Analysis of lipid peroxidation in animal and plant tissues as field-based biomarker in Mediterranean irrigated agroecosystems (Extremadura,Spain)

ABSTRACT

The development of field-based biomarkers can allow for a more reliable assessment of the exposure of organisms to pollutants. Different sampling sites, along two streams running through an irrigable agricultural area, were selected to evaluate the effect of agrochemical load on the measured endpoints. The levels of lipid peroxidation were evaluated in several organs of Procambarus clarkii. The same method was applied to leaves of two woody species. Determining levels of MDA (malonaldehyde) by thiobarbituric acid reactive substances assay allows measuring the levels of lipid peroxidation. Differences in lipid peroxidation levels were observed in P. clarkii individuals collected at different sites; however, the patterns varied depending on the organ (when accounting for variations due to sex). The use of a MDA-gills/MDA-hepatopancreas index allowed for discrimination between reference and polluted sites. Significant differences in oxidative damage between sites were found in the leaves of Quercus rotundifolia but not in Salix sp. The lipid peroxidation of crayfish organs and holm oak leaves as a suitable biomarker of environmental pollution deserves further investigation.     

Effects of Ni, Cr, Hg, Cu, Zn, Al on the dissolved oxygen balance of streams

Inorganic metals besides their direct toxic effects, carry the potential of causing serious variations on existing ecosystems in receiving waters. Self purification mechanism is vital for the continuity of the existing micro and macro living organisms in the streams. This mechanism is effected from the existence of metals. In this study, interferences of HgCl2, HgSO4, Ni(NO3)2, CrCl3, CuSO4.5H2O, K2Cr2O7, ZnSO4.7H2O and Al2(SO4)3.18H2O metal compounds on respirometric BOD and related effects on the self purification, are investigated with the representative formations of DO deficit curves. In the presence of these metals, streamwater BOD parameters and related DO deficit curve formations are significantly effected.     

Ecotoxicological impact assessment of the brine discharges from a desalination plant in the marine waters of the Algerian west coast,using a multibiomarker approach in a limpet,Patella rustica

The aim of our study is to evaluate the impact of Bousfer desalination plant brine discharges on the Algerian west coast, on a natural population of the marine gastropod mollusc Patella rustica. The effects of a chronic exposure to such discharges are complex to understand due to the combined effects of environmental physico-chemical parameters (e.g., high salinity) and different pollutants that can modulate the physiological responses of this species to stress. In this context, we assessed the biological effects in a marine species P. rustica, by a multibiomarker approach that provided information on the health status of organisms in response to such an environmental stress. We measured biomarkers in the whole soft tissues of limpets as indicators of neurotoxicity (AChE activity), oxidative stress (CAT, SOD, GR, and GPx activities), biotransformation (GST), oxidative damage (LPO through TBARS levels), and genotoxicity (CSP 3-like activity). In parallel, hydrological parameters were measured in the Bay of Oran, at four selected sites: site H considered as a “hotspot,” located at Bousfer desalination plant; two other sites E and W, at the east and the west of H respectively; finally, site R “reference” located in Madragh, which is considered as a remote clean site. Our analyses revealed that the activities of antioxidant defense enzymes reached the highest levels in P. rustica collected from site H. The activation of antioxidant defense system in these organisms translated the alteration of their status health, reflecting a level of environmental disruption generated by the desalination plant brine discharges and the high salinity in this area. We also observed that the tissues of limpets collected from site H as well as the two other sites, E and W, had undergone molecular damage, confirmed by the high levels of CSP 3-like activity. This damage resulted from chronic exposure to environmental conditions, potentially genotoxic, due to the desalination plant discharges. The present results indicate the adverse impact of brine effluents from desalination plants on marine fauna and suggest the need for a more consistent approach to environmental management of brine discharges.

     

The influence of untreated wastewater to aquatic communities in the Balatuin River,The Philippines

A risk assessment of chemical constituents in rivers that receive untreated wastewater should take into account the adverse effects of increased biological oxygen demand (BOD), ammonia and reduced dissolved oxygen (DO). This concept was tested via a field study in the Balatuin River, The Philippines, where the influence of physical and chemical factors, including the consumer product chemical linear alkylbenezene sulfonate (LAS), to aquatic communities (algae, invertebrates, fish) was determined. Periphytic algae were found to be insensitive to high BOD (>10 mg/l) and ammonia (>0.01 mg unionized NH(3)/l), concentrations from organically enriched untreated wastewater discharges. However, taxa richness and abundance of macroinvertebrates were influenced greatly by the discharges. Where BOD and ammonia concentrations were elevated, the dominant taxa were oligochaete worms and chironominds. Fish and crustaceans (freshwater crabs and prawns) were found only in sites with the least BOD concentrations (furthest upstream and downstream). The maximum concentration of LAS (0.122 mg/l) was less than that expected to affect 5% of taxa (0.245 mg/l), whereas exceedences of DO and ammonia criteria were observed in several sites. The lack of recovery observed was attributed to influences of low DO, high ammonia and poor colonization from upstream and downstream reaches due to organically-enriched discharges     

Eco-assessment of streams of Konya closed river basin (Turkey) using various ecoregional diatom indices

Ecological assessment of freshwater ecosystems based on diatom metrics is an important issue for attaining environmental sustainability. The present study aimed to evaluate differences in the diatom–stressor relationship in relatively least disturbed streams in the Konya closed river basin using multivariate analyses and to bio-assess streams by the application of different ecoregional diatom indices. Cocconeis euglypta, Cymbella excisa, Cocconeis placentula, and Achnanthidium minutissimum are the most contributing species to the dissimilarity of sampling stations between rainy (spring) and dry (summer and fall) seasons and also between altitude (A2 800- < 1600 m and A3 ≥ 1600 m) groups. The first two axes of canonical correspondence analysis revealed a significant (82.8%) relationship between diatom species and stressors. Diatom species displayed distinct responses to environmental variables (electrical conductivity, Ni, Cu, B, and altitude) playing important roles on the distribution of species. Diatom indices indicate different ecological statuses of stations, from bad to high. European diatom indices except Duero Diatom Index (DDI) and Trophic Diatom Index (TDI) showed good responses to the eco-assessment of streams and indicated high ecological status for the least disturbed sampling stations symbolized as S16, S20, S24, S25, S27-29, S37, and S39. These results were also supported by abiotic evaluation. Although TIT was more competitive in the bio-assessment of streams among diatom indices, it is necessary to increase its species list by determining their trophic weights in future studies. Therefore, the use of ecoregion-specific diatom indices is suggested along with increasing the number of used species to correctly interpret the water quality.

Graphical Abstract

     

Algae community and trophic state of subtropical reservoirs in southeast Fujian, China

Background,

aim, and scope Fujian reservoirs in southeast China are important water resources for economic and social sustainable development, although few have been studied previously. In recent years, growing population and increasing demands for water shifted the focus of many reservoirs from flood control and irrigation water to drinking water. However, most of them showed a rapid increase in the level of eutrophication, which is one of the most serious and challenging environmental problems. In this study, we investigated the algae community characteristics, trophic state, and eutrophication control strategies for typical subtropical reservoirs in southeast Fujian.

Materials and methods

Surface water samples were collected using polyvinyl chloride (PVC) plastic bottles from 11 Fujian reservoirs in summer 2010. Planktonic algae were investigated by optical microscopy. Water properties were determined according to the national standard methods.

Results and discussion

Shallow reservoirs generally have higher values of trophic state index (TSI) and appear to be more susceptible to anthropogenic disturbance than deeper reservoirs. A total of 129 taxa belonging to eight phyla (i.e., Bacillariophyta, Chlorophyta, Chrysophyta, Cryptophyta, Cyanophyta, Euglenophyta, Pyrrophyta, Xanthophyta) were observed and the most diverse groups were Chlorophyta (52 taxa), Cyanophyta (20 taxa), Euglenophyta (17 taxa), Chrysophyta (14 taxa). The dominant groups were Chlorophyta (40.58%), Cyanophyta (22.91%), Bacillariophyta (21.61%), Chrysophyta (6.91%). The species richness, abundance, diversity, and evenness of algae varied significantly between reservoirs. TSI results indicated that all 11 reservoirs were eutrophic, three of them were hypereutrophic, six were middle eutrophic, and two were light eutrophic. There was a strong positive correlation between algal diversity and TSI at P?4-N, NO _x -N, TP, and chlorophyll a were significant environmental variables affecting the distribution of algae communities. The transparency and chlorophyll a were the strongest environmental factors in explaining the community data. Furthermore, the degradation of water quality associated with excess levels of nitrogen and phosphorus in Fujian reservoirs may be impacted by interactions among agriculture and urban factors. A watershed-based management strategy, especially phosphorus control, should be developed for drinking water source protection and sustainable reservoirs in the future.

Conclusion and recommendations

All investigated reservoirs were eutrophicated based on the comprehensive TSI values; thus, our results provided an early warning of water degradation in Fujian reservoirs. Furthermore, the trophic state plays an important role in shaping community structure and in determining species diversity of algae. Therefore, long-term and regular monitoring of Euglenophyta, Cyanophyta, TN, TP and chlorophyll a in reservoirs is urgently needed to further understand the future trend of eutrophication and to develop a watershed-based strategy to manage the Cyanophyta bloom hazards.     

A novel thermotolerant Pediococcus acidilactici B-25 strain for color, COD, and BOD reduction of distillery effluent for end use applications

The present study was aimed to characterize physico-chemical and microbial population of distillery effluent and isolate a novel thermotolerant bacterium for color, COD, and BOD reduction of spentwash. The level of alkalinity, TSS, DO, COD, BOD, TN, ammonical nitrogen, nitrate nitrogen, phosphorous, potassium, chloride, and calcium of spentwash (SW), bioreactor effluent (BE), and secondary treated effluent (STE) were well above the permissible limits. The level of color, TS, and TDS were under the permissible limits for STE but not for SW and BE. The microbial population was higher in BE. The results revealed that effluent was highly polluted and require suitable treatment before discharge. A novel thermotolerant bacterium, identified as Pediococcus acidilactici, was isolated which exhibited maximum 79 % decolorization, 85 % COD, and 94 % BOD reduction at 45 °C using 0.1 %, glucose; 0.1 %, peptone; 0.05 %, MgSO4; 0.05 %, K₂HPO₄; pH 6.0 within 24 h under static condition. The ability of this strain to decolorize melanoidin at minimum carbon and nitrogen supplementation warrants its possible application for effluent treatment at industrial level. In addition, it is first instance when melanoidin decolorization was reported by P. acidilactici. This study could be an approach towards control of environmental pollution and health hazards of people in and around the effluent distillery unit.     

International cross-validation of a BOD5 surrogate

BOD5 dates back to 1912 when the Royal Commission decided to use the mean residence time of water in the rivers of England, 5 days, as a standard to measure the biochemical oxygen demand. Initially designed to protect the quality of river waters from extensive sewage discharge, the use of BOD5 has been quickly extended to waste water treatment plants (WWTPs) to monitor their efficiency on a daily basis. The measurement has been automatized but remains a tedious, time- and resource-consuming analysis. We have cross-validated a surrogate BOD5 method on two sites in France and in the USA with a total of 109 samples. This method uses a fluorescent redox indicator on a 96-well microplate to measure microbial catabolic activity for a large number of samples simultaneously. Three statistical tests were used to compare surrogate and reference methods and showed robust equivalence.     

Development and Integration of Quantitative Real-Time PCR Methods for Detection of Mitochondrial DNA and Methanobrevibacter smithii nifH Gene as Novel Microbial Source Tracking Tools

A novel microbial source tracking (MST) method based on the detection of human and non-human markers was developed and applied to track the origin of fecal pollution in water systems. Mitochondrial DNA sequences were used to develop new quantitative real-time polymerase chain reaction (qPCR) assays for dog, poultry, and gull. The targets were included as part of a toolbox including human, cow, pig, and sheep assays. A primer and probe set for the detection of the human-specific nifH gene of Methanobrevibacter smithii was also designed as an indicator of human fecal contamination. The assays were tested for specificity and applied to fecal-spiked surface waters and environmental samples collected from two river catchments impacted by sources of human and non-human fecal contamination. The MST methods described were applicable to both spiked waters and environmental samples, and using the two approaches the origin of fecal pollution could be successfully determined in mixed source fecally polluted waters.     

The impact of climate change on three indicator Galliformes species in the northern highlands of Pakistan

The rise in global temperature is one of the main threats of extinction to many vulnerable species by the twenty-first century. The negative impacts of climate change on the northern highlands of Pakistan (NHP) could change the species composition. Range shifts and range reduction in the forested landscapes will dramatically affect the distribution of forest-dwelling species, including the Galliformes (ground birds). Three Galliformes (e.g., Lophophorus impejanus, Pucrasia macrolopha, and Tragopan melanocephalus) are indicator species of the environment and currently distributed in NHP. For this study, we used Maximum Entropy Model (MaxEnt) to simulate the current (average for 1960–1990) and future (in 2050 and 2070) distributions of the species using three General Circulation Models (GCMs) and two climate change scenarios, i.e., RCP4.5 (moderate carbon emission scenario) and RCP8.5 (peak carbon emission scenario). Our results indicated that (i) under all three climate scenarios, species distribution was predicted to both reduce and shift towards higher altitudes. (ii) Across the provinces in the NHP, the species were predicted to average lose around one-third (35%) in 2050 and one-half (47%) by 2070 of the current suitable habitat. (iii) The maximum area of climate refugia was projected between the altitudinal range of 2000 to 4000 m and predicted to shift towards higher altitudes primarily?>?3000 m in the future. Our results help inform management plans and conservation strategies for mitigating the impacts of climate change on three indicator Galliforms species in the NHP.

     

Metabolic diversity of the heterotrophic microorganisms and potential link to pollution of the Rouge River

The heterotrophic microbial communities of the Rouge River were tracked using Biolog Ecoplates to understand the metabolic diversity at different temporal and spatial scales, and potential link to river pollution. Site less impacted by anthrophogenic sources (site 1), showed markedly lower metabolic diversity. The only substrates that were utilized in the water samples were carbohydrates. Sites more impacted by anthrophogenic sources (sites 8 and 9) showed higher metabolic diversity. Higher functional diversity was linked to the physico-chemical and biological properties of the water samples (i.e. higher concentrations of DO, DOC, chlorophyll, and bacterial density). Biolog analysis was found to be useful in differentiating metabolic diversity between microbial communities; in determining factors that most influence the separation of communities; and in identifying which substrates were most utilized by the communities. It can also be used as an effective ecological indicator of changes in river function attributable to urbanization and pollution.     

The effect of residential and agricultural runoff on the microbiology of a Hawaiian ahupua'a.

The objective of this project was to study the relationship between environmental runoff and the incidence of antibiotic-resistant microorganisms (ARMO) in freshwater streams. Five water systems along the windward coast of the island of O'ahu were evaluated. Samples were collected from sites upstream of residential or agricultural areas, throughout these areas, and at sites of entrance into oceans or bays. It was hypothesized that the incidence of ARMO would increase as the stream received runoff from residential and agricultural areas. The percentage of ARMO did not increase as the streams passed through residential or agricultural areas. Surprisingly, pristine sites, well upstream from residential or agricultural areas, contained bacteria resistant to at least one antibiotic. Areas most affected by runoff did not show a significant increase in the incidence of antibiotic-resistant organisms, suggesting that the incidence of antibiotic resistance is not simply a function of contamination with agricultural or residential runoff. The correlation of antibiotic resistance with heavy metal resistance was evaluated, because others (Fasim et al., 1999; Lazar et al., 2002; Nies, 1999) have shown that antibiotic and heavy metal resistance are each carried on extrachromosomal plasmids. The vast majority of ARMO were also resistant to concentrations of heavy metals reported in the sediments of indicator streams (Waihee, system III), suggesting that an antibiotic-resistant bacterium has a high probability of having dual resistance to a heavy metal. A 3.2-kb plasmid (pSTAMP) was isolated from a bacterium with dual antibiotic and heavy metal resistance. Further analysis of the plasmid is currently in progress.     

Cultivation of freshwater microalgae in biodiesel wash water

Biodiesel wash water is a contaminating industrial effluent that must be treated prior to disposal. The use of this effluent as a low-cost alternative cultivation medium for microalgae could represent a viable supplementary treatment. We cultivated 11 microalgae species with potential use for biodiesel production to assess their growth capacities in biodiesel industrial washing waters. Only Monoraphidium contortum, Ankistrodesmus sp., Chlorococcum sp., and one unidentified Chlorophyceae species grew effectively in that effluent. M. contortum showed the highest growth capacity and had the second highest fatty acid content (267.9 mg g⁻¹ of DW), predominantly producing palmitic (20.9%), 7,10,13-hexadecatrienoic (14%), oleic (16.2%), linoleic (10.5%), and linolenic acids (23.2%). In the second phase of the experiment, the microalgae were cultivated in biodiesel wash water at 75% of its initial concentration as well as in WC (control) medium. After 21 days of cultivation, 25.8 and 7.2% of the effluent nitrate and phosphate were removed, respectively, and the chemical oxygen demand was diminished by 31.2%. These results suggest the possibility of cultivating biodiesel producing microalgae in industrial wash water effluents.

     

Biodegradation of s-triazine herbicide atrazine by Enterobacter cloacae and Burkholderia cepacia sp. from long-term treated sugarcane-cultivated soils in Kenya

In this study soils from sugarcane-cultivated fields were screened for bacterial species capable of atrazine (6-chloro-N²-ethyl-N⁴-isopropyl-1,3,5-triazine-2,4-diamine) degradation due to long exposure of the soils to this herbicide. To enrich for atrazine degraders, Minimal Salt Medium containing atrazine as the sole N source and glucose as the C source was inoculated with soils impacted with this herbicide and incubated. Bacterial growth was monitored by measuring optical density. The degradation of atrazine was followed by measuring residual atrazine in liquid cultures over a given time period by high performance liquid chromatography. Bacterial strains isolated from the enrichment cultures were characterized by biochemical tests and identified by 16S rRNA gene sequencing. Two bacterial strains coded ISL 8 and ISL 15 isolated from two different fields were shown to have 94 and 96% 16S rRNA gene sequence similarity to Burkholderia cepacia respectively. Another bacterial sp., ISL 14 was closely related to Enterobacter cloacae with a 96% 16S rRNA gene sequence similarity. There was not much difference between the extents of atrazine degradation by the enrichment cultures with communities (79–82% applied amount) from which pure strains were isolated and the pure strains themselves in liquid cultures that showed a degradation of 53–83% of applied amount. The study showed existence of bacterial strains in different sugarcane-cultivated fields which can use atrazine as a nitrogen source. The bacterial strains isolated can be used to enhance the degradation of atrazine in contaminated soils where atrazine is still considered to be recalcitrant.     

Biodegradation of persistent environmental pollutants by Arthrobacter sp.

Persistent environmental pollutants are a growing problem around the world. The effective control of the pollutants is of great significance for human health. Some microbes, especially Arthrobacter, can degrade pollutants into nontoxic substances in various ways. Here, we review the biological properties of Arthrobacter adapting to a variety of environmental stresses, including starvation, hypertonic and hypotonic condition, oxidative stress, heavy metal stress, and low-temperature stress. Furthermore, we categorized the Arthrobacter species that can degrade triazines, organophosphorus, alkaloids, benzene, and its derivatives. Metabolic pathways behind the various biodegradation processes are further discussed. This review will be a helpful reference for comprehensive utilization of Arthrobacter species to tackle environmental pollutants.

     

Fungal biodegradation of CCA-treated wood and removal of its metal components

In the present study, 5 isolates of brown-rot fungi were used for fungal bioprocessing (FB) of chromated copper arsenate (CCA)-treated wood wastes: Antrodia vaillantii SEL8501, Fomitopsis palustris TYP0507 and TYP6137, and Crustoderma sp. KUC8065 and KUC8611. The isolates showed notable capacity for the degradation of treated wood and removal of CCA components via the American Wood Protection Association soil block test. Among them, Crustoderma sp. KUC8611 effectively decayed the treated wood, causing a mass loss of up to 60%. F. palustris caused extensive leaching of CrO₃ of up to 79% and As₂O₅ of up to 87%, but only moderate leaching of CuO of up to 50%. This high capacity for removal of CrO₃ and As₂O₅ showed a strong logarithmic relationship with the amount of oxalic acid produced in the decayed wood. The majority of metals removed from treated wood during the decay process were deposited in the soil and feeder strip. Further investigation will be required to establish the capability of selected fungi for FB of full-sized lumber treated with CCA.     

Distribution and trends of mercury in deciduous tree cores

The distribution of total mercury (THg) within common deciduous trees and the applicability of tree cores as biomonitors of historical environmental THg trends were assessed for both contaminated and reference sites around Kingston, Ontario. Samples were collected from Acer spp., Quercus spp. Populus spp. and Salix spp. Bark and wood THg concentrations were found to be highly correlated whereas soil and wood THg concentrations were not. There were no temporal relationships for THg in dated tree rings corresponding with any other known environmental Hg trends. The shoreline speciess, Populus and Salix spp., had the greatest bark and wood Hg concentrations reaching 18 ng/g, significantly higher than for inland trees Quercus and Acer spp. with maximum values of 7 and 1.2 ng/g for bark and wood respectively. While tree cores cannot be reliably used as temporal THg biomonitors, there is promise for tree species such as Populus spp and Salix spp as spatial indicators of local long-term Hg contamination.     

Use of phytoproductivity data in the choice of native plant species to restore a degraded coal mining site amended with a stabilized industrial organic sludge

Coal mining-related activities result in a degraded landscape and sites associated with large amounts of dumped waste material. The arid soil resulting from acid mine drainage affects terrestrial and aquatic ecosystems, and thus, site remediation programs must be implemented to mitigate this sequential deleterious processes. A low-cost alternative material to counterbalance the affected physico-chemical-microbiological aspects of the degraded soil is the amendment with low contaminated and stabilized industrial organic sludge. The content of nutrients P and N, together with stabilized organic matter, makes this material an excellent fertilizer and soil conditioner, fostering biota colonization and succession in the degraded site. However, choice of native plant species to restore a degraded site must be guided by some minimal criteria, such as plant survival/adaptation and plant biomass productivity. Thus, in this 3-month study under environmental conditions, phytoproductivity tests with five native plant species (Surinam cherry Eugenia uniflora L., C. myrianthum–Citharexylum myrianthum, Inga–Inga spp., Brazilian peppertree Schinus terebinthifolius, and Sour cherry Prunus cerasus) were performed to assess these criteria, and additional biochemical parameters were measured in plant tissues (i.e., protein content and peroxidase activity) exposed to different soil/sludge mixture proportions. The results show that three native plants were more adequate to restore vegetation on degraded sites: Surinam cherry, C. myrianthum, and Brazilian peppertree. Thus, this study demonstrates that phytoproductivity tests associated with biochemical endpoint measurements can help in the choice of native plant species, as well as aiding in the choice of the most appropriate soil/stabilized sludge proportion in order to optimize biomass production.

     

Diamondback terrapins as indicator species of persistent organic pollutants: Using Barnegat Bay, New Jersey as a case study

The diamondback terrapin’s (Malaclemys terrapin) wide geographic distribution, long life span, occurrence in a variety of habitats within the saltmarsh ecosystem, predatory foraging behavior, and high site fidelity make it a useful indicator species for contaminant monitoring in estuarine ecosystems. In this study fat biopsies and plasma samples were collected from males and females from two sites within Barnegat Bay, New Jersey, as well as tissues from a gravid female and blue mussels (Mytilus edulis), which are terrapin prey. Samples were analyzed for persistent organic pollutants (POPs), including polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), chlorinated pesticides, and methyl-triclosan. Terrapins from the northern site, Spizzle Creek, closest to influences from industrial areas, had higher POP concentrations for both tissues than terrapins from the less impacted Forsythe National Wildlife Refuge. Sex differences were observed with males having higher contaminant concentrations in fat and females in plasma. PCB patterns in terrapin fat and plasma were comparable to other wildlife. An atypical PBDE pattern was observed, dominated by PBDEs 153 and 100 instead of PBDEs 47 and 99, which has been documented in only a few other turtle species. The typical PBDE patterns measured in mussels, terrapin prey, suggests that the terrapin may efficiently biotransform or eliminate PBDE 47 and possibly PBDE 99. Plasma contaminant concentrations significantly and positively correlated with those in fat. This study addresses several aspects of using the terrapin as an indicator species for POP monitoring: site and sex differences, tissue sampling choices, maternal transfer, and biomagnification.     

Seasonal variation of pollution biomarkers to assess the impact on the health status of juvenile Pacific oysters <Emphasis Type="BoldItalic">Crassostrea gigas</Emphasis> exposed in situ

Background, aim, and scope  

In this study, a suite of sublethal stress biomarkers were analyzed in juveniles of the sentinel species, the Pacific oyster Crassostrea gigas, with a view to using them as pollution monitoring tools. The aim of this work was (1) to study baseline seasonal variations of biomarkers in different body compartments of C. gigas in the reference site and, after selecting biomarkers presenting no seasonal variations, (2) to compare responses of these biomarkers between contaminated and reference sites.