Organohalogen compounds in blubber of Indo-Pacific bottlenose dolphin (Tursiops aduncus) and spinner dolphin (Stenella longirostris) from Zanzibar, Tanzania

Environmental Pollution 158 (2010) 2200e2207

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Organohalogen compounds in blubber of Indo-Pacific bottlenose dolphin (Tursiops aduncus) and spinner dolphin (Stenella longirostris) from Zanzibar, Tanzania Haji Mwevura a, *, Omar A. Amir b, c, Michael Kishimba a, Per Berggren c, f, Henrik Kylin d, e a

Department of Chemistry, University of Dar es Salaam, Dar es Salaam, Tanzania Institute of Marine Sciences, University of Dar es Salaam, P O Box 668, Zanzibar, Tanzania Department of Zoology, Stockholm University, SE-106 91 Stockholm, Sweden d Department of Aquatic Sciences and Assessment, Swedish University of Agricultural Sciences, P O Box 7050, SE-750 07 Uppsala, Sweden e Norwegian Institute of Air Research, Polar Environmental Centre, NO-9296 Tromsø, Norway f School of Marine Science and Technology, University of Newcastle upon Tyne, NE1 7RU, United Kingdom b c

Biogenic brominated organic compounds were found at higher concentrations than anthropogenic organochlorine pesticides in dolphins off Zanzibar.

a r t i c l e i n f o

a b s t r a c t

Article history: Received 30 December 2009 Received in revised form 19 February 2010 Accepted 25 February 2010

Blubber samples of Indo-Pacific bottlenose (Tursiops aduncus) and spinner (Stenella longirostris) dolphins from Zanzibar, East Africa, were analyzed for a wide range of organohalogen compounds. Methoxylated polybrominated diphenyl ethers (MeO-BDEs), presumably biogenic, were found at higher concentrations than anthropogenic organochlorine pesticides (OCPs). Only traces of industrial pollutants, such as polychlorinated biphenyls, were detected. The OCP levels found off Zanzibar were lower than those reported from other regions while MeO-BDE levels were higher. The relative composition of the OCPs indicated recent use of lindane (g-hexachlorocyclohexane) and aged residues of DDT and technical HCH. Placental transfer was estimated to 2.5% and 0.5% of the total burden of OCPs and MeO-BDEs, respectively. Overall transfer from mother to calf in Indo-Pacific bottlenose dolphins was estimated to 72% and 85% for the OCPs and MeO-BDEs burdens, respectively. Health effects of MeO-BDEs are not known, but structural similarities with well-known environmental toxins are cause for concern. Ó 2010 Elsevier Ltd. All rights reserved.

Keywords: Methoxylated polybrominated diphenyl ethers MeO-BDEs Organochlorine pesticides Maternal transfer

1. Introduction The coastal waters off Tanzania have a diverse fauna with many species of fish and marine mammals. Around Unguja Island (the main island of Zanzibar) seven species of dolphins occur regularly, of which Indo-Pacific bottlenose dolphin (Tursiops aduncus) and spinner dolphin (Stenella longirostris) are the two most common (Amir et al., 2005a). The near-shore distribution of these species make them particularly vulnerable to human activities such as hunting, incidental capture in fisheries and habitat degradation from anthropogenic organic contaminants (Amir et al., 2005a). Persistent organic pollutants (POPs) are among the most wellknown environmental contaminants and many are regulated * Corresponding author. Present address: State University of Zanzibar, P O Box 146, Zanzibar, Tanzania. E-mail addresses: [email protected], [email protected] (H. Mwevura), [email protected] (O.A. Amir), [email protected] (M. Kishimba), [email protected] (P. Berggren), [email protected]. se (H. Kylin). 0269-7491/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.envpol.2010.02.027

internationally within the “Stockholm Convention” (www.pops.int). Typical legacy POPs that have been studied for many years are, e.g., organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCB). Among POPs that have gained scientific interest more recently are, e.g., brominated flame retardants such as polybrominated diphenyl ethers (PBDE). These compounds are hydrophobic, most are halogenated, and, because of their refractory nature, they accumulate in lipid-rich tissues. Dolphins have lipid-rich blubber and feed at a high trophic level, which makes them prone to accumulate significant amounts of POPs (Boon et al.,1994). The likelihood that dolphins accumulate hydrophobic persistent compounds makes them ideal to monitor organohalogen compounds in the marine environment. Although structurally similar to metabolites of anthropogenic PBDEs, methoxylated polybrominated diphenyl ethers (MeO-BDEs) found in biota are of biogenic origin (Teuten and Reddy, 2007). MeO-BDEs have been found as natural products in marine sponges (Agrawal and Bowden, 2005), and in algae (Malmvärn et al., 2005). Although little is known about the biological role of these compounds, researchers are concerned about possible health

H. Mwevura et al. / Environmental Pollution 158 (2010) 2200e2207

effects at high trophic levels, pointing out that their structural similarity with PCBs may result in similar environmental problems (McDonald, 2002). The build-up of high concentrations of anthropogenic POPs in marine mammals is the result of biomagnifications from the invertebrates and fish they consume (Gray, 2002). The transfer of natural organohalogens from the identified sources to higher trophic levels is not yet clearly understood, but higher concentrations of MeO-BDEs in carnivores (dolphins and whales) than in herbivores (dugongs and green turtles) show that the accumulation is associated with biomagnifications through the food chain (Vetter et al., 2001b). Information on organohalogens in the Tanzanian environment is scarce (Kishimba et al., 2004). To our knowledge, no previous study has assessed the status of organohalogens at high trophic levels of the marine ecosystem. Such an assessment was made possible by a programme to archive samples from dolphins stranded or trapped and killed as bycatch during fishing activities. For archiving and research purposes animals killed as bycatch is better than dead stranded animals; the samples are fresh and the cause of death is known. In this study, the accumulation and

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composition of organohalogen compounds in blubber of IndoPacific bottlenose (TA) and spinner (SL) dolphins from the coastal waters off Zanzibar were determined. Sex and maturity-related differences were investigated within species, and maternal transfer of the quantified organohalogens was estimated. 2. Materials and methods 2.1. Sampling area Unguja (Zanzibar Island, Fig. 1) is located on the continental shelf, separated from mainland Tanzania by the Zanzibar Channel. Rivers emptying into the waters of the Zanzibar Channel are either small or ephemeral, why there is little runoff carrying pollutants into the waters. However, the area is densely populated (URT, 2003), with some 0.7 million inhabitants on Unguja (0.2 million in Zanzibar Town), and >3 million inhabitants on the mainland, including two major cities Dar es Salaam (2.5 million), and Tanga (0.2 million). There is little heavy industry in the area, but both Unguja and the coastal plain are under intense agriculture (Mmochi and Mberek, 1998). Water movements in the Zanzibar Channel are complex and affected by seasonally changing monsoons. Along the coast runs the Mozambique/Aghulas Current southwards. The “winter monsoon” which blows from the northeast (NovembereMarch) creates a surface current from the Arabian Sea, while the

Fig. 1. Map of Zanzibar showing approximate sampling locations for T. aduncus (closed circles) and S. longirostris (open circles).

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“summer monsoon” blowing from the southeast (AprileOctober) drives surface waters in a clockwise current in the southern Indian Ocean bringing water from the open ocean to Zanzibar (Newell, 1959; Shaghude and Wannäs, 1998). The majority of dolphins sampled in this study were caught as bycatch on the shelf west of Unguja between Unguja and the mainland, but a few also on the shelf off the east coast of Unguja (Fig. 1). 2.2. Sample collection Blubber samples from 18 Indo-Pacific bottlenose dolphins and 18 spinner dolphins were collected from dolphins trapped and killed in gillnets as bycatch during the period 2000e2002, approximate locations are shown in Fig. 1. Biological and physical data were collected during dissections of the dead dolphins performed at the Institute of Marine Sciences (IMS), Zanzibar (Table 1). Prior to dissection, the total body length was measured to the nearest centimetre in a straight line from the tip of the upper jaw to the fluke notch and the mass was recorded to the nearest kilogram. The sex of the specimens was determined externally and confirmed by examination of gonads. Blubber samples were taken from the left side in front of the dorsal fin, wrapped in aluminium foil, put into polythene bags and frozen at
20  C until analysis. Since there was no targeted sampling, sample consistency in species, age, sex and location could not be maintained. 2.3. Determination of maturity The sexual maturity of females was determined from evidence of ovulation, pregnancy and lactation (Amir, 2010). In males, sexual maturity was indicated by the presence of sperm in the testicular tissue as determined by histological investigations (Amir, 2010).

2.4. Sample preparation Blubber samples were defrosted and cut into pieces and a subsample (5 g) was extracted. The blubber was ground with anhydrous sodium sulphate and extracted successively by shaking for 10 min with 50 þ 20 þ 20 þ 20 ml of dichloromethane using an overhead shaker. The extracts were filtered through anhydrous sodium sulphate and the solvent removed in a rotary evaporator. After extraction, two lipid subsamples (0.5 g) were dissolved in cyclohexane (4 ml), the internal standard hexabromobenzene (HBB) was added to each, after which one was cleaned up by sulphuric acid for the determination of acid stable compounds and the other with potassium hydroxide for determination of compounds, e.g., the dieldrin and aldrin, that are not stable to treatment with concentrated sulphuric acid (Åkerblom, 1995).

2.5. Instrumental analysis OCPs were quantified with gas chromatography (GC) on an Agilent 6890 (Agilent, Wilmington, Delaware, USA) equipped with two electron capture detectors. Separation was performed simultaneously on two capillary columns of different polarity (CP-Sil 5CB and CP-Sil 19CB, 30 m  0.32 mm  25 mm, Chrompak, Middelburg, The Netherlands) attached to the same split-splitless injector, but with separate detectors. The column temperature program employed was 90  C (equilibrium time 1 min), rising to 180  C at 30  C/min, then to 260  C at 4  C/min and held for 10 min. The injector and detector were held at 250  C and 300  C, respectively. The analytes were identified and quantified by comparisons of the retention times and peak heights of the analytes and authentic reference standards relative to HBB on the two columns. Identity was considered confirmed if the relative retention times were correct on both columns, and quantification was done by using the value from whichever column gave the lowest quantity. Linearity was

Table 1 Summary data on sample characteristics and the concentrations (ng/g lipid) of different organohalogen groups in individual samples of Indo-Pacific bottlenose (TA) and spinner (SL) dolphins. Data on each individual compound is given in the electronic supplement, Table E1. Sample Indo-Pacific TA 04 TA 12 TA 18 TA 29 TA 10 TA 24 TA 28 TA 26 m TA 26 f TA 16 TA 17 TA 19 TA 21 TA 22 TA 13 TA 27 TA 08 TA 20

Location

Sex

Maturity status

bottlenose dolphin (Tursiops aduncus) \ Imm U. Membea Pungume Is. \ Imm M. Mwanab \ Imm Nungwi \ Imm Kitami \ Mat Nungwi \ Mat Changuu Is \ Mat Nungwi \ Mat Nungwi _ Foetus U. Membe _ Imm M. Mwana _ Imm U. Membe _ Imm Tanga _ Imm Uroa _ Imm U. Membe _ Mat Tanga _ Mat Nungwi _ Mat Nungwi _ Mat

Spinner dolphin (Stenella longirostris) SL 06 Tanga \ Imm SL 12 Mnemba Is. \ Imm SL 25 U. Membe \ Imm SL 22 U. Membe \ Mat SL 17 M. Mwana \ Mat SL 26 Nungwi \ Mat SL 16 Nungwi \ Mat SL 15 Nungwi _ Imm SL 23 U. Membe _ Mat SL 24 U. Membe _ Mat SL 14 Nungwi _ Mat SL 18 Nungwi _ Mat SL 19 Nungwi _ Mat SL 08 Nungwi _ Mat SL 27 Nungwi _ Mat SL 28 Mnemba Is. _ Mat SL 29 Mnemba Is. _ Mat SL 30 Mnemba Is. _ Mat *ND ¼ not detected. a Uso wa Membe. b Mwana wa Mwana.

Weight (kg)

Length (cm)

Lipid (%)

HCB (ng/g)

SHCH (ng/g)

SCyclodienes (ng/g)

Methoxychlor (ng/g)

SDDT (ng/g)

SMeO-BDE (ng/g)

165 31 57 32 122 110 120 149 16 29 60 24 38 25 112 120 118 158

194 136 149 139 221 219 219 221 104 130 154 126 149 122 205 202 213 225

39.9 35.6 32.1 37.8 39 34.6 39.4 46.4 29.7 31 34.9 36.6 38.5 30.6 48 42.8 38.6 44.7

40 76 28 380 180 160 94 71 6 58 76 39 83 52 69 41 85 61

160 310 36 200 200 130 150 200 34 220 160 140 87 180 110 180 150 170

210 450 290 970 220 400 260 150 20 200 280 290 230 94 590 160 260 520

360 330 240 350 170 ND* ND 120 ND 130 500 380 190 110 580 150 170 150

17 000 8000 7100 46 000 13 000 20 000 10 000 2800 570 2700 24 000 29 000 1000 500 93 000 10 000 29 000 38 000

33 15 72 160 21 29 22 18

20 14 13 61 66 62 67 39 63 72 72 73 55 77 58 69 67 70

125 105 101 189 187 175 189 150 180 184 191 185 176 207 177 183 175 174

40.8 28.7 34.1 34.2 37.6 31.5 36.8 29.6 32.6 43.4 33.2 39.7 33.8 43.5 33.5 35.3 31.7 36

52 20 34 26 19 41 31 39 59 47 69 53 39 85 36 81 57 35

87 79 62 100 89 200 77 110 160 160 220 140 120 160 130 130 96 93

300 180 150 340 240 290 99 65 330 420 370 500 350 750 220 300 480 330

310 ND 160 ND 320 930 120 78 240 410 310 230 350 720 180 200 270 170

8900 5500 3800 5800 1700 7800 7700 3800 11 000 23 000 11 000 19 000 8600 76 000 8900 8900 11 000 9400

61 000 67 000 10 000 31 000 32 000 6800 51 000 52 000 55 000 210 000 72 000 99 000 100 000 190 000 94 000 110 000 44 000 42 000

000 000 000 000 000 000 000 000 600 37 000 65 000 25 000 5800 8300 190 000 85 000 150 000 180 000

H. Mwevura et al. / Environmental Pollution 158 (2010) 2200e2207 tested with a five point calibration curve. For quantification of analytes with concentrations too high to be within linearity, the extracts were diluted with addition of a known quantity of HBB before reinjection on the GC. Limits of detection and recoveries are given in Table E1 in the electronic supplement; limit of quantification was assigned as five times the limit of detection. Brominated compounds were identified by gas chromatography-mass spectrometry (GCeMS) with electron capture negative ion (ECNI) ionization in the full scan mode. Mass spectra and retention times were compared with the spectra of MeO-BDEs reference standards provided by Åke Bergman, Environmental Chemistry, Stockholm University. After identification, quantifications were performed with GC-ECD. To accommodate quantification of the very high concentrations the samples had to be diluted to be within the linear range of the detector response. The dilution was done with a known amount of HBB added to the diluting solvent. 2.6. Data analysis Statistical analyses were performed using the STATISTICA 8 (StatSoft Inc., Tulsa, OK, USA), with statistical significance for all tests set at p  0.05. Levene's equality of variances F-test was used to determine the homogeneity of variances. Where necessary, the data were log-transformed to increase the homogeneity of variance. Student's t-test and KruskaleWallis Anova test were used to examine the difference between means by gender and maturity classes within species. Spearman correlation (r) was used to examine the relationships between accumulation of OCPs and MeO-BDEs with age of both sexes. Due to the lack of data on developmental stage for spinner dolphins, we used length as a proxy for immature individuals as length increases with age until the asymptotic length is reached. 2.7. Estimation of maternal transfer Maternal transfer to offspring in Indo-Pacific bottlenose dolphins was estimated as described by Tanabe et al. (1982). The blubber mass of the mother/foetus pair, mature male and female dolphins was calculated as described in McLellan et al. (2002). Briefly, the whole blubber was stripped from the animals and weighed to calculate the % contribution of blubber to total body mass. The blubber mass was calculated as 23.9% for the foetus and 13.1% for the mother, and the average contribution of blubber to total mass of the mature males (n ¼ 5) was 14.8% and mature females (n ¼ 7) was 14.2%. The body burdens of the foetus/mother pair were used to calculate transfer from mother to offspring during gestation (Table 2), while the body burden differences between mature males and females were used to estimate the combined gestational and lactational transfer (Table 3) using the equation: Female burden ¼ male burden ð1
0:01PÞ where P (%) is the overall maternal transfer (Nakata et al., 1995)

3. Results and discussion 3.1. Compounds detected The OCPs identified in the blubber included hexachlorobenzene (HCB), hexachlorocyclohexanes (HCHs), viz, a-HCH, b-HCH, g-HCH and d-HCH, compounds of the DDT group, viz. 1,1,1-trichloro2,2ebis(4-chlorophenyl) ethane (p,p'-DDT), 1,1-dichloro-2,2-bis(4chlorophenyl) ethene (p,p'-DDE) and 1,1-dichloro-2,2-bis (4-chlorophenyl) ethane (p,p'-DDD), the DDT analogue methoxychlor, and the cyclodienes aldrin, dieldrin, heptachlor, heptachlor epoxide and g-chlordane. Essentially all of the anthropogenic compounds detected at quantifiable concentrations were pesticides, while only traces of compounds with industrial or other uses, such as polychlorinated biphenyls (PCBs), were found. The virtual absence of PCBs in these Tanzanian samples stand out from what has been reported for pantropical spotted dolphins (S. attenuata) off the eastern coast of South Africa (Cockcroft and Ross, 1991) where PCBs

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were the dominating organochlorines. This difference is not surprising as Zanzibar, in contrast to South Africa, has no heavy industry. The only anthropogenic compound found that may have other than pesticide use was HCB, which in addition to its pesticide use also may be formed during combustion (Müller, 1982). Summary data are given in Table 1, while the concentrations of the individual compounds in each dolphin are given in the electronic supplement Table E1. In addition to OCPs, both dolphin species contained MeO-BDEs, most likely of biogenic origin. These were identified with GCeMS by fragments at m/z 79, 81, 159, 161 516 and 530 (Vetter, 2001). Although the presence of several brominated compounds was indicated, including dimethoxy and chloro-bromo compounds, only two, 20 -methoxy-2,30 ,4,50 -tetrabromodiphenyl ether (20 -MeO-BDE68) and 6-methoxy-2,20 ,4,40 -tetrabromodiphenyl ether (6-MeO-BDE47), could be confirmed and quantified with authentic standards. 3.2. Organochlorine pesticides In both species, residues of the DDT group dominated among the OCPs (54e98%), followed by cyclodienes, HCHs and HCB. The sum of DDT residues (SDDT ¼ p,p'-DDE þ p,p'-DDD þ o,p'-DDT þ p,p'-DDT, concentration range 500e93 000 ng/g lipid, Table E1 in the electronic supplement) from both species was dominated by p,p'-DDE which comprised between 82% and 99% in Indo-Pacific bottlenose and between 86% and 95% in spinner dolphins (Fig. 2A). This contribution of p,p'-DDE was higher than found in Indo-Pacific humpback dolphins (Sousa chinensis) (13e43.5%) from the waters off Southern China (Leung et al., 2005) and common bottlenose dolphins (Tursiops truncatus) (48.6e89.1%), from the western Mediterranean Sea (Borrell and Aguilar, 2007). The high proportion of p,p'-DDE in dolphins from Zanzibar may indicate that DDT has not been used recently in the area or that the metabolism of DDT to DDE is very rapid. DDT was officially banned in Tanzania in 1988, but some was sold illegally in the beginning of the 1990s under the name Chlorphenethane (Mmochi and Mberek, 1998). The general lack of environmental data from the region makes any firm conclusion on the reason for the dominance of DDE over DDT in these samples difficult. Among the HCH isomers, b-HCH accounted for about 61% and 66% of the total concentration of all HCH isomers (SHCH) in IndoPacific bottlenose and spinner dolphins, respectively (Fig. 2B, Table E1 in electronic supplement). Detection of all four major isomers with b-HCH dominating and low levels of a-HCH, which dominates in the technical product, is an indication of past use of technical HCH. In technical HCH, a-HCH contributes 60e70% of the total (Buser and Müller, 1995), but its average contributions to the SHCH was low in both Indo-Pacific bottlenose (9%) and spinner dolphins (11%). g-HCH accounted for 24% and 19% of the SHCH in IndoPacific bottlenose and spinner dolphins, respectively. b-HCH is highly persistent in cetaceans since it resists enzymatic degradation and its predominance has also been reported in other species (Karuppiah et al., 2005; Leung et al., 2005). The cyclodienes showed similar concentration profiles in both Indo-Pacific bottlenose and spinner dolphins (dieldrin > aldrin > heptachlor epoxide > g-chlordane > heptachlor). Dieldrin, which

Table 2 Estimated whole-body burdens (mg) of organohalogen compounds and gestational transfer (%) in an Indo-Pacific bottlenose (Tursiops aduncus) mother/foetus pair.

Mother body burden (mg)a Foetus body burden (mg) Gestation transfer (%)b a b

HCB

b-HCH

SHCH

Aldrin

Dieldrin

SCyclodienes

Methoxychlor

p,p'-DDE

SDDT

SOCP

SMeO-BDE

0.57 0.0067 1.2

0.74 0.027 3.5

1.6 0.038 2.3

0.053 ND e

1.1 0.022 2.0

1.2 0.022 1.8

0.96 ND e

22 0.53 2.4

24 0.63 2.6

28 0.7 2.4

140 0.67 0.48

Body burden ¼ blubber weight  organohalogen concentration (ng/g lipid)  % lipid content/100. Gestational transfer ¼ burden in foetus  100/burden in (foetus þ mother) (Tanabe et al., 1982).

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Table 3 Estimated whole-body burdens (mg) of organohalogen compounds in mature individuals and overall mother-to-calf transfer during gestation and lactation (%) in Indo-Pacific bottlenose dolphins. HCB

b-HCH

Indo-Pacific bottlenose dolphin Tursiops aduncus 1.0 0.89 Mature male body burden (mg)a Mature female body burden (mg) 0.43 0.71 57 21 Overall transfer P (%)b a

SHCH

Aldrin

Dieldrin

SCyclodienes

Methoxychlor

p,p'-DDE

SDDT

SOCP

SMeO-BDE

1.4 1.1 22

0.21 0.16 23

3.0 1.5 47

5.2 2.7 48

0.63 0.43 31

282 80 71

337 92 72

340 96 72

1300 179 85

Body burden ¼ blubber weight  organohalogen concentration (ng/g lipid)  % lipid content/100. Overall transfer (P) ¼ (1
female burden/male burden)100 (Nakata et al., 1995).

b

was detected in all blubber samples, strongly dominated among the cyclodienes with average contributions of 88% in Indo-Pacific bottlenose and 77% in spinner dolphins. Dieldrin has high persistence and hydrophobicity (log KOW in the range 3.7e6.2, Ritter et al., 1996), typical for compounds that may undergo bioconcentration and biomagnification. The presence of aldrin is interesting. Aldrin is transformed to dieldrin (Bann et al., 1956) and the presence of aldrin in environmental samples has been taken to indicate recent sources, e.g., active use or emissions from stocks. Dieldrin residues, on the other hand, have been seen as a combination of residues of dieldrin used as insecticide in its own right and transformed residues of aldrin. This could, in part, explain the dominance of dieldrin among the cyclodienes. However, dieldrin may also be transformed to aldrin by intestinal bacteria (Kitamura et al.,1999). The presence of aldrin in the dolphins may, therefore, be a consequence of conversion of dieldrin residues in the food, but other sources cannot be excluded at this time. Similar to aldrin, heptachlor is transformed to heptachlor epoxide, which may explain why heptachlor epoxide is the third most common cyclodiene in the samples. All cyclodienes are hydrophobic and bioaccumulate (Ritter et al., 1996). As cyclodienes are no longer registered for use in Tanzania, the contribution of these highly persistent compounds to the total contaminant levels are low. In the mid 1980s the average dieldrin concentration in Tanzanian fish was 55 ng/g lipid (Paasivirta et al., 1988), similar to the concentration found in dolphins in this study. As dolphins feed at a higher trophic level than most fish and dieldrin is known to biomagnify through the food chain, the general levels of dieldrin in the Tanzanian environment most likely have gone down since the mid 1980s.

S. longirostris

T. aduncus

A

0%

p,p'-DDT

o,p'-DDT

p,p'-DDE

p,p'-DDD

20%

40%

60%

80%

α-HCH

β-HCH

γ-HCH

δ-HCH

100%

S. longirostris

T. aduncus

B

The most likely sources of the contaminants are local, but influence of remote sources on the occurrence of organochlorine pesticides, especially transport from the Indian subcontinent during the summer monsoon November to March, should not be disregarded. However the high temperatures of these waters favours volatilization (Wania and Mackay, 1996), making deposition of pollutants airborne from remote sources less likely. 3.3. Brominated compounds 6-MeO-BDE47 and 20 -MeO-BDE68 were the dominating organohalogen compounds in all spinner dolphin samples and all but two Indo-Pacific bottlenose dolphin samples. However, these MeOBDEs were found at significantly higher concentrations than anthropogenic OCPs in both spinner dolphins (t-test ¼
6.40, df ¼ 34, p ¼ 0.0001) and Indo-Pacific bottlenose dolphins (t-test ¼
2.28, df ¼ 34, p ¼ 0.03). The overall mean concentration of MeO-BDEs in spinner dolphins (74 000 ng/g lipid) was more than five times the mean concentration of OCPs (14 000 ng/g lipid). Similarly, the mean concentration of MeO-BDEs in Indo-Pacific bottlenose dolphins (62 000 ng/g lipid) was about three times higher than the OCPs (20 000 ng/g lipid). These findings indicate that brominated biogenic compounds are capable of bioaccumulation in animal tissues similar to the classical persistent organic pollutants with which they share physico-chemical properties. Both 6-MeO-BDE47 and 20 -MeO-BDE68 have been widely reported in different marine organisms (Malmvärn, 2007), but the concentrations reported here are higher than levels reported in any other marine mammal worldwide. For example, food products of cetaceans bought as retail in Japan had 48e2900 ng/g lipid (Marsh et al., 2005), dolphins from Mediterranean Sea 15e808 ng/g lipid (Pettersson et al., 2004), sea lions (Zalophus californianus) from California < 0.3e62 ng/g lipid (Stapleton et al., 2006), bottlenose dolphins (940e11 000 ng/g lipid) and dugong (Dugong dugon) (240 ng/g lipid) from North-eastern Australia (Vetter et al., 2001b). The concentrations of MeO-BDEs in dolphins from Zanzibar suggest the presence of potent sources of MeO-BDEs in the Western Indian Ocean. Identifying these sources would give insights in the ecological significance of these compounds. In addition to the MeO-BDEs identified these samples contained many brominated compounds that we could not identify due to lack of reference standards. Particularly noteworthy are at least one dimethoxy-BDE and a mixed chloro-bromo compound. Both dimethoxy-BDEs (Marsh et al., 2005) and a mixed chloro-bromo compounds (Vetter et al., 2001a) have been identified previously, but we do not know if the compounds found off Zanzibar are the same as those previously identified. 3.4. Inter-specific variations

0%

20%

40%

60%

80%

100%

Fig. 2. Relative proportions of A) DDT and B) HCH residues in blubber of Indo-pacific bottlenose (T. aduncus) and spinner (S. longirostris) dolphins off Zanzibar.

The inter-specific variations of residues in the two dolphin species indicate similarities in important aspects of their feeding ecology. The two species contained the same types of residues and

H. Mwevura et al. / Environmental Pollution 158 (2010) 2200e2207

similar composition trends of OCPs. These similarities suggest that these two dolphin species are subjected to common sources of organochlorines. All specimens were from the coastal habitats off the northern coast of Unguja Island and in the waters between Unguja Island and Tanga on the Tanzanian mainland (Amir et al., 2005a). A comparison of the food preference between the two species from the same geographical area indicated that although both species fed mainly on fish (TA 81%, SL 84% by weight), Indo-Pacific bottlenose dolphins feed mainly on small and medium-size neritic fish and squid (Amir et al., 2005b), while the diet of spinner dolphins consisted mainly of mesopelagic fish and squid of smaller size (S. Hamed, pers. comm.). However, the two dolphin species have little dietary overlap of fish species (4.5%), but greater overlap of cephalopods species 67% (S. Hamed, pers. comm.). Given this relatively small overlap in food preferences, further studies on the POP concentrations in other species are necessary to understand the flow of contaminants in the respective food chains of the two dolphin species.

3.5. Correlation between organohalogens The two quantified MeO-BDEs showed similar concentration patterns in the two dolphin species. In Indo-Pacific bottlenose dolphins, the concentration of 6-MeO-BDE47 (mean ¼ 37 000 ng/g lipid) was not significantly difference to that of 20 -MeO-BDE68 (mean ¼ 24 000 ng/g lipid) (t-test ¼
0.47, df ¼ 34, p ¼ 0.64). Likewise, in spinner dolphins the concentration of 20 -MeO-BDE68

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(mean ¼ 42 000 ng/g lipid) was not significantly different to that of 6-MeO-BDE47 (mean ¼ 31 000 ng/g lipid) (t-test ¼ 0.67, df ¼ 34, p ¼ 0.51). Further, the MeO-BDEs in both species were positively correlated (R2 ¼ 0.70, Fig. 3A), suggesting similar routes of exposure or mechanism of accumulation. There was positive correlation (R2 ¼ 0.52, Fig. 3B) between OCPs and MeO-BDEs measured in two dolphin species, though not as strong as that of the two MeO-BDEs. These correlations may suggest that the two organohalogen classes share some of the accumulation route in dolphin tissues.

3.6. Correlation with sex and maturity, and maternal transfer Lipid-normalized concentrations of OCPs and MeO-BDEs in both Indo-Pacific bottlenose and spinner dolphins showed sex and sexual maturity-related variations (Fig. 4). In the present study levels of SOCP showed a significant increased accumulation with age in both male Indo-Pacific bottlenose dolphins (r ¼ 0.67, p ¼ 0.05) and male spinner dolphins (r ¼ 0.77, p ¼ 0.005). Similarly, the SMeO-BDE levels increased significantly with age in both male Indo-Pacific bottlenose dolphins (r ¼ 0.85, p ¼ 0.004) and male spinner dolphins (r ¼ 0.68, p ¼ 0.02). In contrast, there was no correlation between levels of SOCP with age in females (TA r ¼
0.18, p ¼ 0.67; SL r ¼ 0.14, p ¼ 0.76). However, the SMeO-BDE levels decreased significantly with age in female Indo-Pacific bottlenose dolphins (r ¼
0.31, p ¼ 0.045), but not in female spinner dolphins although there was a trend (r ¼
0.72, p ¼ 0.07). In other studies of marine mammals, POP levels increase with age in males, but decrease in females due to transfer from mother to offspring (Borrell and Aguilar, 2005; Kajiwara et al., 2008). The sex/age

A A

5

150

2

Mean conc. (μg/g lipid) in T. aduncus

log [6-MeO-BDE47]

R = 0.7023

4

3

2 2

3

4

5

log [2'-MeO-BDE68]

B

Immature female Mature female

120

Immature male Mature male

90

60

30

0

ΣOCPs

ΣMeO-BDE

6

B Mean conc. (μg/g lipid) S. tenuirostris

log [ Σ MeO-BDE]

100 2

5

R = 0.5153

4

3

2 2

3

4

5

log [ Σ OCs]

Immature female

80

Mature female Immature male Mature male

60

40

20

0 20 -MeO-

Fig. 3. Correlation between the concentrations of A) 6-MeO-BDE47 and BDE68, and B) S-MeO-BDEs and organochlorine pesticides (S OCPs) in samples of dolphin blubber off Zanzibar.

ΣOCPs

ΣMeO-BDE

Fig. 4. Concentrations of OCPs and MeO-BDEs with sex and maturity in A) Indo-Pacific bottlenose and B), spinner dolphins.

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H. Mwevura et al. / Environmental Pollution 158 (2010) 2200e2207

variations observed in this study are likely also attributed to maternal transfer to the offspring. Transfer of hydrophobic contaminants from mother to offspring (maternal transfer) occurs during gestation and lactation (Kajiwara et al., 2008). The actual amounts transferred of each compound differ greatly depending on transfer route and the compounds' properties and is also affected by the metabolism rate of the respective compound in the species. For most of the compound groups about 2% of the mother's body burden had been transferred to the foetus (Table 2). b-HCH was transferred to the highest extent (3.5%), while HCB (1%) and particularly MeO-BDEs (0.5%) were transferred to the lowest extent during gestation. However, it must be pointed out that we had only access to one mother-foetus pair. Investigations of additional pairs are needed to gain accurate information on maternal transfer process. The total transfer of organohalogens from mother to offspring during gestation and lactation (Kajiwara et al., 2008) as indicated by the body burdens of mature males and females were 20e30% for the HCHs, aldrin, and methoxychlor, 40e60% for HCB and dieldrin, and around 70% for the DDTs (Table 3). The highest estimated total transfer (85%) was of the MeO-BDEs. Notably these compounds were transferred to a less extent than other compounds during gestation. The most important observation, however, is that the estimated total transfer is higher than the gestational transfer. Thus, the total transfer is dominated by lactational transfer with limited contribution of gestational transfer. Similar observations have been made for other species (Borrell et al., 1995; Nakata et al., 1995). The difference between gestational and lactational transfer was also possible to study due to the presence of three young calves (SL 006, SL 012 and SL 025, age indicated by milk in their stomachs). The concentrations of OCPs in the calves were 10 times higher than in foetus while MeO-BDEs were 76 times higher in the calves than in the foetus. However, in this first set of data the number of samples, particularly of mother/foetus pair and calves, was limited, and further studies with more samples are needed to fully understand the transfer processes. A female striped dolphin (Stenella coeruleoalba) was estimated to lose between 4 and 9% (Tanabe et al., 1982) of her total body burden of chlorinated contaminants during gestation and the transfer can go up to more than 90% after the lactational period (Tanabe et al., 1981). These estimations of maternal transfer of OCPs in striped dolphin were higher than those estimated in this study (Table 2). The overall transfer of MeO-BDEs (84%) estimated for Indo-Pacific bottlenose dolphins in this study is similar to that estimated for anthropogenic PBDEs in melon-headed whale (85%) (Kajiwara et al., 2008). However, transfer during gestation estimated in this study (0.5%) is lower than estimated in the melonheaded whale (2.6e3.5%) (Kajiwara et al., 2008). The observed differences in maternal transfer might be caused by differences in the lactation period between the species and the number of times the female dolphin has given birth. Other possible causes of differences in transfer may be attributed to the method used for calculation as well as age and sex composition of the analyzed samples used for calculations. As a consequence of maternal transfer, levels of pollutants in mature male dolphins are normally higher than in other age/sex classes. However, in both species the two contaminant groups followed the same concentration pattern: mature males > immature females > mature females > immature males. Transfer of a large proportion of the organohalogen body burden from female dolphins to their offspring during gestation and lactation show that dolphin calves are exposed to high doses of organohalogens. The mother's firstborn calf is most affected, and is expected to receive more than the second or subsequent calves (Wells et al., 2005). This type of reproductive transfer has been

associated with low chance of survival of the firstborn and sometimes second calves (Reddy et al., 2001). 4. Conclusion As the dominating organohalogens were presumably of natural origin, further studies on the identity and ecological consequences of natural organohalogen compounds in the Western Indian Ocean is called for. It is also necessary to follow the concentration changes of anthropogenic compounds in the future by further environmental monitoring programmes. These should include further studies of the possible health effects on marine mammals and other coastal top predators. Acknowledgments Anna Hellström and Lutufyo Mwamtobe gave technical assistance in the lab, and Dr. Narriman S. Jiddawi, Institute of Marine Sciences, Zanzibar, logistic support. Henk Bouwman gave valuable input to the manuscript. Financial support came from the International Programmes in Chemical Sciences (IPICS), Uppsala University, and the Swedish International Development Agency (SIDA) regional marine science programme administered by Swedmar. Appendix. Supplementary material Supplementary material associated with this paper can be found, in the online version, at doi:10.1016/j.envpol.2010.02.027. References Agrawal, M.S., Bowden, B.F., 2005. Marine sponge Dysidea herbacea revisited: another brominated diphenyl ether. Marine Drugs 3, 9e14. Åkerblom, M., 1995. Environmental Monitoring of Pesticide Residues, Guideline for the SADC Region, SADC/ELMS Monitoring Technique Series 3, Maseru, Leshoto. Amir, O.A., Jiddawi, N.S., Berggren, P., 2005a. The occurrence and distribution of dolphins in Zanzibar, Tanzania, with comments on the differences between two species of Tursiops. Western Indian Ocean Journal of Marine Sciences 4, 85e93. Amir, O.A., Berggren, P., Ndaro, S.G.M., Jiddawi, N.S., 2005b. Feeding ecology of the Indopacific bottlenose dolphins (Tursiops aduncus) incidentally caught in the Gillnet fisheries off Zanzibar, Tanzania. Journal of Estuarine, Coastal and Shell Sciences 63, 429e437. Amir, O., 2010. Biology, ecology and anthropogenic threats of Indo-pacific bottlenose dolphis in east Africa. PhD thesis, Stockholm University, Sweden. Bann, J.M., DeCino, T.J., Earle, N.W., Sun, Y.-P., 1956. The fate of aldrin and dieldrin in the animal body. Journal of Agricultural and Food Chemistry 11, 937e941. Boon, J.P., Oostingh, I., van der Meer, J., Theo, M., Hillebrand, J., 1994. A model for the bioaccumulation of chlorobiphenyl congeners in marine mammals. European Journal of Pharmacology 270, 237e251. Borrell, A., Aguilar, A., 2005. Mother-calf transfer of organochlorine compounds in the common dolphin (Delphinus delphis). Bulletin of Environmental Contamination and Toxicology 75, 149e156. Borrell, A., Aguilar, A., 2007. Organochlorine concentrations declined during 1987e2002 in western Mediterranean bottlenose dolphins, a coastal top predator. Chemosphere 66, 347e352. Borrell, A., Bloch, D., Desportes, G., 1995. Age trends and reproductive transfer of organochlorine compounds in long-finned pilot whale from the Faeroe Islands. Environmental Pollution 88, 293e298. Buser, H.R., Müller, M.D., 1995. Isomer and enantioselective degradation of hexachlorocyclohexane isomers in sewage sludge under anaerobic conditions. Environmental Science & Technology 29, 664e672. Cockcroft, V.G., Ross, G.J.B., 1991. Occurrence of organochlorines in stranded cetaceans and seal from the east coast of southern Africa. UNEP. Marine Mammals Technical Report 3, 271e276. Gray, J.S., 2002. Biomagnification in marine systems: the perspective of an ecologist. Marine Pollution Bulletin 45, 46e52. Kajiwara, N., Kamikawa, S., Amano, M., Hayano, A., Yamada, T.K., Miyazaki, N., Tanabe, S., 2008. Polybrominated diphenyl ethers (PBDEs) and organochlorines in melon-headed whales, Peponocephala electra, mass stranded along the Japanese coasts: maternal transfer and temporal trend. Environmental Pollution 156, 106e114. Karuppiah, S., Subramanian, A., Obbard, J.P., 2005. Organochlorine residues in odontocete species from southeast coast of India. Chemosphere 60, 891e987.

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