Marine Pollution Bulletin 58 (2009) 1025–1031

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Marine Pollution Bulletin

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Stable isotopes and metal contamination in caged marine musselMytilus galloprovincialis

S. Deudero a,*, A. Box a, S. Tejada a, J. Tintoré b

a Laboratory of Marine Biology, Department of Biology, University of Balearic Islands, Campus Universitari, Ctra. de Valldemossa s/n km. 7,5, 07022 Palma deMallorca, Balearic Islands, Spainb IMEDEA (CSIC-UIB) Instituto Mediterráneo de Estudios Avanzados, C/Miquel Marqués, 21. 07190 Esporles, Illes Balears, Spain

a r t i c l e i n f o a b s t r a c t

Keywords:MusselsIsotopesPollutionMetalsLeadCadmium

0025-326X/$ - see front matter � 2009 Elsevier Ltd. Adoi:10.1016/j.marpolbul.2009.02.011

* Corresponding author. Tel.: +34 971173138; fax:E-mail address: salud.deudero@uib.es (S. Deudero

Metal concentrations and isotopic composition were measured in different tissues of the mussel Mytilusgalloprovincialis in waters of the Balearic Islands (Western Mediterranean) in order to assess pollutionlevels. The isotopic composition was correlated with lead, cadmium, selenium and nickel obtained fromthe digestive gland and foot of the mussels. Significant negative correlations were found between cad-mium, selenium and zinc and the mussel foot, mainly for 13C. Significant correlations were also foundbetween lead and cadmium and the digestive gland. Pearson correlations indicated that the 13C isotopicsignal in foot is a good proxy for the concentration of metals such as lead, cadmium, selenium and zinc.Similarly, 15N isotopic signatures in the digestive gland reflected the lead and cadmium concentration.

� 2009 Elsevier Ltd. All rights reserved.

1. Introduction

Pollutants from industrial and agricultural sources that cancause adverse effects are increasing in marine ecosystems (Fuet al., 2003; Maanan, 2008). Knowledge of the sources of pollutionand their impacts on marine waters are essential for a betterunderstanding of the ecosystem response to pollutants. Marinepollution evaluation is based on direct measurements of the abioticcomponents and on measurements of the abundance and bioaccu-mulation of metals in selected marine organisms (Cope et al.,1996). Pollution of the environment can affect the health of livingorganisms, directly by uptake from the water or indirectly throughtheir diet. Consequently, the concentration of a pollutant in an-organism is a balance between intake and regulation. The accumu-lation of metals in organisms is used as a bioindicator for environ-mental quality assessment (Langston and Spence, 1995; Mertenset al., 2005).

Mussels are widely used as sentinel species for monitoringcoastal environments (Andral et al., 2004; Box et al., 2007; Deuderoet al., 2007a,b; Saavedra et al., 2004; Santovito et al., 2005) for sev-eral reasons: they have a large capacity to accumulate pollutants intheir tissues to a degree that is suitable for measurement, as seden-tary filter-feeders (Catsiki and Florou, 2006; Phillips, 1976) that fil-ter the water column (Angelo et al., 2007); they have lowdecontamination kinetics (Wang et al., 1996); they have suitabledimensions for their manipulation; easy to identify and collect (Ce-

ll rights reserved.

+34 971173184.).

vik et al., 2008); and they provide enough tissue for chemical anal-ysis (Angelo et al., 2007). The used of caged mussel Mytilusgalloprovincialis for the assessment of water quality is widespread(Andral et al., 2004; Romeo et al., 2003a,b) since mussels are com-mon in the Mediterranean, and the use of cages prevents geneticshifts between populations and makes it possible to control thesource and age of the samples (Andral et al., 2004).

Metals are major pollutants in the industrial world (Jarup,2003). These pollutants can cause long-term effects on marine eco-systems even if their impacts have no visible influence comparedto other pollutants. Their capacity to bind with short carbon chainscan be dangerous for marine organisms since they continuouslyaccumulate and concentrate metals in their tissues (Shahidul-Islam and Tanaka, 2004). Moreover, metals can interfere in the met-abolic processes of the organisms because most of these pollutantshave high affinities for sulphur, tending to bind with sulphydrylgroups of proteins and thus altering their structure (Davies, 1978).

The levels of metals in marine organisms depend on environ-mental contamination and also on several ecological factors suchas diet and trophic position (Das et al., 2004b). Through the diet,marine organisms assimilate and discriminate stable isotopes(Pinnegar and Polunin, 1999). In fact, several studies have appliedstable carbon and nitrogen isotopes as an effective method tostudy the food web (Fisher et al., 2001; Lorrain et al., 2002; Pinne-gar and Polunin, 2000). Stable isotope d13C values have allowed toindicate primary food sources, whereas d15N values have been usedto determine the trophic level that a specific organism occupies(Post, 2002; Vander Zanden et al., 1997). Moreover, changes instable isotopes have also been used to establish the relationship

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between the pollutants and the trophic level and pollutant transferinto the food web (Cheung and Wang, 2008; Das et al., 2004a;Domi et al., 2005). Previous studies have also described biomagni-fication of contaminants in food webs by relating contaminant con-centrations to stable nitrogen isotope values (Gray, 2002; Rigetet al., 2007). Therefore, assessing the uniformity of stable isotopedistribution among organs is essential to study the effects of pollu-tants on the marine organisms. Indeed, all parameters that affectmetabolism may affect fractionation processes directly linked tophysiological processes and may vary according to different envi-ronmental parameters (Dubois et al., 2007; Trueman et al., 2005)and locations (Dubois et al., 2007; Jennings and Warr, 2003).Therefore, enrichment processes are tissue-specific (Logan et al.,2006; Yokoyama et al., 2005). Previous studies have demonstratedfractionation in d13C and d15N signatures across tissues in marineorganisms ranging from primary producers (Vizzini et al., 2003)to consumers such as different fishes (Gaston and Suthers, 2004;Pinnegar and Polunin, 2000; Sweeting et al., 2007), turtles (Semi-noff et al., 2006) and dolphins (Das et al., 2000). In the case of fil-ter-feeders, it has been demonstrated that different tissues(digestive gland, muscle and gills) of the marine mussel M. gallo-provincialis showed different fractionation patterns (Deuderoet al., 2008).

The aim of this study was to determine metal concentrationsand stable isotope ratios in the caged mussel M. galloprovincialisin waters of the Balearic Islands and to assess pollution levels insentinel organisms by these complementary methodologies.

2. Material and methods

2.1. Sampling design

The mussels M. galloprovincialis were harvested in the Sete re-gion (South East France) under pristine conditions and were main-tained in the Thau lagoon (France) for one week beforetransplantation. The batch was composed of caged mussels of thesame genetic origin, age (18–24 months) and shell length (50 mm).

During March 2005, M. galloprovincialis were actively trans-planted (Andral et al., 2004) to different areas of the Balearic Is-lands (see Section 2.2) at depths of 20–35 m. The immersionperiod after transplantation was 3 months. This allowed the adap-tation of the mussels to the new environmental conditions (oligo-trophic waters) and a response in the isotopic composition of theirtissues (Holmer et al., 2007).

2.2. Sampling location

Fifteen sampling stations were selected surrounding the Bale-aric Islands, according to a gradient of anthropogenic load and de-gree of human impact (Box et al., 2007): Santa Maria Bay (Cabrera)and Espardell (Formentera) were selected as pristine areas, bothbeing protected marine areas; areas subject to human impact weresites in the vicinity of the harbours of Palma de Mallorca (Mallor-ca), Eivissa and San Antoni (Eivissa), Maó and Ciutadella (Menor-ca); other locations with different degrees of human impact wereSanta Eulàlia (Eivissa), Santa Ponsa, Porto Colom, Sóller, Calad’Or, Alcúdia (Mallorca), Cala Trebelutja and Cala Fornells (Menor-ca) (Fig. 1).

2.3. Sample collection and processing

The mussels M. galloprovincialis were sampled by experiencedscuba divers at each study site. After collection, M. galloprovincialisindividuals (n = 45; three samples for each location) were cleanedof epiphytes and immediately frozen at �20 �C until tissue extrac-

tion for chemical and isotopic analyses. Mussels were pre-pro-cessed in accordance to standard procedures (Andral et al., 2004).Fresh and dry weight measurements of the whole organism andshell were performed for each mussel. Mussels were opened andflesh was scraped out of the shell with a stainless steel scalpel.Shells were dried at 60 �C for 48 h before obtaining the dry weight.Flesh was weighed after freeze-drying. Measurements of fresh anddry weight of the digestive gland, foot and gills were performed ineach mussel in order to refer the results to the dry weight.

2.4. Metal determination

The tissue samples of the M. galloprovincialis were analysedfor the following metals: lead, cadmium, mercury, nickel, copper,selenium, zinc, arsenic and chromium. Analyses were performedon the mussel tissues that were freeze-dried and homogenised.The methodology used was described by Rodríguez et al.(1995). Mineralization was carried out at 130 �C for 5 h with ni-tric acid in a microwave oven using high pressure Teflon reac-tors. The quantitative analyses were carried out by atomicabsorption spectrophotometry (Perking–Elmer 4110-ZL) usinggraphite furnace or flame. Mercury was determined by flow-injection cold-vapour atomic absorption spectrometry (FIMS,Perking–Elmer). All chemical analyses were performed followingthe guidelines recommended for chemical contaminant monitor-ing in marine organisms (UNEP/FAO/IOC/IAEA, 1993). The inter-nal quality control was assured by testing control, blanks andduplicated samples in each analytical series. The external accu-racy of the analytical procedures was tested by using certifiedreference material and by participation in international inter-comparison exercises, such as the QUASIMEME programme. Onlyresults with a |Z| 6 2 were considered to be satisfactory in thestatistical analyses.

2.5. Stable isotope analyses

Mussels (n = 3 individuals per location) were dissected in orderto separate the digestive gland, foot and gills. All tissue samples(three samples for each tissue and each location) were dried at60 �C for 24 h and then grounded to a fine powder using a mortarand pestle. Homogeneous dried powder (2 mg ± 0.1) from each tis-sue was placed into cadmium tin cups and then combusted to ana-lyse d15N and d13C stable isotope composition by continuous flowisotope ratio mass spectrometry (CF-IRMS) using a THERMO DELTAX-PLUS mass spectrometer. Two samples of an internal referencematerial were analysed after every eighth sample in order to recal-ibrate the system and compensate for drift over time. The referencematerial used for carbon and nitrogen stable isotope analysis wasBovine Liver Standard (1577b) and Peach Leave Standard (1547),respectively (US Department of Commerce, National Institute ofStandards and Technology, Gaithersburg, MD 20899). The analyti-cal precision was based on the standard deviation of internal stan-dard replicates; these deviations were 0.09‰ and 0.21‰ for thePeach Leave Standard (PLS) for d13C, and 0.10‰ and 0.09‰ forthe BLS (Bovine Liver Standard) for d15N.

Stable isotope abundances were measured by comparing the ra-tio of the most abundant isotopes (13C:12C and 15N:14N) in the sam-ple with the international isotopic standards. Carbon and nitrogenstable isotopic ratios are expressed in d notation in terms of partsper thousand (‰) deviations from the standards according to thefollowing equation:

dX ¼ ½ðRsample=RreferenceÞ � 1� � 103

where X is 13C or 15N and R is the corresponding 13C/12C or 15N/14Nratio.

Fig. 1. Sampling locations distributed around the Balearic Islands. The mussels M. galloprovincialis were caught by scuba diving in March 2005.

S. Deudero et al. / Marine Pollution Bulletin 58 (2009) 1025–1031 1027

2.6. Statistical analysis

Relationships between stable isotope d13C and d15N values andthe studied metals were analysed by Pearson correlations (p < 0.05was considered statistically significant). Canonical analysis of prin-cipal coordinates (CAP) was applied to investigate the relationshipbetween d13C and d15N values of the tissues extracted from themussel M. galloprovincialis and the metal patterns in the differentstudied areas. All statistical computations were performed usingstatistical package PRIMER� 6.0 software.

3. Results

3.1. Mussel metal concentrations

The metal concentrations (expressed as mg kg�1 of dry mus-sel flesh) in the mussel M. galloprovincialis at fifteen differentsampling stations are shown in Table 1. The tissues of the mus-sels contained all analysed metals, showing a concentration upto the detection limit. Zinc, arsenic, selenium and copper werethe pollutants with the highest concentrations (154.5–289.5,6.02–11.56, 6.335–8.5, 4.295–5.19 mg/kg, respectively) in themussel M. galloprovincialis. Lead (1.816–3.345 mg/kg) and cad-mium (1.566–3.374 mg/kg) also presented high levels, followedby nickel (0.513–1.252 mg/kg), while mercury and chromium(0.158–0.256, 0.439–0.774 mg/kg, respectively) had the lowestconcentrations (Table 1).

3.2. Relations between isotopic composition and metals

The tissues of the mussel M. galloprovincialis differentially re-sponded to the bioaccumulation of metals as shown by Pearsoncorrelations (Table 2). The lead concentration in the digestive glandwas positively correlated with d15N; similarly, lead concentrationin the foot was positively correlated with the d13C isotopic compo-sition. Cadmium concentration in the digestive gland and foot wasinversely correlated with the d15N and d13C isotopic composition,respectively. The concentrations of selenium and zinc in the footof the mussels were also inversely correlated the d13C isotopiccomposition. With regards the digestive gland, the isotopic compo-sition of nitrogen portrayed a negative correlation, being the cad-mium the metal with a more statistical significance for allstudied Pearson correlations (Table 2).

Canonical analysis of principal coordinates (CAP) was per-formed with m = 4 constrained axes. CAP for the studied metalsshowed a different trend of distribution among the studied locali-ties for the carbon (Fig. 2A) and nitrogen (Fig. 2B) isotopic signa-tures. CAP performed on the 13C dataset (Fig. 2A) showed a shiftof the sampling locations that were strongly associated with theenvironmental variation in zinc, cadmium and arsenic, while leadwas strongly correlated with the harbour of Palma. The patternemerging from the 15N correlations differed from that of 13C inthe sense that most stations presented a strong correlation withnickel, lead and mercury and on the first axis a strong correlationwith zinc and arsenic.

Table 1Metal concentrations (mg/kg of dry weight) of lead (Pb), cadmium (Cd), mercury (Hg), nickel (Ni), copper (Cu), selenium (Se), zinc (Zn), arsenic (As) and chromium (Cr) quantifiedin the collected mussels M. galloprovincialis from different localities of the Balearic Islands (Western Mediterranean). Heavy metals in waters of the Balearic Islands (WesternMediterranean).

Locality Heavy metals (mg/kg dry weight)

Pb Cd Hg Ni Cu Se Zn As Cr

Alcúdia 2.307 3.008 0.256 1.124 5.150 7.685 238.5 9.930 0.542Santa Maria 2.306 2.470 0.213 1.252 4.560 7.275 289.5 11.56 0.509Cala d’Or 1.816 3.374 0.206 0.759 4.495 6.630 233.0 6.955 0.562Ciutadella 2.229 3.310 0.204 1.016 5.115 6.505 238.0 8.210 0.478Eivissa 2.101 2.791 0.188 0.768 4.860 6.450 274.5 8.515 0.573Espardell 3.212 3.212 0.245 1.263 4.660 7.930 251.0 9.555 0.566Fornells 3.252 3.252 0.194 1.148 4.915 7.080 220.5 6.020 0.516Maó 2.965 2.965 0.181 0.711 4.675 6.335 223.0 7.895 0.439Palma 3.345 1.556 0.158 0.513 4.620 5.220 154.5 7.745 0.454Porto Colom 2.046 2.721 0.194 0.728 4.645 6.890 230.5 10.145 0.495Sant Antoni 2.479 2.792 0.181 0.858 4.735 8.500 225.0 10.995 0.534Santa Eulàlia 2.135 2.699 0.219 0.980 4.445 7.880 227.5 8.515 0.507Santa Ponsa 2.113 2.460 0.190 1.11 4.295 6.635 226.5 10.14 0.494Sóller 2.291 2.535 0.190 0.837 5.190 7.090 229.0 10.85 0.774Trebelutja 1.896 3.229 0.237 0.880 5.060 7.025 251.5 8.695 0.444

Table 2Pearson correlations (rho values) between lead (Pb), cadmium (Cd), mercury (Hg), nickel (Ni), copper (Cu), selenium (Se), zinc (Zn), arsenic (As) and chromium (Cr) and the tissuesdigestive gland, foot and gills of the mussels M. galloprovincialis.

Tissue Heavy metals

(‰) Pb Cd Hg Ni Cu Se Zn As Cr

Digestive gland d13C ns ns ns ns ns ns ns ns nsd15N 0.614a �0.653b ns ns ns ns ns ns ns

Foot d13C 0.551a �0.595a ns ns ns �0.553a �0.616a ns nsd15N ns ns ns ns ns ns ns ns ns

Gills d13C ns ns ns ns ns ns ns ns nsd15N ns ns ns ns ns ns ns ns ns

ns indicates non significant values.a Indicates p < 0.05.b Indicates p < 0.01.

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4. Discussion

In the present work, the pollution levels in waters of the Bale-aric Islands were assessed by comparing metal concentrations(lead, cadmium, mercury, nickel, copper, selenium, zinc, arsenicand chromium) with the carbon and nitrogen isotopic compositionof the digestive gland, foot and gill tissues from the mussel M. gal-loprovincialis. Correlations showed that the digestive gland andfoot of the mussel are good indicators of lead and cadmium con-centration, while the gills did not show significant correlationsfor any studied metals. CAP illustrated a shift in the 13C structureof sampling locations that was strongly associated with zinc, cad-mium, and arsenic, while lead was correlated with Palma harbour.The 15N pattern showed a strong correlation between most stationsand nickel, lead and mercury, and on the first axis with zinc andarsenic.

The decontamination kinetic is usually slower than that of con-tamination (Wang and Wang, 2006). Metal concentrations in themussel M. galloprovincialis can provide a measure of the effects ofpotential pollutants in the marine environment. However, littledata are available on the pollutant content in mussels (Ceviket al., 2008) in the localities studied in the current work. Previousstudies on chemical pollutants in cultured mussels in the WesternMediterranean (Balearic Islands) showed a decreasing trend overthe last ten years (Deudero et al., 2007a,b). In the present study,pollutants were assessed at a mesoscale level (1428 km of coast-line) to avoid the influence of factors such as variations in physico-chemical characteristics and food availability in the immersionzones (Andral et al., 2004). Metal concentrations in M. galloprovin-

cialis were also low in the current study compared with previousworks on metal pollution in mussels (Andral et al., 2004; Ceviket al., 2008). Cadmium and lead were the exceptions since the con-centrations found in the present work were higher than those de-scribed in the literature (Campbell et al., 2005; Conti and Cecchetti,2003; Pedersen et al., 1997). These values can be considered asindicative of pollution in the mussels since cadmium and leadare potential immuno-suppressants (Cantillo, 1998; Law et al.,1999). Zinc reached high concentration levels in the mussels M.galloprovincialis in the present work, which could be related tothe regulation of the bivalves since they are capable of accumulat-ing this metal in their tissues, and it may not be transferred alongthe food web; consequently, increasing concentrations in the foodweb are limited (Cheung and Wang, 2008).

Several studies have demonstrated that the pollution factor(Rogers, 2003) and tissue turnovers (Herzka and Holt, 2000; Mac-Avoy et al., 2001; Maruyama et al., 2001) are linked to the isotopiccomposition. In fact, the measurement of carbon and nitrogen iso-topic ratios has been used as a technique to investigate the rela-tionship between the trophic level and pollution in marineorganisms (Cheung and Wang, 2008; Das et al., 2004a; Domiet al., 2005). The carbon isotopic signatures have been suggestedto be a more appropriate sewage tracer for mussels, which filterthe effluent’s particulate organic matter from the water. In con-trast, nitrogen signatures are a better tracer of pollution in sea-weed and associated grazers (Rogers, 2003). In addition, the useof bivalves and the study of isotopic signatures might enable chro-nological assessment of pollutant exposure (Gustafson et al., 2007).The present study showed the existence of differences in the isoto-

Fig. 2. Canonical analysis of principal coordinates (CAP) plot of the metal concentrations, relating sampling stations 13C data (A) and 15N (B) for digestive gland, foot and gilltissues of the mussel M. galloprovincialis around the Balearic Islands (Western Mediterranean Sea). Symbols: filled triangle: digestive gland; open circles: foot; and opensquares: gills. Locations: Alc, Alcúdia; Cab: Cabrera; Or: Cala d’Or; Ciu: Ciutadella; Ei: Eivissa; Esp: Espardell; Fll: Fornells; Mh: Maó; PC: Porto Colom; SA: Sant Antoni; SE:Santa Eulàlia; SP: Santa Ponsa; Tre: Trebelutja.

S. Deudero et al. / Marine Pollution Bulletin 58 (2009) 1025–1031 1029

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pic composition of the analysed tissues of M. galloprovincialisacross sampling stations with a different degree of contamination.This is consistent with previous studies in which differences in M.galloprovincialis isotopic composition of 13C and 15N across stationswith different degree of contamination emerged in M. galloprovin-cialis (Rogers, 2003). Similarly, spatial variation in the isotopic sig-nal in fish tissues between contaminated and non-contaminatedzones has been described (Gaston and Suthers, 2004).

The correlations between metals and the isotopic compositionfrom the different studied tissues suggest that the signal obtainedby the stable isotope 13C for the digestive gland represents the bestproxy for lead and cadmium, which were positively and inverselycorrelated with the isotopic signal, respectively. Previous studieshave established that the isotopic signature of 13C is a good estima-tor of contamination in mussels (Rogers, 2003) and that the diges-tive gland shows quick turnover that reflects short-termassimilation and thus generates a fast response to environmentalconditions (Raikow and Hamilton, 2001). Similarly, the signal of-fered by the stable isotope 15N for the foot of the mussels can beconsidered as a chemical contamination proxy for lead, cadmium,selenium and zinc, which were inversely correlated with nitrogensignatures, except for lead, which presented a positive correlation.Bivalves mainly bioaccumulate mercury available from the envi-ronment in their gills (Lakshmanan and Nambisan, 1989; Micallefand Tyler, 1990) indicating the existence of contamination by thismetal. In fact, in a previous study the highest mercury concentra-tion in the bivalve Perna viridis was found in gill tissue (Roesijadiand Fellingham, 1987). In the current work, it is remarkable thatmercury did not show any significant correlations in the threestudied tissues (digestive gland, foot and gills), even though it isone of the most pollutant metals in the marine environment dueto its bioaccumulation capacity in the tissues of organisms. Fur-thermore, the gills did not show any statistically significant corre-lation with the studied metals. Mercury levels in the mussels M.galloprovincialis were low in comparison with other studies onmussels (Andral et al., 2004; Deudero et al., 2007b) and P. viridis(Liu and Kueh, 2005). For this reason, it is possible that correlationfailed to indicate the degree of pollution in these marine organismsbecause of the low bioaccumulation in the tissues due to the lowmercury concentration.

Different pollutant loads may be responsible for differences inthe isotopic signal across localities in the studied M. galloprovincial-is. The stations with similar pollution conditions showed minimalvariation in the isotopic composition in M. galloprovincialis (Rogers,2003). Consequently, the presence of metals and the site character-istics will be responsible for the changes in the isotopic composi-tion of the mussel M. galloprovincialis and other marineorganisms by bioaccumulation through the food web. Secondly,the metal concentrations found in the present study can be addedto the bibliography of pollution measures, providing new dataabout the metal levels in the waters around the Balearic Islands.Knowledge from the study of metals in seawater, together withtraditional biomonitoring investigations, are complementary forthe development of environmental protection policies in thefuture.

Acknowledgements

The authors thank M. Ribas (IUNICS) for the collaboration in iso-topic analysis, D. March for mussel samples preparation, and IFR-EMER and the Nantes laboratory for the analytical lipiddeterminations. We also thank the collaboration of M. Cabanellasand J. Alós. This study was partly financed by the Interreg MedocIIIB, UE Project: ‘Development d’un réseau de surveillance de la qua-lité des eaux côtières par des biointégrateurs pour la protection dura-ble de la Méditerranée Occidentale (MYTILOS)’. A. Box and S. Tejada

received a fellowship sponsored by the MYTILOS project InterregMedoc IIIB. This study complies with the present Spanish law.

References

Andral, B., Stanisiere, J.Y., Sauzade, D., Damier, E., Thebault, H., Galgani, F., Boissery,P., 2004. Monitoring chemical contamination levels in the Mediterranean basedon the use of mussel caging. Marine Pollution Bulletin 49, 704–712.

Angelo, R.T., Cringan, M.S., Chamberlain, D.L., Stahl, A.J., Haslouer, S.G., Goodrich,C.A., 2007. Residual effects of lead and zinc mining on freshwater mussels in theSpring River Basin (Kansas, Missouri, and Oklahoma, USA). Science TotalEnvironmental 384, 467–496.

Box, A., Sureda, A., Galgani, F., Pons, A., Deudero, S., 2007. Assessment ofenvironmental pollution at Balearic Islands applying oxidative stressbiomarkers in the mussel Mytilus galloprovincialis. Comparative Biochemistryand Physiology C-Toxicology and Pharmacology 146, 531–539.

Campbell, L.M., Norstrom, R.J., Hobson, K.A., Muir, D.C., Backus, S., Fisk, A.T., 2005.Mercury and other trace elements in a pelagic Arctic marine food web(Northwater Polynya, Baffin Bay). Science Total Environmental 351–352, 247–263.

Cantillo, A.Y., 1998. Comparison of results of mussel watch programs of the UnitedStates and France with worldwide mussel watch studies. Marine PollutionBulletin 36, 712–717.

Catsiki, V.A., Florou, H., 2006. Study on the behavior of the heavy metals Cu, Cr, Ni,Zn, Fe, Mn and 137Cs in an estuarine ecosystem using Mytilus galloprovincialis asa bioindicator species: the case of Thermaikos gulf, Greece. Journal ofEnvironmental Radioactivity 86, 31–44.

Cevik, U., Damla, N., Kobya, A.I., Bulut, V.N., Duran, C., Dalgic, G., Bozaci, R., 2008.Assessment of metal element concentrations in mussel (Mytilusgalloprovincialis) in Eastern Black Sea, Turkey. Journal of Hazardous Materials.doi:10.1016/j.jhazmat.2008.03.010.

Conti, M.E., Cecchetti, G., 2003. A biomonitoring study: trace metals in algae andmolluscs from Tyrrhenian coastal areas. Environmental Research 93, 99–112.

Cope, W.G., Wiener, J.G., Steingraeber, M.T., 1996. Test system for exposing fish toresuspended, contaminated sediment. Environmental Pollution 91, 177–182.

Cheung, M.S., Wang, W.X., 2008. Analyzing biomagnification of metals in differentmarine food webs using nitrogen isotopes. Marine Pollution Bulletin 56, 2082–2105.

Das, K., Holsbeekc, L., Browninga, J., Siebertb, U., Birkun, Jr.d.A., Bouquegneau, J.M.,2004a. Trace metal and stable isotope measurements (d13C and d15N) in theharbour porpoise Phocoena phocoena relicta from the Black Sea. EnvironmentalPollution 131, 197–204.

Das, K., Lepoint, G., Loizeau, V., Debacker, V., Dauby, P., Bouquegneau, J.M., 2000.Tuna and dolphin associations in the North-east Atlantic: evidence of differentecological niches from stable isotope and heavy metal measurements. MarinePollution Bulletin 40, 102–109.

Das, K., Siebert, U., Fontaine, M., Jauniaux, T., Holsbeek, L., Bouquegneau, J.M., 2004b.Ecological and pathological factors related to trace metal concentrations inharbour porpoises Phocoena phocoena from the North Sea and adjacent areas.Marine Ecology Progress Series 281, 283–295.

Deudero, S., Box, A., March, D., Valencia, J.M., Grau, A.M., Tintore, J., Calvo, M.,Caixach, J., 2007a. Organic compounds temporal trends at some invertebratespecies from the Balearics, Western Mediterranean. Chemosphere 68, 1650–1659.

Deudero, S., Box, A., March, D., Valencia, J.M., Grau, A.M., Tintore, T., Benedicto, J.,2007b. Temporal trends of metals in benthic invertebrate species from theBalearic Islands, Western, Mediterranean. Marine Pollution Bulletin 54, 1545–1558.

Deudero, S., Cabanellas, M., Blanco, A., Tejada, S., 2008. Stable isotope fractionationin the digestive gland, muscle and gills tissues of the marine mussel Mytilusgalloprovincialis. Journal of Experimental Marine Biology and Ecology 31, 181–188.

Domi, N., Bouquegneau, J.M., Das, K., 2005. Feeding ecology of five commercialshark species of the Celtic Sea through stable isotope and trace metal analysis.Marine Environmental Research 60, 551–569.

Dubois, S., Jean-Louis, B., Bedrtrand, B., Lefebvre, S., 2007. Isotope trophic-stepfraction of suspension-feeding species: implications for food partitioning incoastal ecosystems. Journal of Experimental Marine Biology and Ecology 351,121–128.

Fisher, S.J., Brown, M.L., Willis, D.W., 2001. Temporal food web variability in anupper Missouri River backwater: energy origination points and transfermechanisms. Ecology of Freshwater Fish 10, 154–167.

Fu, J., Mai, B., Sheng, G., Zhang, G., Wang, X., Peng, P., Xiao, X., Ran, R., Cheng, F., Peng,X., Wang, Z., Tang, U.W., 2003. Persistent organic pollutants in environment ofthe Pearl River Delta, China: an overview. Chemosphere 52, 1411–1422.

Gaston, T.F., Suthers, I.M., 2004. Spatial variation in d13C and d15N of liver, muscleand bone in a rocky reef planktivorous fish: the relative contribution of sewage.Journal of Experimental Marine Biology and Ecology 304, 17–33.

Gray, J.S., 2002. Biomagnification in marine systems: the perspective of an ecologist.Marine Pollution Bulletin 45, 46–52.

Gustafson, L., Showers, W., Kwak, T., Levine, J., Stoskopf, M., 2007. Temporal andspatial variability in stable isotope compositions of a freshwater mussel:implications for biomonitoring and ecological studies. Oecologia 152, 140–150.

Herzka, S.Z., Holt, G.J., 2000. Changes in isotopic composition of red drum (Sciaenopsocellatus) larvae in response to dietary shifts: potential applications to

S. Deudero et al. / Marine Pollution Bulletin 58 (2009) 1025–1031 1031

settlement studies. Canadian Journal of Fisheries and Aquatic Sciences 57, 137–147.

Holmer, M., Marba, N., Diaz-Almela, E., Duarte, C.M., Tsapakis, M., Danovaro, R.,2007. Sedimentation of organic matter from fish farms in oligotrophicMediterranean assessed through bulk and stable isotope (d13C and d15N)analyses. Aquaculture 262, 268–280.

Jarup, L., 2003. Hazards of heavy metal contamination. British Medical Bulletin 68,7–182.

Jennings, S., Warr, K.J., 2003. Environmental correlates of large-scale spatialvariation in the d15N of marine animals. Marine Biology 142, 1131–1140.

Lakshmanan, P.T., Nambisan, P.N.K., 1989. Bioaccumulation and depuration of sometrace-metals in the mussel, Perna viridis (Linnaeus). Bulletin of EnvironmentalContamination and Toxicology 43, 131–138.

Langston, W.J., Spence, S.K., 1995. Biological factors involved in metal concentrationobserved in aquatic organisms. In: Tessier, A.T.D. (Ed.), Metal Speciation andBioavailability in Aquatic Systems. Wiley, Chichester, England, pp. 407–478.

Law, R.J., Blake, S.J., Spurrier, C.J.H., 1999. Butylene compounds in liver tissues ofpelagic cetaceans stranded on the coasts of England and Wales. MarinePollution Bulletin 38, 1258–1261.

Liu, J.H., Kueh, C.S., 2005. Biomonitoring of heavy metals and trace organics usingthe intertidal mussel Perna viridis in Hong Kong coastal waters. MarinePollution Bulletin 51, 857–875.

Logan, J., Haas, H., Deegan, L., Gaines, E., 2006. Turnover rates of nitrogen stableisotopes in the salt marsh mummichog, Fundulus heteroclitus, following alaboratory diet switch. Oecologia 147, 391–395.

Lorrain, A., Paulet, Y.M., Chauvaud, L., Savoye, N., Donval, A., Saout, C., 2002.Differential d13C and d15N signatures among scallop tissues: implications forecology and physiology. Journal of Experimental Marine Biology and Ecology275, 47–61.

Maanan, M., 2008. Heavy metal concentrations in marine molluscs from theMoroccan coastal region. Environmental Pollution 153, 176–183.

MacAvoy, S.E., Macko, S.A., Garman, G.C., 2001. Isotopic turnover in aquaticpredators: quantifying the exploitation of migratory prey. Canadian Journal ofFisheries and Aquatic Sciences 58, 923–932.

Maruyama, A., Yamada, Y., Rusuwa, B., Yuma, M., 2001. Change in stable nitrogenisotope ratio in the muscle tissue of a migratory goby, Rhinogobius sp., in anatural setting. Canadian Journal of Fisheries and Aquatic Sciences 58, 2125–2128.

Mertens, J., Luyssaert, S., Verheyen, K., 2005. Use and abuse of trace metalconcentrations in plant tissue for biomonitoring and phytoextraction.Environmental Pollution 138, 1–4.

Micallef, S., Tyler, P.A., 1990. Effect of mercury and selenium on the gill function ofMytilus edulis. Marine Pollution Bulletin 21, 288–292.

Pedersen, B., Cofino, W., Davies, I., 1997. The 1993–1995 QUASIMEME laboratoryperformance study: trace metals in sediment and biota. Marine PollutionBulletin 35, 42–51.

Phillips, D.J.H., 1976. Common mussel Mytilus edulis as an indicator of pollution byzinc, cadmium, lead and copper II. Relationship of metals in mussel to thosedischarged by industry. Marine Biology 38, 71–80.

Pinnegar, J.K., Polunin, N.V.C., 1999. Differential fractionation of d13C and d15Namong fish tissues: implications for the study of trophic interactions. FunctionalEcology 13, 22–231.

Pinnegar, J.K., Polunin, N.V.C., 2000. Contributions of stable-isotope data toelucidating food webs of Mediterranean rocky littoral fishes. Oecologia 122,399–409.

Post, D.M., 2002. Using stable isotopes to estimate trophic position: models,methods, and assumptions. Ecology 83, 703–718.

Raikow, D.F., Hamilton, S.K., 2001. Bivalve diets in a midwestern US stream: a stableisotope enrichment study. Limnology and Oceanography 46, 514–522.

Riget, F., Moller, P., Dietz, R., Nielsen, T.G., Asmund, G., Strand, J., Larsen, M.M.,Hobson, K.A., 2007. Transfer of mercury in the marine food web of WestGreenland. Journal of Environmental Monitoring 9, 877–883.

Roesijadi, G., Fellingham, G.W., 1987. Influence of Cu, Cd, and Zn preexposure on Hgtoxicity in the mussel Mytilus edulis. Canadian Journal of Fisheries and AquaticSciences 44, 680–684.

Rogers, K.M., 2003. Stable carbon and nitrogen isotope signatures indicate recoveryof marine biota from sewage pollution at Moa Point, New Zealand. MarinePollution Bulletin 46, 821–827.

Romeo, M., Hoarau, P., Garello, G., Gnassia-Barelli, M., Girard, J.P., 2003a. Musseltransplantation and biomarkers as useful tools for assessing water quality in theNW Mediterranean. Environmental Pollution 122, 369–378.

Romeo, M., Mourgaud, Y., Geffard, A., Gnassia-Barelli, M., Amiard, J.C., Budzinski, H.,2003b. Multimarker approach in transplanted mussels for evaluating waterquality in Charentes, France, coast areas exposed to different anthropogenicconditions. Environmental Toxicology 18, 295–305.

Saavedra, Y., Gonzalez, A., Fernandez, P., Blanco, J., 2004. The effect of size on tracemetal levels in raft cultivated mussels (Mytilus galloprovincialis). Science TotalEnvironmental 318, 115–124.

Santovito, G., Piccinni, E., Cassini, A., Irato, P., Albergoni, V., 2005. Antioxidantresponses of the Mediterranean mussel, Mytilus galloprovincialis, toenvironmental variability of dissolved oxygen. Comparative Biochemistry andPhysiology C-Toxicology and Pharmacology 140, 321–329.

Seminoff, J.A., Jones, T.T., Eguchi, T., Jones, D.R., Dutton, P.H., 2006. Stable isotopediscrimination (d13C and d15N) between soft tissues of the green sea turtleChelonia mydas and its diet. Marine Ecology Progress Series 308, 271–278.

Shahidul-Islam, M., Tanaka, M., 2004. Impacts of pollution on coastal and marineecosystems including coastal and marine fisheries and approach formanagement: a review and synthesis. Marine Pollution Bulletin 48, 624–649.

Sweeting, C.J., Barry, J.T., Polunin, N.V.C., Jennings, S., 2007. Effects of body size andenvironment on diet-tissue d13C fractionation in fishes. Journal of ExperimentalMarine Biology and Ecology 352, 165–176.

Trueman, C.N., Field, J.H., Dortch, J., Charles, B., Wroe, S., 2005. Prolongedcoexistence of humans and megafauna in Pleistocene Australia. Proceedingsof the National Academy of Sciences of the Unites States of America 102, 8381–8385.

Vander Zanden, M.J., Cabana, G., Rasmussen, J.B., 1997. Comparing trophic positionof freshwater fish calculated using stable nitrogen isotope ratios (d15N) andliterature dietary data. Canadian Journal of Fisheries and Aquatic Sciences 54,1142–1158.

Vizzini, S., Sara, G., Mateo, M.A., Mazzola, A., 2003. D13C and d15N variability inPosidonia oceanica associated with seasonality and plant fraction. AquaticBotany 76, 195–202.

Wang, W.X., Fisher, N.S., Luoma, S.N., 1996. Kinetic determinations of trace elementbioaccumulation in the mussel Mytilus edulis. Marine Ecology Progress Series140, 91–113.

Wang, X., Wang, W.-X., 2006. Bioaccumulation and transfer of benzo(a)pyrene in asimplified marine food chain. Marine Ecology Progress Series 312, 101–111.

Yokoyama, H., Tamaki, A., Harada, K., Shimoda, K., Koyama, K., Ishihi, Y., 2005.Variability of diet-tissue isotopic fractionation in estuarine macrobenthos.Marine Ecology Progress Series 296, 115–128.