with chemical analysis of soils and elutriates, as well as with bioaccumulation studies in earthworms. The sensitivity of the test species and the toxicity endpoints varied depending on the soil. It was concluded that whilst Zugaztieta soil showed very little or no toxicity, Legazpi soil was toxic according to almost all the toxicity tests (solid phase Microtox®, D. discoideum inhibition of fruiting body formation and developmental cycle solid phase assays, lettuce seed germination and root elongation test, earthworm acute toxicity and reproduction tests, D. discoideum cell viability and replication elutriate assays). Thus, albeit both soils had similar SVs, their ecotoxicological risk, and therefore the need for intervening, was different for each soil as unveiled after toxicity profiling based on multiple endpoint bioassays. Such a toxicity profiling approach is suitable to be applied for scenario-targeted soil risk assessment in those cases where applicable national/regional soil legislation based on SVs demands further toxicity assessment.