environmental and human health risk

Environ Sci Pollut Res DOI 10.1007/s11356-016-7945-x

RESEARCH ARTICLE

Metal bioaccumulation in two edible cephalopods in the Gulf of Gabes, South-Eastern Tunisia: environmental and human health risk assessment Lotfi Rabaoui 1,2 & Radhouan El Zrelli 3 & Rafik Balti 4 & Lamjed Mansour 1,5 & Pierre Courjault-Radé 3 & Nabil Daghbouj 6 & Sabiha Tlig-Zouari 1

Received: 31 March 2016 / Accepted: 20 October 2016 # Springer-Verlag Berlin Heidelberg 2016

Abstract Samples of Octopus vulgaris and Sepia officinalis were collected from four areas in the Gulf of Gabes, southeastern Tunisia, and their edible tissues (mantle and arms) were analyzed for cadmium, copper, mercury, and zinc. While the concentrations of metals showed significant differences between the sampling sites, no differences were revealed between the tissues of the two species. The spatial distribution of metals analyzed showed similar pattern for both tissues of the two species, with the highest concentrations found in the central area of Gabes Gulf, and the lowest in the northern and/or southern areas. From a human health risk point of view, the highest values of estimated daily intake, target hazard quotient, and hazard index were found in the central area of Gabes Gulf. Although the results of these Responsible Editor: Philippe Garrigues * Lotfi Rabaoui [email protected]; [email protected]

1

Research Unit of Integrative Biology and Evolutionary and Functional Ecology of Aquatic Systems, Faculty of Science of Tunis, University of Tunis El Manar, B.P 94 Cité Rommana, University Campus, 1068 Tunis, Tunisia

2

Marine Studies Section, Center for Environment and Water, King Fahd University of Petroleum and Minerals, Dhahran, Saudi Arabia

3

Geosciences Environment Toulouse (GET), UMR 5563 CNRS/UPS/ IRD/CNES, Université de Toulouse, 14 avenue Edouard Belin, 31400 Toulouse, France

4

Enzymes and Bioconversion Unit, National School of Engineering of Sfax, University of Sfax, Km 4 Road Soukra, 3038 Sfax, Tunisia

5

Zoology Department, College of Science, King Saud University, Riyadh, Saudi Arabia

6

CEMES-CNRS and Université de Toulouse, 29 rue J. Marvig, 31055 Toulouse, France

indices were, in general, not alarming, the health risks posed by the consumption of cephalopods on local consumers cannot be excluded. Keywords Trace metals . Cephalopods . Gulf of Gabes . Human health risk assessment . Estimated daily intake . Target hazard quotient

Introduction Because of the worldwide continuous development of human activities, various anthropogenic contaminants, in particular trace metals, end in marine coastal environments (Ruilian et al. 2008). Trace metals which are considered as major pollutants of the marine systems pose a serious risk when the environmental concentrations of these contaminants exceed standard values (Larsen 1992; Readman et al. 1993; Buchholtz Ten Brink et al. 1996). Because of their toxicity, absorbability, persistence, and bioaccumulation properties, trace metals can be incorporated in food chains of marine systems (Muirhead and Furness 1988; Bustamante et al. 1998; Lahaye et al. 2005) posing serious threats not only to living organisms but also to humans (De Forest et al. 2007; Amiard et al. 2008). In marine systems, numerous species, in particular benthic organisms, accumulate not only essential but also non-essential metals. Many of these metals bioaccumulators constitute prey for many carnivorous predators including the edible cephalopods which were reported to accumulate higher concentrations of trace metals in their tissues (Miramand and Bentley 1992; Bustamante et al. 1998, 2000; Seixas et al. 2005a, b; Raimundo et al. 2008; Pereira et al. 2009). Most of these latter studies focused on the common octopus, Octopus vulgaris Cuvier 1797 and the common cuttlefish Sepia officinalis Linnaeus 1758 which have a wide distribution area and are among the most edible cephalopod

Environ Sci Pollut Res

species. In the Mediterranean Sea, several studies have been carried out about the trace metals bioaccumulation in O. vulgaris (Miramand and Guary 1980; Nessim and Riad 2003; Rjeibi et al. 2014, 2015) and S. officinalis (LacoueLabarthe et al. 2009a, b; Ayas and Ozogul 2011; Duysak et al. 2013; Rjeibi et al. 2014, 2015). Similar to the case of most southern Mediterranean countries, only few studies have been recently conducted in Tunisia in this regard (Rjeibi et al. 2014, 2015). However, it is worth noting that these latter works did not cover the entire Tunisian coastline and they were limited only to the coasts extending between Bizerte, located at the north, and Sfax, located at the northern border of Gabes Gulf (GG). The latter authors did not examine samples from other areas including the Gulf of Gabes which is considered as one of the heavily trace metals-enriched environment in Tunisia due to the pressures posed by the industrial complex of Ghannouch-Gabes on the marine environment (Lahbib et al. 2013; Ayadi et al. 2014; Rabaoui et al. 2014, 2015; El Zrelli et al. 2015). Besides, the exploitation of cephalopod stocks is very important in the Gulf of Gabes, in particular those of O. vulgaris and S. officinalis. These two latter species are widely consumed by the people living in the area of Gabes Gulf, in particular by local fishermen. Therefore, it is important to conduct a study on the bioaccumulation of trace metals in the edible parts of O. vulgaris and S. officinalis in the Gulf of Gabes in order not only to assess the health status of the marine environment in this area but also to assess the human health risk of the consumption of these two latter cephalopod species. The present study was conducted in this regard with the main objectives of (i) evaluating the metal bioaccumulation in the two edible muscle tissues (mantle and arms) of O. vulgaris and S. officinalis collected from four different sites covering almost the entire area of Gabes Gulf, (ii) discussing the metallic pollution of the marine environment in this area based on the concentrations of trace metals found, and (iii) assessing the human health risk that may come from the consumption of these two cephalopods in Tunisia.

Material and methods Sample collection In winter–spring 2015, 39 specimens of O. vulgaris and S. officinalis have been collected, with the help of local fishermen, from four areas (3 to 7 specimens of each species from each area) extending from the south to the north of Gabes Gulf (GG), south-eastern Tunisia: Elbibane Lagoon, located in the south of GG: 5 O. vulgaris + 7 S. officinalis; Gabes, located in the central area of GG: 4 O. vulgaris + 5 S. officinalis; and Kerkennah Island and Chebba, both located in the north of GG: 6 O. vulgaris + 5 S. officinalis and 3 O. vulgaris + 4 S. officinalis, respectively (Fig. 1). The average dorsal mantle lengths of all O. vulgaris and S. officinalis collected were

127 ± 19 and 116 ± 21 mm, respectively. From each specimen collected, samples of edible tissues (mantle and arms) were collected, preserved separately in clean polythene bags, kept in an ice chest, and transported immediately to the laboratory for further processing. Sample preparation and trace metal analyses In the laboratory, collected tissues were freeze-dried separately. Thereafter, subsamples (1–2 g of freeze-dried tissues) were taken from each tissue sample (mantle or arms), weighted, placed in polyethylene tubes, submerged with 5 mL concentrated HNO3, and then left in the covered tubes for 24 h at room temperature. The samples were thereafter digested by exposing them to a temperature of 95 °C for 30 min. After removing the samples from the heat and cooling them at room temperature, additional 2 mL HNO3 were added and the samples were heated again for approximately 30 min. The same procedure was repeated until the tissue fibers were visibly broken down. After that, 5 mL of HCl were added and the samples were heated for 15 min. The obtained—digested— samples were then allowed to cool to room temperature and their volumes were adjusted at 50 mL by adding distilled water. The samples were then centrifuged and the obtained digests were decanted and analyzed in triplicate for Cd, Cu, and Zn using ICP-MS. For mercury (Hg) analysis, homogenized freeze-dried subsamples were analyzed by direct combustion on a direct mercury analyzer. Prior to chemical analysis, the moisture content (MC: weight of water (g)/weight of edible tissue (g)) of the analyzed tissues (i.e., mantle and arms) were determined for both cephalopod species, and the dry matter (DM: estimated as the dried weight (g)/weight of the wet edible tissue (g)) was calculated as: DM ¼ 1−MC

ð1Þ

Each sample (edible tissue) was thereafter lyophilized in order to prevent the loss of mercury and other possible volatile metallic compounds to facilitate sieving and make the sample ready for grinding. The obtained dry weights (DW) of lyophilized samples were then recorded for calculation of trace metal concentrations on a dry weight basis (μg g−1 DW), and thereafter on a wet weight basis (μg g−1 WW), using the following formulas, respectively: . DWConcx ¼ ðCxV Þ W ð2Þ WWConcx ¼ DWConcx xDM

ð3Þ

where Concx is the concentration of the analyzed metal (Cd, Cu, Hg, or Zn) in a dry weight (DW) or wet weight (WW) basis, C is the concentration of the extract (μg mL−1), V is the volume of the extract (mL), and W is the weight of the sample

Environ Sci Pollut Res Fig. 1 Location of the sampling areas of the two cephalopod species, Octopus vulgaris and Sepia officinalis, in the Gulf of Gabes

aliquot extracted (g). The concentrations of trace metals analyzed given here were reported as micrograms per gram on a wet weight basis (μg g−1 WW). The quality control of the analysis results was enabled using triplicate samples and blanks which were processed similarly to samples. Besides the standard reference materials, DOLT-2 and DORM-2 were used for the instrument calibration and for quality control. The quantitative recoveries of the four trace metals analyzed ranged between 77 and 120 %. The detection limits were 0.001, 0.003, 0.047, and 0.1 μg kg−1 for Cd, Cu, Zn (using ICPMS), and Hg (using direct mercury analyzer), respectively. Data analysis of trace metals concentrations The concentrations of trace metals analyzed in the Bmantle^ and Barms^ tissues of the two species were compared between the four sampling sites using Kruskal-Wallis (KW) test, due to the non-normality and/or non-homogeneity of the data relative to each of the cephalopods considered separately. Considering the pooled data of the two species, the conditions of normality and homogeneity of variables were satisfied (tested with Shapiro-Wilk test) and the comparison of the concentrations of trace metals analyzed between the two species was performed using one-way ANOVA. Besides, the principal component analysis (PCA) was conducted using the trace metals data obtained with each of the two cephalopod species (O. vulgaris or S. officinalis) and those obtained with both species (O. vulgaris + S. officinalis) and taking into consideration the Bsampling site^ factor (Elbibane Lagoon, Gabes, Kerkennah Island, and Chebba). In addition, the hierarchic classification dendrogram (based on Euclidean distance) was also conducted based on the data obtained with all trace metals and with both species, taking into consideration the Bsampling

site^ (Elbibane Lagoon, Gabes, Kerkennah Island, and Chebba) and Blocation^ (Southern GG, Central GG and Northern GG) factors. All statistical analyses were performed using MS Office Excel 2013, SPSS and Primer 6.0 software packages. Indices of human health risk assessment The human health risk due to the cephalopod consumption was evaluated by calculating the estimated daily intake (EDI), the target hazard quotient (THQ), and the hazard index (HI). These indices were used to evaluate the health risk posed to adult humans by the ingestion of trace metals analyzed (Cd, Cu, Hg, and Zn) via the consumption of both edible tissues (mantle + arms) of the two studied cephalopod species. &

Estimated Daily Intake (EDI): In the present study, EDI represents the daily intake of the trace metals analyzed (i.e., Cd, Cu, Hg, and Zn) through the consumption of the two cephalopod species. The EDI calculation was considered dependent on the amount of consumed cephalopods that can be ingested daily and its metals’ concentrations. According to Bortey-Sam et al. (2015), EDI of the four metals considered can be determined using the following equation: EDI ¼

MC FDC BW

ð4Þ

where MC is the average concentration of each trace metal in seafood (μg g−1 WW), FDC is the average food daily consumption of the two cephalopod species in Tunisia (g person−1 day−1) so that 33 g person−1 day−1 (FAO 2009) and BW is the average body weight. EDI was computed for an adult (so that 60 kg in BW).


&

Target Hazard Quotient (THQ): The THQ was developed in 1989 by the US Environment Protection Agency as a tool of human health risk assessment, associated with the consumption of contaminated products. The THQ can be calculated using the following equation: THQ ¼

&

EF ED MS C 10−3 RfDo BW At

ð5Þ

where EF is the exposure frequency (365 days year−1), ED is the exposure duration, so that 75 years according to NIS (2014) which is equivalent to the average life time in Tunisia, MS is the food meal size (33 g person−1 day−1 according to FAO (2009)), C is the average metal concentration in both edible parts (mantle and arms) of the cephalopod species (μg g−1 wet weight), RfDo is the oral reference dose (μg g−1 day−1), BW is the average adult body weight so that 60 kg, and At is the averaging exposure time for non-carcinogens, which is given by multiplying the exposure frequency (EF) by the exposure duration (ED). Hazard Index (HI): HI was developed to assess the total risk that may generate from the ingestion of the four trace metals analyzed in both mantle and arms tissues of the two cephalopod species. According to Jian et al. (2013), HI’s equation is the following: HI ¼ ∑nn¼1 THQi

ð6Þ

where HI is the total hazard index, THQi is the target hazard quotient of an individual trace metal, and n is the total number of trace metals considered (n = 4).

Results The average concentrations of trace metals analyzed in the mantle and arms of both species O. vulgaris and S. officinalis are given in Figs. 2 and 3. The spatial variations of trace metal concentrations were found to follow a similar pattern for both species and both tissues examined. With the mantle of O. vulgaris, average Cd metal concentrations oscillated between the minimal values of 0.03 ± 0.01 μg g−1 WW (in Elbibane lagoon) and 0.03 ± 0.01 μg g−1 WW (in Chebba) and the maximum of 0.14 ± 0.03 μg g−1 WW (in Gabes) (Fig. 2a). For Cu, average mantle concentrations of O. vulgaris varied from 3.11 ± 0.78 μg g−1 WW (in Gabes) to 6.16 ± 0.96 μg g−1 WW (in Elbibane lagoon) (Fig. 2b). The highest average mantle concentration of Hg varied between 0.03 ± 0.01 μg g−1 WW (in Chebba) and 0.05 ± 0.01 μg g−1 WW (in Gabes) (Fig. 2c). For Zn, the lowest and highest average contents estimated in the mantle of the Tunisian common octopus were found to be 8.40 ± 1.25 μg g−1 WW (in Elbibane lagoon) and 21.42 ± 2.02 μg g−1 WW (in Gabes), respectively (Fig. 2d).

With the arms of O. vulgaris, the lowest and highest concentrations of Cd were noted in Elbibane lagoon and Chebba (0.02 ± 0.01 μg g−1 WW) and Gabes (0.13 ± 0.04 μg g−1 WW), respectively (Fig. 2e). As for Cu, its average arms concentrations were found to vary between 2.45 ± 0.72 μg g−1 WW (in Gabes) and 5.04 ± 1.10 μg g−1 WW (in Elbibane lagoon) (Fig. 2f). Regarding Hg, the lowest average arms concentrations were recorded in Chebba (0.03 ± 0.01 μg g−1 WW) and Kerkennah Island (0.03 ± 0.01 μg g−1 WW); whereas the highest in Gabes (0.05 ± 0.01 μg g−1 WW) (Fig. 2g). With Zn, average concentrations were found to vary between 9.24 ± 1.78 μg g − 1 WW (in Elbibane lagoon) and 21.94 ± 0.01 μg g−1 WW (in Gabes) (Fig. 2h). As for the mantle of the cuttlefish S. officinalis, we found that the lowest average Cd concentrations were found to be 0.03 ± 0.01 μg g − 1 WW (in Elbibane lagoon) and 0.03 ± 0.01 μg g−1 WW (in Chebba); whereas the highest was estimated of 0.13 ± 0.02 μg g −1 WW (in Gabes) (Fig. 3a). As for Cu, average mantle concentrations, estimated with S. officinalis, were found to vary from a minimum of 3.28 ± 0.85 μg g−1 WW (in Gabes) to a maximum of 5.33 ± 1.51 μg g−1 WW (in Elbibane lagoon) (Fig. 3b). In the case of Hg, we found that mantle average concentrations varied from a minimum of 0.03 ± 0.01 μg g−1 WW recorded in both sites of Kerkennah Island and Chebba and a maximum of 0.05 ± 0.02 μg g−1 WW noted in Gabes (Fig. 3c). For Zn, while the lowest average concentration was found in Elbibane lagoon (8.43 ± 1.40 μg g−1 WW), the highest was recorded in Gabes (20.86 ± 2.14 μg g−1 WW) (Fig. 3d). The results of trace metals analyses in the arms of S. officinalis showed that the average Cd concentrations oscillated between a minimum of 0.02 ± 0.01 μg g−1 WW (in Chebba) and a maximum of 0.12 ± 0.02 μg g−1 WW (in Gabes) (Fig. 3e). As for Cu, lowest and highest average concentrations were found in Gabes (3.03 ± 0.87 μg g−1 WW) and Elbibane lagoon (4.82 ± 1.52 μg g−1 WW), respectively (Fig. 3f). Regarding Hg, the average concentration estimated in S. officinalis arms was almost constant (0.03 μg g−1 WW) in most of the sampling sites (i.e., Elbibane lagoon, Kerkennah Island, and Chebba). The Hg concentration noted in Gabes was the highest (0.05 ± 0.01 μg g−1 WW) (Fig. 3g). As for Zn, our results indicated that average concentrations varied between 7.14 ± 1.55 μg g−1 WW (in Elbibane lagoon) and 19.36 ± 0.02 μg g−1 WW (in Gabes) (Fig. 3h). The comparison of the concentrations of trace metals analyzed with each cephalopod species and with each muscle tissue between the four sampling sites, with the test of Kruskal-Wallis, indicated significant differences in all cases (Figs. 2 and 3). In contrast, the comparison of the contents of trace metals in mantle and arms tissues between the two species showed no significant differences with all cases (ANOVA, p > 0.5).


Fig. 2 Spatial variations of mean concentrations ± standard deviations of trace metals analyzed in the edible muscle tissues of the common octopus Octopus vulgaris in the Gulf of Gabes, south-eastern Tunisia. (a–d mantle

concentrations of Cd, Cu, Hg, and Zn, respectively; e–h arm concentrations of Cd, Cu, Hg, and Zn, respectively)

The principal components analyses (PCA) conducted using the trace metals concentrations in the edible tissues of the two cephalopod species, analyzed separately and pooled together showed a general segregation between the sampling sites (Fig. 4). Taking into account O. vulgaris, the principal component analysis conducted with the data of trace metals analyzed in both tissues showed, at an Euclidean distance of 15, the segregation of three clusters: one first cluster composed of all samples of Elbibane lagoon and the majority of those

collected from Chebba in addition to only two samples from Kerkennah Island; one second cluster formed by all samples of Chebba and Kerkennah Island and almost the half of Gabes samples and one third cluster consisting of all samples of Gabes and some of Kerkennah Island samples. At a distance of 5.9, all these latter clusters subdivided in different and overlapping sub-clusters highlighting a general segregation between the four sampling sites and/or their geographic locations in GG (Fig. 4a). Similar PCA results were found with


Fig. 3 Spatial variations of mean concentrations ± standard deviations of trace metals analyzed in the edible muscle tissues of the common cuttlefish Sepia officinalis in the Gulf of Gabes, south-eastern Tunisia.

(a–d mantle concentrations of Cd, Cu, Hg, and Zn, respectively; e–h arm concentrations of Cd, Cu, Hg, and Zn, respectively)

S. officinalis, showing a clear separation between two clusters, at level 19 of Euclidean distance: one first cluster formed by all samples of Elbibane Lagoon and few samples of Chebba and one second cluster which overlapped partially with the first one and it was then formed by few samples of Elbibane Lagoon and all the samples of Gabes, Kerkennah, and Chebba. At a distance of 13, the first cluster subdivided into two overlapped sub-clusters formed mainly by Elbibane Lagoon samples. As for the second cluster, it separated into two overlapping sub-clusters: one sub-cluster represented by

all the samples collected from Gabes, almost all samples of Kerkennah Island and around the half of Chebba samples and one second sub-cluster consisting of all Chebba samples and the majority of samples coming from Kerkennah Island (Fig. 4b). Considering the trace metals concentrations in the two studied muscle tissues of both cephalopod species, the PCA showed similar results to those obtained with the PCAs conducted with the two species separately. At a Euclidean distance of 18 to 25, the samples analyzed segregated into two main clusters: while the first one grouped all samples of

Environ Sci Pollut Res Fig. 4 Principal component analyses (PCA) of trace metals concentrations in the mantle and arms of Octopus vulgaris (a), Sepia officinalis (b), and both species (c), conducted based on the Bsampling site^ factor

Gabes, Kerkennah Island, and Chebba in addition to most of the samples of Elbibane Lagoon, the second one grouped all samples collected from this latter site (i.e., Elbibane Lagoon) with few samples from Chebba. At a distance level of 15, this latter second cluster showed the same grouping; whereas the first cluster was subdivided into two overlapping sub-clusters: one first sub-cluster represented by almost all samples from Gabes and Kerkennah and most of the samples of Chebba, and one second sub-cluster consisting of all Chebba samples and around the half of samples collected from Kerkennah Island and Elbibane Lagoon (Fig. 4c). In addition, the dendrogram of hierarchic classification conducted using all trace metals data relative to both mantle and arms tissues of the two cephalopod species highlighted a clear separation between two main groups: one group formed by only the sampling site located in southern GG (i.e., Elbibane Lagoon) and one second group formed by the central (i.e., Gabes) and northern (i.e., Kerkennah Island and Chebba) sampling sites of GG. At closer Euclidean distance, this latter group subdivided into two sub-groups: one first sub-group formed by only Gabes (central GG) and one second formed by the other two sites, located in

northern GG (i.e., Kerkennah Island and Chebba). These latter sites subdivided at closer Euclidean distance (Fig. 5). The results of estimated daily intake (EDI) and total hazard quotients (THQ) as well as of hazard index (HI) for each of the trace metals analyzed in each of the four sampling areas in the GG are given in Table 1. The highest O. vulgaris and S. officinalis EDIs of Cd (0.073 and 0.068 μg kg−1 BW day−1, respectively), Hg (0.026 μg kg −1 BW day −1 for both species) and Zn (11.924 and 11.063 μg kg−1 BW day−1, respectively) were recorded in Gabes (central GG). In the case of Cu, the highest EDIs of the two cephalopods (3.082 and 2.792 μg kg−1 BW day−1 for O. vulgaris and S. officinalis, respectively) were recorded for both species in Elbibane Lagoon (southern GG). The lowest EDIs of Hg and Cd were recorded for both species in Chebba (northern GG). For Zn and Cu, the lowest EDIs were noted for both cephalopod species in Elbibane Lagoon (southern GG) and Gabes (central GG), respectively. It is worth noting that all EDI values, estimated for each trace metal in each cephalopod species, were found to be lower than the provisional

Environ Sci Pollut Res Fig. 5 Dendrogram of hierarchic classification of sampling sites, conducted based on Euclidean distance and using the trace metals concentrations assessed in the mantle and arms of both cephalopod species studied (Octopus vulgaris and Sepia officinalis), with respect to location of these sampling sites in the Gulf of Gabes

tolerable daily intake (PTDI) for human adults (Table 1). The spatial variations of THQs were found to follow the same pattern than that of EDIs. In the case of Hg and Cd, while the highest THQs estimated with both O. vulgaris and S. officinalis were found in Gabes (central GG); the lowest records were noted in Chebba (northern GG). For Zn and Cu, the highest and lowest THQs were recorded for both species in Gabes (central GG) and Elbibane Lagoon (southern GG), respectively (Table 1). Regarding HI, the estimated values for all trace metals analyzed followed, for the two cephalopod species, the following descending order: Gabes > Kerkennah Island > Elbibane Lagoon > Chebba (Table 1).

Discussion This is the first study on the environmental and human risk assessment of trace metal pollution using cephalopods of the GG area. The findings on the trace metals distribution in the coastal area of GG are in agreement with previous studies carried out recently in the same region on the assessment of trace metals in gastropod and bivalve molluscs (Lahbib et al. 2013; Rabaoui et al. 2014) as well in marine surface sediments (Ayadi et al. 2014; Rabaoui et al. 2015; El Zrelli et al. 2015). It is worth noting that the higher Cu concentrations found in Elbibane Lagoon, compared to the other sites, may be due

Table 1 Average concentrations of trace metals analyzed in the edible tissue of Octopus vulgaris and Sepia officinalis, the corresponding estimated daily intake (EDI) and total hazard quotients (THQ) and their total (HI) in each of the sampling sites of Gabes gulf. The provisional tolerable daily intake (PTDI) for each of the trace metals analyzed is given at the bottom (last line) of the table Species

Sampling sites

Octopus vulgaris Elbibane Lagoon Gabes Kerkennah Island Chebba Sepia officinalis El Bibane Lagoon Gabes Kerkennah Island Chebba PTDIa

Average concentrations of trace metals in EDI both edible tissues (μg g−1 WW)

THQ

Cd

Cu

Hg

Zn

Cd

Cu

Hg

Zn

0.030 0.132 0.094 0.027 0.032 0.123 0.107 0.025

5.603 2.782 4.387 4.102 5.077 3.155 4.612 4.085

0.037 0.047 0.036 0.031 0.034 0.047 0.033 0.029

8.817 21.680 17.260 12.313 7.784 20.114 15.796 11.706

0.016 0.073 0.051 0.015 0.017 0.068 0.059 0.014 1.00

3.082 1.530 2.413 2.256 2.792 1.735 2.536 2.247 0.500

0.020 4.849 0.026 11.924 0.020 9.493 0.017 6.772 0.019 4.281 0.026 11.063 0.018 8.688 0.016 6.438 0.10 300.00

HI

Cd

Cu

Hg

Zn

0.016 0.073 0.051 0.015 0.017 0.068 0.059 0.014

0.006 0.003 0.005 0.005 0.006 0.003 0.005 0.004

0.201 0.257 0.197 0.168 0.188 0.261 0.179 0.159

0.016 0.040 0.032 0.023 0.014 0.037 0.029 0.021

0.240 0.373 0.285 0.209 0.225 0.368 0.272 0.199

RfD of Hg = 0.1 μg kg−1 day−1 , Cd = 1 μg kg−1 day−1 (USEPA 2000), and Zn = 0.3 mg kg−1 day−1 (USEPA 2005); The PTDI value for Cu was assessed based on the provisional maximum tolerable daily intake (PMTDI) established by the Joint FAO/WHO Expert Committee on Food Additives (JECFA): PMTDI of Cu = 0.5 mg kg−1 day−1 (JECFA 1982) a

PTDI values for Hg, Cd, and Zn were estimated based on the oral reference dose data (RfD) from the United States Environmental Protection Agency (2000, 2005)


to the treatment effect of the surrounding olive groves with special fungicide (Bordeaux mixture) formed mainly by a copper sulfate mixture (CuSO4) and slaked lime (Ca(OH)2), leading most likely to the increase of Cu contents in the soil and the lagoon. The main results, obtained with the two cephalopod species studied, highlighted that the central area of GG (i.e., Gabes) is the one hosting the highest concentrations of trace metals, in particular Cd, Hg, and Zn. The lowest concentrations of these latter trace metals as well in the mantle as in the arms of both species were recorded in the northernmost (i.e., Chebba) and southernmost (i.e., Elbibane Lagoon) sampling sites of GG. In the case of Cu, the highest contents were recorded (with both muscle tissues and both cephalopod species) in Elbibane Lagoon, followed by Kerkennah Island, and the lowest contents in Gabes. Similar pattern of trace metal distribution in the marine environment of GG (especially Cd, Hg, and Zn) was also reported by Rabaoui et al. (2014, 2015) and El Zrelli et al. (2015). Several factors represented mainly by coastal anthropogenic activities were considered responsible of the high enrichment of the central area of GG with trace metals, in particular, the industrial wastes of the Gabes-Ghannouch industrial complex, including phosphogypsums (El Zrelli et al. 2015; Rabaoui et al. 2015), which are mostly released Buntreated^ directly in the open sea (Kharroubi et al. 2012). In fact, phosphogypsum of the industrial complex in Gabes was reported to be composed of various pollutants including metals (Soussi et al. 1995; Zaghden et al. 2005). Based on a recent study on metallic pollution in the GG, it was reported that Cd and Zn may derive mainly from phosphogypsum of Gabes industrial complex (El Zrelli et al. 2015). Several authors reported the environmental consequences of the coastal industrialization in the coastal area of GG, in particular, the impact of phosphogypsum wastes which affects different levels of local marine ecosystems, from organisms (Lahbib et al. 2013; Messaoudi et al. 2009) to communities (Darmoul and Vitiello 1980; Aloulou et al. 2012; Rabaoui et al. 2015) and even to key habitats like Posidonia oceanica meadows (Darmoul et al. 1980; Darmoul 1988; Zaouali 1993). The non-significant differences obtained between the concentrations of trace elements in O. vulgaris and S. officinalis may be explained by the benthic-living behavior of the two species. The two species have almost the same depth range (up to 200 m) and same migratory behavior, occurring mainly in shallow coastal waters in spring/summer and in deep waters in autumn/winter (FAO species catalogue 1984), and may then project similar assessment of metallic pollution of the environment where they live. Within this context, it is worth noting that benthic cephalopods were reported to accumulate more Cd, Cu, and Zn than pelagic ones, due to the differences

in physiological structures and feeding behavior (Bustamante et al. 2002). Similar results were also reported in Portugal (Lourenço et al. 2009) and Tunisia (Rjeibi et al. 2014). A comparison of the concentrations of trace metals in the muscle tissues of the two cephalopods species with the results of other studies carried out in Tunisian and other Mediterranean and Atlantic areas is given in Table 2. Considering the concentrations of trace metals analyzed in the mantle of O. vulgaris, the mean concentrations ranges of most of trace metals considered (Cd, Hg, and Zn) were more or less quite similar to the records of Rjeibi et al. (2015). The highest mean concentrations of Hg, Zn, and Cd recorded in the GG exceeded those noted in Bizerte and Monastir, located at northern and eastern Tunisian coasts, respectively (Rjeibi et al. 2015), but they were almost equal or lower than those recorded by these latter authors in Sfax, located in the northern part of GG. As for Cu, the records obtained herein were found to not vary significantly from those recorded in Monastir, located at the eastern coast of Tunisia (Rjeibi et al. 2015), but they were low compared to the concentrations reported in Bizerte (northern Tunisian coast) and Sfax (northern part of GG) by Rjeibi et al. (2015) (Table 2). The comparison of the trace metals concentrations in the arms of O. vulgaris showed also some geographical variations. The highest mean concentrations of Zn and Cd found in the arms of GG common octopuses are evidently higher that the ones reported by Rjeibi et al. (2015) in Bizerte and Monastir, but lower than those recorded in Sfax by the same latter authors. For the highest mean concentration of Hg, it was found higher than those found in Monastir and Sfax, but lower than that noted in Bizerte by Rjeibi et al. (2015). As for Cu, the highest mean concentration noted in the arms of GG octopus was found to be lower than that reported recently in Monastir (Rjeibi et al. 2015) but slightly high compared to those noted by the same latter authors in Bizerte and Sfax (Table 2). The trace metals mean concentrations reported with O. vulgaris arms and mantle, by other authors in other Mediterranean (Nesim and Riad 2003) and Atlantic (Lourenço et al. 2009) areas were found to be in the same range of Tunisian records (present study; Rjeibi et al. 2014), except for Hg which was found to be very high in Portugal (Lourenço et al. 2009). With S. officinalis, the mean concentrations of Hg analyzed in the mantle and arms during this study followed a similar pattern to that of O. vulgaris. They were almost similar to those noted in Bizerte, but lower than those reported in Monastir (Rjeibi et al. 2015) and Sfax (Rjeibi et al. 2015; Mezghani-Chaari et al. 2011). In the case of Zn, the lowest and highest mean concentrations recorded herein were, in general, higher than those noted by the same latter authors along the northern and eastern Tunisian coasts. Regarding Cd, the mean mantle concentration ranges of Tunisian GG common cuttlefishes were almost similar to those recorded in Monastir (Rjeibi et al. 2015), lower than those of Bizerte

0.028 ± 0.012–0.033 ± 0.015 0.018 ± 0.011–0.050 ± 0.013 0.062 ± 0.024–0.173 ± 0.031 0.02 ± 0.01–0.13 ± 0.04

0.016 ± 0.008–0.023 ± 0.014 0.004 ± 0.002–0.040 ± 0.012 0.047 ± 0.019–0.154 ± 0.032 1.2–1.9

0.38 ± 0.39

Mantle Mantle Mantle Arms

Arms Arms Arms Mantle + arms

Mantle + arms

b

a

0.02 ± 0.01–0.12 ± 0.02 0.020 ± 0.007–0.051 ± 0.025 0.017 ± 0.006–0.053 ± 0.019 0.018 ± 0.007–0.052 ± 0.008 0.31 ± 0.28

Arms Arms Arms Arms Mantle + arms

Mantle + arms

0.03 ± 0.01–0.13 ± 0.02 0.024 ± 0.015–0.10 ± 0.037 0.031 ± 0.017–0.124 ± 0.036 0.023 ± 0.012–.062 ± 0.016 1.73 ± 0.57–2.39 ± 0.37

Mantle Mantle Mantle Mantle Mantle 3.03 ± 0.02–4.82 ± 1.52 4.36 ± 1.55–7.21 ± 1.71 4.84 ± 1.54–6.86 ± 2.41 4.14 ± 1.03–5.83 ± 1.12 4.5 ± 2.5a

3.28 ± 0.85–5.33 ± 1.51 2.12 ± 1.20–9.59 ± 4.95 2.70 ± 1.41–6.40 ± 2.58 2.93 ± 0.65–6.16 ± 1.92 11.4 ± 19–12.3 ± 18

3.8 ± 1.6

3.00 ± 0.78–4.27 ± 2.42 2.09 ± 0.72–6.02 ± 2.89 2.58 ± 0.67–4.36 ± 1.32 5.3–12.6

5.68 ± 1.26–8.64 ± 4.03 3.01 ± 1.71–5.18 ± 1.39 4.30 ± 1.92–6.51 ± 1.17 2.45 ± 0.72–5.04 ± 1.10

3.11 ± 0.78–6.16 ± 0.96

Cu

0.43 ± 0.12–0.55 ± 0.23

0.03 ± 0.01–0.05 ± 0.01 0.033 ± 0.007–0.051 ± 0.064 0.051 ± 0.011–0.083 ± 0.029 0.035 ± 0.011–0.076 ± 0.010 0.15 ± 0.10

0.03 ± 0.01–0.05 ± 0.01 0.035 ± 0.014–0.045 ± 0.012 0.058 ± 0.011–0.085 ± 0.029 0.041 ± 0.016–0.091 ± 0.014

0.13 ± 0.06

0.020 ± 0.009–0.060 ± 0.035 0.024 ± 0.004–0.033 ± 0.011 0.022 ± 0.003–0.039 ± 0.012

0.023 ± 0.010–0.045 ± 0.021 0.029 ± 0.005–0.039 ± 0.014 0.025 ± 0.003–0.051 ± 0.006 0.03 ± 0.01–0.05 ± 0.01

0.03 ± 0.01–0.05 ± 0.01

Hg

Mean concentrations ranges defined with two genders of S. officinalis

Mean concentrations ranges defined based on a seasonal assessment of trace metals in cephalopods

Sepia officinalis

0.03 ± 0.01–0.14 ± 0.03

Mantle

Octopus vulgaris

Cd

Trace metals

Tissues

Cephalopod species

7.14 ± 1.55–19.36 ± 1.67 10.78 ± 5.64–21.77 ± 2.76 12.04 ± 10.48–18.98 ± 2.34 11.90 ± 9.23–18.55 ± 4.15 17.7 ± 2.3

8.43 ± 1.40–20.86 ± 2.14 10.60 ± 2.13–18.34 ± 6.20 9.22 ± 0.98–11.70 ± 1.37 9.35 ± 0.74–18.55 ± 6.80 7.60 ± 0.79–8.18 ± 1.05

17.7 ± 2.2

11.60 ± 2.76–14.46 ± 0.59 13.30 ± 1.79–17.27 ± 5.21 12.83 ± 1.40–27.19 ± 12.42 13.3–20.0

6.82 ± 1.47–12.07 ± 1.05 10.48 ± 2.46–13.88 ± 6.07 13.38 ± 0.68–23.98 ± 9.87 9.24 ± 1.78–21.94 ± 2.40

8.40 ± 1.25–21.42 ± 2.02

Zn

Gulf of Gabes (Tunisia) Bizerte (Northern Tunisia) Monastir (Eastern Tunisia) Sfax (Eastern Tunisia) Iskenderun Bay (Southern Coast of Turkey) Gulf of Gabes (Tunisia) Bizerte (Northern Tunisia) Monastir (Eastern Tunisia) Sfax (Eastern Tunisia) Portugal (Atlantic Northeastern European coast) Sfax (Eastern Tunisia)

Portugal

Gulf of Gabes (South-eastern Tunisia) Bizerte (Northern Tunisia) Monastir (Eastern Tunisia) Sfax (Eastern Tunisia) Gulf of Gabes (South-eastern Tunisia) Bizerte (Northern Tunisia) Monastir (Eastern Tunisia) Sfax (Eastern Tunisia) Alexandria (Egypt)

Study areas (Location)

Rjeibi et al. (2015)a Rjeibi et al. (2015)a Rjeibi et al. (2015)a Nessim and Riad (2003) Lourenço et al. (2009) Present study Rjeibi et al. (2015)a Rjeibi et al. (2015)a Rjeibi et al. (2015)a Duysak et al. (2013)b Present study Rjeibi et al. (2015)a Rjeibi et al. (2015)a Rjeibi et al. (2015)a Lourenço et al. (2009) Mezghani-Chaari et al. (2011)

Rjeibi et al. (2015)a Rjeibi et al. (2015)a Rjeibi et al. (2015)a Present study

Present study

References

Table 2 Mean concentrations range or mean concentrations ± standard deviation per sampling sites of trace metals (μg g−1 WW) in muscle tissues (mantle, arms, mantle + arms) of Octopus vulgaris and Sepia officinalis in the Mediterranean and Atlantic areas



and Sfax (Rjeibi et al. 2015) and higher than that reported in Iskenderun Bay, Turkey (Duysak et al. 2013). In the case of S. officinalis arms, the highest Zn mean concentration recorded herein were, in general, higher than those reported in other Tunisian areas (Rjeibi et al. 2015). For Cu, the highest mantle and arms concentrations found in GG were, in general, low compared to those recorded in northern and eastern Tunisian coasts (Rjeibi et al. 2015) and in Iskenderun Bay, Turkey (Duysak et al. 2013) (Table 2). In addition, it seems from these results that the concentrations of trace metals in the two cephalopod species vary with respect to geographic locations and elements considered. This is mainly due to the variability of trace metals enrichment sources from one area to another and to the fact that some trace metals may originate from different anthropogenic releases. Based on our study, the highest concentrations of trace metals were, in general, found in Gabes, located in the central area of GG, confirming the findings of previous studies which lead to deduce that there are many sources of enrichment of the marine environment with trace metals (Lahbib et al. 2013; El Zrelli et al. 2015; Rabaoui et al. 2014, 2015). Moreover, the studies of Rjeibi et al. (2014, 2015), conducted on trace metal bioaccumulation in three cephalopod species including O. vulgaris and S. officinalis highlighted that the highest concentrations were recorded in Sfax, located in the northern part of GG. Sfax city is among the biggest cities of the GG hosting one of the well-developed coastal industrial and urban zones in the area. All these anthropogenic developments may exert various pressures on the environment including the enrichment with trace metals. In our case, the BKerkennah Island^ sampling site was not found to host high concentrations of trace metals analyzed. Rabaoui et al. (2015) found that in spite of the proximity of Kerkennah Island to the big city of Sfax, it does not seem to be reached by the organic/urban and industrial wastes of Sfax industrial zone explaining that by the effects of the north-south marine currents which may transfer the pollutants released (from Sfax city) in the open sea to the south and by the fact that this latter Island does not host a well-developed urban area. Considering the samples collected from Gabes (central GG) which is considered as the most polluted area in GG, none of the trace metals found in the edible tissues of both cephalopod species was found to be worrying and/or exceeding the standard values fixed by international organisms. This is different to the findings of some other previous studies carried out in the same area using gastropod and bivalve molluscs (Lahbib et al. 2013; Rabaoui et al. 2014) and fishes (Mezghani-Chaari et al. 2011). This may be explained by the very efficient detoxification processes that cephalopods have developed, especially their capacity to store and sequester many trace metals in their digestive gland. This has been already reported as well for Cd and Zn (Bustamante et al. 2002, 2006; Koyama et al. 2000) as well as for Hg (Bustamante et al. 2006). Within this context, Bustamante

et al. (2002) demonstrated that the bioaccumulation of Cd and Zn in S. officinalis is primarily due to food ingestion and that the digestive gland of this cuttlefish is responsible on the subsequent storage and presumed detoxification of these latter metals (Bustamante et al. 2002). This can be also due to the motile and migratory lifestyle of cephalopod species which do not seem to occur in a same area for long periods. Besides, we cannot be sure about the exact area from which Gabes samples were collected because the central coastal area of GG was reported to exhibit different levels of metallic pollution based on the location/proximity of sampling sites to Ghannouch-Gabes industrial complex (El Zrelli et al. 2015). Moreover, cephalopod species including O. vulgaris and S. officinalis were reported to accumulate more metals in their digestive glands (Miramand and Guary 1980; Bustamante et al. 2008; Rjeibi et al. 2014). Based on the results of human health risk assessment, the values of EDIs, THQs, and HIs found herein are, in general, not alarming and do not reveal risks for local consumers. However, we think that at a long-term continuous consumption of these cephalopod species, toxicity risk can take place especially for inhabitants of the central area of GG (Gabes inhabitants, mainly fishermen), since the highest health risk generated by the consumption of O. vulgaris and S. officinalis was found in Gabes, followed by Kerkennah Island, Elbibane Lagoon, and then Chebba (Table 1). This can be accentuated through the biomagnification of Hg or other trace metals not considered herein, across the marine web chain in the central area of GG leading probably to the human health risk aggravation of local consumers, in particular those living along the coast of Gabes city (i.e., fishermen Fig. 6). Within this context, it is worth noting that the mean HI values estimated for each cephalopod species considering the four trace metals analyzed showed higher records with O. vulgaris (mean HI = 0.277 ± 0.071) compared to S. officinalis (mean HI = 0.266 ± 0.074), although the difference between the two values was not found to be significant (ANOVA: F = 0.043; p = 0.842). Moreover, we remind here that the present study was carried out considering only four trace metals (Cd, Cu, Hg, and Zn) and that other metals including lead (Pb), chromium (Cr), arsenic (As), fluorine (F), and radionuclides, which may be hazardous and threatening the human health, were reported to enrich the marine environment of GG mainly through the phosphogypsum marine discharge (El Zrelli et al. 2015). According to Jović and Stancković (2014), these latter pollutants can synergistically pose a serious threat to human health and be toxic even at low doses. On the other hand, it is worth noting that apart from cephalopods, other marine species are also consumed by locals including bivalve and gastropod molluscs as well as fishes. Previous studies on the trace metals bioaccumulation of some of these latter taxa collected from the central areas of GG indicated higher concentrations that exceed the standard levels fixed by

Environ Sci Pollut Res Fig. 6 Diagram representing the bioaccumulation and biomagnification of trace metals in the trophic chain of the coastal ecosystem in the Gulf of Gabes

international organizations like WHO, FAO, USEPA, and EU (Mezghani-Chaari et al. 2011; Lahbib et al. 2013; Rabaoui et al. 2014). Local inhabitants, particularly fishermen, usually consume various seafood products, which may lead to some negative consequences on their health, in particular in case of high concentrations of trace metals. Within this context, it is worth noting that high rates of numerous diseases (including fetuses malformations, abortion, infertility, impotence, cardiovascular diseases, various cancers, and many other chronic diseases) were highlighted by doctors (through local media) in the families living in the coasts of Gabes city, compared to the rates observed in other Tunisian areas. The consumption of contaminated marine products can be one of the responsible factors explaining these high rates of diseases observed in the local population, as is the case of autism signs observed in a four-year-old child in relation with the consumption of some mercury-contaminated marine products from Sfax coasts (Mezghani-Chaari et al. 2011). According to a recent report, the pollution caused by the chemicals sector including mining and phosphate processing, in some Tunisian coastal cities mainly Gabes and Sfax, is responsible for the increase in cardiovascular diseases, respiratory infections, and the appearance of new types of cancer (EEA 2014). Summarizing, the present study is one of the preliminary contributions to the knowledge of environmental health status and human health risk assessment in the area of GG using two of the most edible cephalopod species. While the central area of GG (Gabes) showed the highest concentrations of trace metals analyzed and the highest EDIs, THQs, and HIs, the results found are, in general, not worrying; however, given the motile and migratory lifestyle of O. vulgaris and S. officinalis and because this

study did not consider other metals, we think that the long-term consumption of GG cephalopods in addition to other marine products may be responsible for some health risks on local consumers. Deeper studies considering the bioaccumulation of other trace metals are needed to make a better and complete environmental and health risk assessment of the GG area. Acknowledgements The authors would like to express their sincere appreciation to the Deanship of Scientific Research at King Saud University for funding the research group no. RG-1435-023. We are also grateful to the three referees who helped to improve the quality of the manuscript through their constructive comments and suggestions and to all those who helped in collecting samples and laboratory analyses.

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