This article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and education use, including for instruction at the authors institution and sharing with colleagues. Other uses, including reproduction and distribution, or selling or licensing copies, or posting to personal, institutional or third party websites are prohibited. In most cases authors are permitted to post their version of the article (e.g. in Word or Tex form) to their personal website or institutional repository. Authors requiring further information regarding Elsevier’s archiving and manuscript policies are encouraged to visit: http://www.elsevier.com/copyright
Author's personal copy S CIE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
a v a i l a b l e a t w w w. s c i e n c e d i r e c t . c o m
w w w. e l s e v i e r. c o m / l o c a t e / s c i t o t e n v
Trace element concentrations in Proteocephalus macrocephalus (Cestoda) and Anguillicola crassus (Nematoda) in comparison to their fish host, Anguilla anguilla in Ria de Aveiro, Portugal C. Eira a,c,⁎, J. Torres b , J. Miquel b , J. Vaqueiro c,d , A.M.V.M. Soares a , J. Vingada a,c,d a
CESAM & Departamento de Biologia, Universidade de Aveiro, Campus de Santiago 3810-193 Aveiro, Portugal Facultat de Farmàcia, Laboratori de Parasitologia i Microbiologia Sanitàries, Universitat de Barcelona, Avinguda Joan XXIII, s/n 08028 Barcelona, Spain c Sociedade Portuguesa de Vida Selvagem, Estação de Campo de Quiaios, Apartado 16 EC Quiaios 3081-101 Figueira da Foz, Portugal d Departamento de Biologia, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal b
AR TIC LE D ATA
ABSTR ACT
Article history:
The use of some fish parasites as bioindicators of heavy metal pollution has been demonstrated
Received 3 June 2008
as particularly adequate due to their capacity of bioconcentration. This study evaluated the effect
Received in revised form
of Proteocephalus macrocephalus on the accumulation of trace elements in the edible fish, Anguilla
21 September 2008
anguilla, in a contaminated area in Portugal (Ria de Aveiro). Also, the model P. macrocephalus/A.
Accepted 15 October 2008
anguilla was assessed as a bioindicator system in the presence of the highly prevalent nematode Anguillicola crassus. Samples (kidney, liver, muscle, A. crassus and P. macrocephalus) of 20 eels
Keywords:
harbouring A. crassus and another 20 harbouring both A. crassus and P. macrocephalus were
Heavy metals
selected for element analysis by ICP-MS. The highest concentrations of Cr, Ni and Zn were
European eel
detected in P. macrocephalus. However, there was a higher liver and muscle Cr concentration in
Helminth parasite/host interaction
eels not infected by P. macrocephalus. Also, the nematode A. crassus presented higher Cr
Estuarine pollution
concentrations in those eels harbouring P. macrocephalus. Results suggest that P. macrocephalus
Aveiro lagoon
individuals accumulate Cr and Ni while levels of Cr in eel livers and Ni levels in eel kidney are reduced. The system P. macrocephalus/A. anguilla yielded bioaccumulation factors for Cr, Ni, Pb and Zn, whereas bioaccumulation of Cu, Cr and Pb in A. crassus varied according to eel co-infection with P. macrocephalus, thus emphasising the possible role of cestode infection in metal metabolization/storage processes in host tissues. Results suggest that heavy metal pollution in Ria de Aveiro has been decreasing although it is still higher than in other contaminated areas in Europe. Nevertheless, eel consumption in Ria de Aveiro represents no risk for humans although they may represent a real contamination risk for wildlife. The system P. macrocephalus/A. anguilla is proposed as another promising bioindicator system to evaluate environmental Cr, Ni, Pb and Zn exposure in estuarine areas where both species co-occur. © 2008 Elsevier B.V. All rights reserved.
1.
Introduction
Severe population declines of the European eel Anguilla anguilla have been recorded along European waters over the
last two decades (Santillo et al., 2005; Bevacqua et al., 2007), and in Portugal it is presently considered an endangered species (Cabral et al., 2006). The eel population decline has been related to climate change, over-fishing, habitat loss, dam
⁎ Corresponding author. SPVS, Estação de Campo de Quiaios, Apartado 16, EC Quiaios 3081-101 Figueira da Foz, Portugal. Tel.: +351 233 910 670. E-mail address:
[email protected] (C. Eira). 0048-9697/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.scitotenv.2008.10.040
Author's personal copy 992
SC IE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
building, poor water quality, including chemical pollution, and increments of parasite populations (Feunteun, 2002). Considering chemical pollution, eels presenting a high contaminant burden, including heavy metals, and low energy stores may reveal migration failure and/or reproduction impairment (Belpaire and Goemans, 2007) and they may also be more susceptible to diseases (Langston et al., 2002). With respect to parasite infection, the nematode Anguillicola crassus is a pathogenic helminth potentially representing an additional factor in the decline of eel populations (Münderle et al., 2004; Kennedy, 2007). In fact, A. anguilla is unable to mount an effective immune response against A. crassus (Taraschewski, 2006). Apart from its concerning conservation status, the European eel has been considered a highly suitable biomonitor for environmental contaminants, as eel tissue concentrations are related to pollution levels of prey species, surface water and sediments (Belpaire and Goemans, 2007; Maes et al., 2008). In fact, A. anguilla has been used in the past to assess environmental levels of heavy metal compounds (Collings et al., 1996; Edwards et al., 1999; Yamaguchi et al., 2003). Furthermore, the European eel is an important food resource, both for wildlife and humans, and thus the assessment of contaminant levels in its tissues can also indicate likely intakes and threats to consumers (Santillo et al., 2005). Although the evaluation of contaminants in fish tissues yields good results, the use of some fish parasites as bioindicators of heavy metal pollution has been demonstrated as particularly adequate due to their capacity of bioconcentration (e.g., Sures et al., 1999; Sures, 2001; Sures and Siddall, 2003). Concerning the European eel, whereas the model A. crassus/A. anguilla is already known to be an inadequate bioindicator system for lead pollution (Zimmermann et al., 1999), the model involving the acanthocephalan Paratenuisentis ambiguus provided high bioconcentration factors (Sures et al., 1994; Zimmermann et al., 1999). Cestode/eel systems have never been evaluated even though some systems involving cestode parasites of other fish have produced good results as potential bioindicator systems (Riggs et al., 1987; Sures et al., 1997). Therefore, the main goal of the present study was to explore the potential effect of the cestode P. macrocephalus on the accumulation of trace elements in A. anguilla and to evaluate the model P. macrocephalus/A. anguilla as a bioindicator system in the presence of the highly prevalent A. crassus, in a contaminated area in Portugal (Ria de Aveiro).
mineral acids, plastics, aromatics, among others. The contamination pathways from anthropogenic sources into the Ria de Aveiro have been extensively documented (e.g. Monterroso et al., 2003; Ramalhosa et al., 2006).
2.2.
Between May and September of 2007, 125 eels were collected from the central area of Ria de Aveiro, which is directly influenced by the main effluent from the Estarreja industrial complex (Estarreja Channel). All eels were frozen (−20 °C) for posterior analysis. After thawing, eels were weighted and measured, and samples of kidney, liver and muscle were collected from all individuals during dissection. These samples were stored in glass vials and deep-frozen until posterior processing for trace element analysis. All swim bladders and digestive tracts were removed and scanned for helminths using a stereomicroscope with the help of stainless steel instruments and Milli-Q water. A. crassus and P. macrocephalus representatives of all infected eels were frozen separately for posterior trace element analysis and the remaining individuals were preserved in 70% alcohol for identification according to standard helminthological methods.
2.3.
Materials and methods
2.1.
Study area
The Ria de Aveiro is a meso-tidal coastal lagoon located in central Portugal with a total area ranging from 66 to 83 km2 according to tidal variation (Dias et al., 2001). Apart from harbour, urban and agricultural wastes, the lagoon receives industrial waste from an industrial chemical complex (located in Estarreja), which includes a chlorine-alkali plant functioning since the 50s (in the past, the main anthropogenic source of Hg in the lagoon) and other industries producing fertilizers,
Analytical procedure
In order to select the individuals for the toxicological analysis, eels were divided into groups including those harbouring both A. crassus and P. macrocephalus and those carrying A. crassus but lacking the cestode. Twenty eels were selected randomly from each of these groups and therefore, samples (kidney, liver, muscle, A. crassus and P. macrocephalus) of 40 eels were used in this analysis. Samples were weighed (±100 mg wet weight) and digested in teflon vessels with HNO3 (2 ml) and H2O2 (1 ml) (Merck, Suprapure), at 90 °C in an oven and left overnight (14 h). All materials used in the digestion process were thoroughly acid-rinsed. After digestion, samples were diluted with Milli-Q water and then analysed for trace elements by ICP-MS (Perkin Elmer Elan 6000). The analytical procedure was checked using standard reference material Dogfish (Squalus acanthias) liver (DOLT-3) and muscle (DORM-2) (National Research Council, Canada). Ten samples of each standard material were analysed. Several analytical blanks were prepared and analysed along with samples in order to determine the detection limits.
2.4.
2.
Sampling
Data analysis
A normal distribution of all data was obtained after log (x + 1) transformation. Differences between element concentrations among the analysed tissues were detected by an ANOVA followed by Tukey's test. An unpaired t-test (two tailed) was used to compare element concentrations in the same host tissue between both eel groups. Linear regression analysis was used to detect relations between trace element levels in eels and parasites. Statistical analysis was performed in Statview 4.5 software package. For all tests, a significance level of P b 0.05 was applied. The bioaccumulation factors were determined according to Sures et al. (1999), as the ratio of the
Author's personal copy 993
S CIE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
Table 1 – Detection limits (ng ml − 1 ) and element concentrations (µg g− 1) in the standard reference material DORM-2 and DOLT-3 determined by inductively coupled mass spectrometry (ICP-MS) Detection Standard limit
As Cd Cr Cu Hg Ni Pb Zn
0.04 b 0.01 0.98 b 0.01 0.04 0.11 0.05 1.92
DORM-2 DOLT-3 DORM-2 DORM-2 DOLT-3 DORM-2 DORM-2 DORM-2
ICP-MS value, mean (± SD) 18.31 17.58 31.26 1.90 3.31 17.77 0.06 23.04
(1.10) (0.47) (3.54) (0.16) (0.81) (3.89) (0.01) (1.56)
Certified value, mean (±CI) 18.00 19.40 34.70 2.34 3.37 19.40 0.07 25.60
Accuracy (%)
(1.10) (0.60) (5.50) (0.16) (0.14) (3.10) (0.01) (2.30)
101.70 90.60 90.10 81.30 98.30 91.60 96.90 90.00
SD, standard deviation; CI, confidence interval.
element concentration in the parasites to that in different host tissues (BF = C[parasite] / C[host tissue]).
3.
Results
The adult swimbladder nematode A. crassus was detected in 73 eels indicating a prevalence of 58.0% and an intensity of 7.5 (1–32) parasites. The cestode P. macrocephalus was detected in 25 eels indicating a prevalence of 20% with an intensity of 4.2 (1–19). The following results refer to a group of 40 eels, all infected with A. crassus, including 20 eels infected with P. macrocephalus.
3.1.
Element distribution in host tissues and parasites
The detection limits (mean blank value plus 3 standard deviations of the mean blank) for each element and accuracy values are presented in Table 1. No data on accuracy values are presently available for Platinum Group Metals (hereby PGMs). Element concentrations detected in liver, kidney and muscle of eels and in their parasites A. crassus and P. macrocephalus are presented in Table 2. Whereas the highest values for As, Cu and Pb were found in liver, the highest values for Cd and Hg were detected, respectively, in kidney and in muscle (see Table 2). In
fact, there was a 2- and 3-fold increase in Hg concentration in eel muscle in relation to values registered in liver and kidney, respectively. The highest concentrations of Cr, Ni and Zn were detected in P. macrocephalus. Also, very low amounts of Pd and Pt (PGMs), were detected in eel liver and muscle. The concentrations of all assessed elements presented significant differences among the analysed tissues (ANOVA, all P b 0.05). In both eel groups, kidney presented the highest Cd concentration (Tukey test, all P b 0.001) and the concentration of Cd in liver was also higher than Cd in muscle and in parasites (all P b 0.001). The concentration of Hg in kidney was lower than that found in muscle (Tukey test, both P b 0.001) in both groups. In eels harbouring P. macrocephalus, Hg in kidney was also lower than that in liver (P b 0.05) but higher than values found in both helminths (both P b 0.01). Also, Hg in P. macrocephalus was higher than in A. crassus (P b 0.05). In those eels not harbouring the cestode, Hg in eel muscle was higher than that in liver (P b 0.01) and A. crassus (P b 0.001). No differences were detected between Cr values in all eel tissues and A. crassus in eels harbouring P. macrocephalus, although the highest Cr concentration was detected in the cestode (Tukey test, all Pb 0.001). On the contrary, significant differences were found between Cr levels in eels not infected by P. macrocephalus. In fact, the lowest amount of Cr was found in A. crassus (all Pb 0.001) while kidney presented less Cr than liver (Pb 0.01) and muscle (Pb 0.001). Furthermore, there was a higher average liver and muscle Cr concentration in eels not infected by P. macrocephalus in comparison to infected eels (t-test, t=3.35, P=0.002 and t=2.18, P=0.035). Also, the nematode A. crassus presented much higher Cr concentrations in eels infected by P. macrocephalus than those in eels lacking the cestode (t=2.56, P=0.015). A higher Ni concentration was found in P. macrocephalus when comparing to the assessed nematode (Tukey test, P b 0.001) and eel kidney (P b 0.01). The nematode also presented more Ni than eel kidney (P b 0.01). In the group of eels not harbouring the cestode, Ni levels in liver and kidney were significantly higher than Ni in A. crassus (both P b 0.01). In both groups, muscle presented lower Pb concentrations than liver and kidney (Tukey test, all Pb 0.001). In eels harbouring P. macrocephalus, the amount of Pb in A. crassus was lower than in liver and kidney (both Pb 0.001) while the amounts of Pb in A. crassus (Pb 0.05) and in P. macrocephalus (Pb 0.001) were
Table 2 – Trace element concentrations in eel tissues, A. crassus and P. macrocephalus (µg g− 1 wet weight) from Ria de Aveiro Liver
As Cd Cr Cu Hg Ni Pb Zn Pd Pt
Kidney
Muscle
A. crassus
P. macrocephalus
Mean
Range
Mean
Range
Mean
Range
Mean
Range
Mean
Range
1.630 0.058 0.659 11.669 0.084 0.380 0.188 43.398 0.00072 0.00033
0.613–2.834 0.003–0.173 0.329–1.132 2.353–32.852 0.030–0.163 0.091–2.117 0.041–0.444 20.289–76.512 0.0003–0.0010 0.00004–0.0007
1.286 0.305 0.545 2.213 0.053 0.278 0.180 52.886 nd nd
0.543–2.879 0.018–1.013 0.327–0.855 1.474–3.396 0.019–0.117 0.092–0.739 0.028–0.426 28.624–108.490
1.274 0.003 0.738 0.220 0.138 0.156 0.023 13.906 0.0005 0.0001
0.387–3.069 0.001–0.011 0.382–1.527 0.131–0.567 0.055–0.285 0.085–0.283 0.008–0.096 8.852–21.588 0.00008–0.00143 0.00001–0.00017
0.657 0.004 0.336 2.015 0.014 0.501 0.065 19.041 nd 0.00016
0.180–5.291 0.000–0.021 0.046–2.011 0.466–12.713 0.002–0.065 0.001–5.705 0.004–0.437 5.573–96.157
0.465 0.004 5.144 1.853 0.034 2.420 0.117 110.109 nd nd
0.105–1.554 0.002–0.009 0.256–24.368 0.402–5.758 0.007–0.080 0.189–8.651 0.022–0.299 20.704–301.821
nd, not detected.
0.00006–0.00031
Author's personal copy 994
SC IE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
Table 3 – Negative linear regressions relating element levels in parasites (P. macrocephalus and A. crassus) in relation to eel tissues in both eel groups Eel group
df
[Cr]P. macrocephalus — [As]liver [Cr]P. macrocephalus — [Cr]liver [Cu]P. macrocephalus — [Zn]liver [Cu]P. macrocephalus — [Mn]liver [Cu]P. macrocephalus — [Cu]liver [Cu]P. macrocephalus — [Cr]liver [Ni]P. macrocephalus — [Ni]kidney [Zn]A. crassus — [Cr]liver [Cu]A. crassus — [Cr]liver [Cr]A. crassus — [Zn]liver [Cr]A. crassus — [Cu]liver [Cr]A. crassus — [Cr]liver [Hg]A. crassus — [Cu]muscle No cestode [Zn]A. crassus — [Cr]kidney [Cu]A. crassus — [Cr]kidney [As]A. crassus — [Cr]kidney With cestode
F
P
R2
1,15 5.083 0.041 0.27 1,15 4.933 0.043 0.26 1,14 6.332 0.026 0.33 1,14 5.745 0.032 0.31 1,14 7.146 0.019 0.36 1,14 6.254 0.027 0.33 1,9 6.005 0.040 0.43 1,13 8.017 0.015 0.40 1,15 8.401 0.012 0.38 1,12 12.060 0.005 0.52 1,12 5.794 0.035 0.35 1,13 11.970 0.005 0.50 1,9 5.965 0.040 0.43 1,19 8.816 0.008 0.33 1,19 5.112 0.036 0.22 1,19 10.449 0.005 0.37
higher than in muscle. In eels not harbouring P. macrocephalus, the amount of Pb in A. crassus was lower than in liver and kidney (both Pb 0.001). No differences were detected in As levels among eel tissues. Nonetheless, eel tissues presented higher As concentrations than values found in A. crassus (Tukey test, all P b 0.001) in both eel groups. Eel tissues also presented higher As concentrations than that found in P. macrocephalus (all P b 0.001). Among all eel tissues, muscle presented lower Zn concentration than levels found in liver and kidney (Tukey test, both P b 0.001). A. crassus also presented lower values than liver and kidney (all P b 0.001). With respect to eels harbouring P. macrocephalus, the cestode presented the highest Zn concentrations in comparison to eel tissues (liver and muscle P b 0.001, kidney P b 0.05) and also in comparison to A. crassus (P b 0.001).
3.2. Relations between element concentrations in eel tissues and parasites Many positive relations were obtained for almost all of the evaluated elements concentrations in P. macrocephalus and A.
crassus and eel tissues (not shown). However, considering element concentrations detected in P. macrocephalus, it was possible to obtain negative relations including the concentrations of Cr, Cu and Ni in the cestode and element levels in eel livers and kidneys. In fact, the Cu levels in P. macrocephalus were negatively related to the levels of Cu, Zn, Mn and Cr in eel livers (see Table 3). Also, the Ni level in the cestode was negatively related to Ni levels in eel kidney. Finally, the Cr level in the cestode was negatively related to the levels of both Cr and As in eel livers. Considering element levels in A. crassus in eels harbouring P. macrocephalus, the concentrations of Zn and Cu were negatively related to the levels of Cr in livers whereas the levels of Zn and Cu in livers were negatively related to Cr levels in A. crassus. In those eels not infected with P. macrocephalus, the levels of Zn, Cu and As in A. crassus were negatively related to Cr levels in eel kidney.
3.3.
Bioaccumulation factors
Considering the trace element levels detected in P. macrocephalus and in the respective host, it was possible to detect that Cr, Ni, Pb and Zn (Table 4), were present in higher levels in the cestode than in eel tissues. In fact, higher amounts of both Cr and Zn were detected in P. macrocephalus in comparison to levels detected in eel liver, kidney and muscle. Although mean Zn bioaccumulation factors indicate approximately 2and 3- fold increases in cestode levels in comparison to kidney and liver, respectively, there was a 9-fold increase in mean Zn concentration in P. macrocephalus in comparison to eel muscle (Table 4). With respect to Cr, while the mean concentration detected in P. macrocephalus was 9-times higher than that found in eel muscle, bioaccumulation factors indicate a 10and 12-fold increase in Cr levels in the cestode in comparison to eel kidney and liver. Also, the mean Pb concentration in P. macrocephalus was over 8-times higher than that in eel muscle. The highest bioaccumulation factor was found for Ni since the concentration of this element in the cestode was almost 16-times higher than that found in eel kidney. Apart from the mean bioaccumulation factors (BFs) detected for P. macrocephalus, relatively lower BFs for Cr, Ni, Pb and Cu were obtained when relating metal levels in A. crassus to those in eel tissues (Table 4). Bioaccumulation of Cu
Table 4 – Mean (SE) and range of bioaccumulation factors, [C]parasite / [C]host macrocephalus and A. crassus in relation to eel tissues Liver
P. macrocephalus
A. crassus
Cr Ni Pb Zn Cr Pb Cu
tissue,
for some elements detected in P.
Kidney
Muscle
Mean
Range
Mean
Range
Mean
Range
12.64 (4.37) – – 2.91 (0.52) 2.56 a (1.41) – – –
0.36–70.55
10.39 (3.06) 15.83 (5.56) – 2.11 (0.32) – – – –
0.54–53.33 0.36–51.20
9.44 (2.88) – 8.42 (2.47) 9.12 (1.68)
0.28–44.90 2.31–18.22 1.63–25.37
5.14 a(2.69) 2.76 b (1.00) 6.74 b (1.42)
0.22–24.63 0.01–9.79 2.91–31.94
0.77–7.66 0.06–20.41
Ratios corresponding to less than a 2-fold increase in concentrations are not presented. a BF values in eels harbouring the cestode. b BF values in eels not harbouring the cestode.
0.36–4.93
Author's personal copy S CIE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
was detected in those eels not infected with P. macrocephalus (BF = 6.7) whereas Cr bioaccumulation (BF = 2.6) was detected in the eel group infected by the cestode. Finally, Pb bioaccumulation was detected in both eel groups (5.1 in infected eels and 2.8 in those eels not infected with P. macrocephalus).
4.
Discussion
4.1.
Element distribution in A. anguilla
A previous study developed in Ria de Aveiro (Pérez Cid et al., 2001) reported Cd, Cu, Ni, Pb and Zn concentrations in muscle of several fish species, including European eels. In the present study, lower general levels were obtained, suggesting that heavy metal pollution in Ria de Aveiro has decreased following the major pollution control measures applied to production processes since the 1990s in the nearby chemical industry compound. In fact, other authors have already referred to the decrease in industrial metal discharges over the last two decades (e.g. Monterroso et al., 2007). The Ni and Zn levels obtained in the present study were similar to some of the average values reported for several areas within Ria de Aveiro (Pérez Cid et al., 2001), whereas Cd and Cu presented a one-third reduction and Pb was reduced to half, approximately. Although eel liver and kidney presented higher Pb concentrations, the muscle Pb range was generally lower than values obtained for other wild eel studies across Europe in moderately or highly polluted waters (see Ureña et al., 2007). The Pb declining trend was also detected in Belgium during a 10-year study on heavy metal contamination of eels, perhaps as a result of the change to unleaded fuels and lower industrial emissions (Maes et al., 2008). Despite the reduction in contaminant levels in eels from Ria de Aveiro, some element concentrations remain relatively intricate. For example, considering trace element levels in eels from salt marshes on the South-Atlantic coast of Spain (Usero et al., 2003), both As and Ni in eel livers correspond to approximately half of the amount detected in the present study. Furthermore, Hg and Cr levels in Ria de Aveiro were, respectively, 7- and 12-times higher (in liver) and 6- and 4-times higher (in muscle) than in the respective tissues of eels from the most polluted area studied by Usero et al. (2003). As described for different fish species (e.g. Pierron et al., 2007; Reynders et al., 2008) the highest Cd concentrations were found in kidneys in both eel groups (infected and not infected with P. macrocephalus) analysed in the present study. Despite the above-mentioned Cd decrease in eel muscle from Ria de Aveiro, it is noteworthy that Cd in eel kidney ranged between 0.018 and 1.013 µg g− 1. Considering the maximum amount of Cd (0.1 µg g− 1 w.w.) in the edible portion of eels allowed by commission regulation EC 466/2001, it was possible to verify that 95% of eel kidneys presented concentrations over the stipulated value, emphasising the need to ensure that kidneys are removed when preparing eels for consumption by Humans. Vinhas and Shirley (1986) detected that eels from the most polluted site in Ria de Aveiro presented liver Hg levels of 0.396 µg g− 1 and muscle levels of 0.386 µg g− 1. Two decades later, the present study shows that the Hg levels in eels have substantially decreased and that levels were well below the
995
maximum Hg value allowed in the edible portion of eels (EC 466/2001). No differences were detected in Cr or in As mean levels among eel tissues, although differences were detected for Cr when considering both eel groups separately (see below). With regard to eel muscle, Cr and As mean levels (respectively, 0.738 and 1.234 µg g− 1) were higher than values reported in Belgium by Maes et al. (2008) (0.255 and 0.168 µg g− 1, respectively). Arsenic was also present in higher amounts in eel muscle from Ria de Aveiro (present study) than mean values detected in the Mersey estuary in England (Collings et al., 1996) and in river Turia in southern Spain (Bordajandi et al., 2003), respectively 0.110 and 0.228 µg g− 1. With respect to Pd and Pt (PGMs), these are released from automobile exhaust catalysts and are present in the environment in low concentrations. Little is known about their bioavailability in aquatic organisms and their accumulation along the food web (Sures et al., 2002, 2003). In this study, Pd and Pt were detected in low amounts in liver and muscle. However, results indicate a relatively higher mean concentration of Pd in liver (0.72 ng g− 1 w.w. ranging from 0.3 to 1.0 ng g− 1 w.w.) than that detected in A. anguilla from a fish farm in Germany (0.18 ng g− 1 w.w.) (Sures et al., 2001). The present results are in accordance with Sures et al. (2002), suggesting that Pd should have a greater bioaccumulation capacity among PGMs and that eels may be suitable indicators of PGM pollution in aquatic systems (Zimmermann et al., 2004).
4.2. Element distribution in A. anguilla in relation to helminths Sures and Siddall (1999) described a lower metal content in fish hosting helmith parasites in comparison to fish without parasites. Also, lower concentrations of Cd, Cu, Zn and As were detected in perch (Perca fluviatilis) infected by the tapeworm Proteocephalus percae in comparison to those not harbouring the cestode (Turcková and Hanzelová, 1999; Turcková et al., 2002). Similarly, the present results suggest that P. macrocephalus individuals accumulate Cr and Ni while reducing the levels of Cr in eel livers and Ni levels in eel kidney. Furthermore, the negative relation between Cu levels in P. macrocephalus and Cu, Zn, Mn and Cr levels (essential micronutrients for endogenous metabolism) in eel livers may be related with metal mobilization/uptake mechanisms in the cestode and in livers, where induction and binding to metallothioneins (MTs) may occur. In the present study, a higher liver Cr concentration was found in those eels not infected by P. macrocephalus in comparison to infected eels. Fernandes et al. (2008) found a negative correlation between eel hepatosomatic index and Cr residues in eel liver, suggesting a negative impact of a high Cr content on fish health. Therefore, considering the lower eel liver Cr contents in infected eels, the present results may suggest a beneficial role of cestode infection. Although no negative relation was detected between Cr concentration in P. macrocephalus and in muscle, results suggest that the presence of the cestode P. macrocephalus may be related with the lower levels of Cr in muscle of infected eels. Data obtained in this study also suggests a possible role of A. crassus infection (or A. crassus and P. macrocephalus co-
Author's personal copy 996
SC IE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
infection) in the distribution of trace metals in eel tissues. Notice that A. crassus presented a higher Ni concentration than liver and kidney in those eels harbouring the cestode, whereas the opposite was true for eels without the cestode. There was also a higher Hg concentration in liver in relation to kidney only in eels infected with the cestode. In this case, the Hg distribution may be related with a higher hepatic MT concentration in infected eels, considering a possible role of parasite infection on MT expression. Despite the variation in the characteristics of metal-binding proteins among fish species (Atli and Canli, 2007) and in elements' relative affinities for hepatic MTs (Langston et al., 2002; Bird et al., 2008), the high binding affinity detected between MTs and Hg in fish (Olsson et al., 1998) may help to explain the higher Hg concentration in livers of eels infected with P. macrocephalus. The negative relation between the concentrations of Zn and Cu in A. crassus and the levels of Cr in livers or in kidney according to cestode infection, may also be related to a higher MT availability in livers of eels infected with P. macrocephalus.
4.3.
Bioaccumulation factors
Although higher bioconcentration factors have been detected in parasite/host systems involving acanthocephalans (e.g. Sures and Taraschewski, 1995; Sures and Siddall, 1999) some cestodes seem to accumulate relatively high amounts of heavy metals and cestode/fish systems may serve as rather useful indicators of heavy metal contamination in the aquatic environment (Riggs et al., 1987; Sures et al., 1997; Jirsa et al., 2008). However, more data are available on parasites of freshwater than on parasites of estuarine or marine fish (Sures, 2001). Furthermore, a study on metal accumulation in Bothriocephalus scorpii infecting turbot Scophthalmus maximus (a marine fish) detected less lead and cadmium than in Monobothrium wageneri infecting tench Tinca tinca, a freshwater fish (Sures et al., 1997), suggesting a lower bioavailability of metals to hosts and parasites in the marine environment, although other factors (parasite size and portion, microhabitat and mode of nutrition) may affect cestode accumulation capacities (Sures et al., 1997; Torres et al., 2006). Therefore, the present study suggests that the system P. macrocephalus/A. anguilla may be an adequate bioindicator model in the case of Cr, Ni, Pb and Zn contamination in estuarine environments. In fact, apart from the heavy metal accumulation potential, the present parasite/host system should be relatively common considering that 20% of the analysed eels were infected with P. macrocephalus. Furthermore, P. macrocephalus is specific to eels and presents a Holarctic distribution (Scholz et al., 2007 and references therein). Previous studies on A. crassus detected either no metal bioconcentration (Sures et al., 1994; Zimmermann et al., 1999) or low metal bioconcentration (Tenora et al., 1999). The latter authors found higher Cd, Cr, Ni and Pb concentrations in A. crassus than in eel muscle. However, muscle Cr and Ni concentrations reported by Tenora et al. (1999) were lower than levels found in eel muscle from Ria de Aveiro. With respect to Pb, the liver levels reported by Sures et al. (1994) in naturally infected eels (0.18 μg g− 1 w.w.) were very similar to those detected in the present study (0.19 μg g− 1 w.w.). Therefore, the higher Pb level in A. crassus from Ria de Aveiro (0.07 μg
g− 1 w.w.) is relatively more significant than the value reported by Sures et al. (1994) (0.02 μg g− 1 w.w.). In general, it was confirmed that the accumulation and concentration values of heavy metals in A. crassus are considerably low, in comparison to other fish parasite species (acanthocephalan and cestode species, in particular). However, it was possible to verify that the bioaccumulation factors for Cu, Cr, Ni and Pb in A. crassus/ A. anguilla varied according to eel co-infection with P. macrocephalus, emphasising the possible role of cestode infection in metal metabolization/storage processes in host tissues.
5.
Conclusion
Presently, eel consumption in Ria de Aveiro represents no risk for humans. In fact, the values obtained in the edible portion of eels in this study stayed way below the maximum limits allowed in EU regulations, and consequently, in regard to possible effects of the studied metals, the consumption of these fish by humans should be safe, once all viscera have been removed. However, eels are prey to birds and carnivores, and therefore they may represent a real contaminant risk for wildlife. Furthermore, although more field and experimental essays are necessary to evaluate the relationship between bioaccumulation in cestode parasites of fish and environmental metal availability, we presently propose the system P. macrocephalus/A. anguilla as another promising bioindicator system to evaluate environmental Cr, Ni, Pb and Zn exposure in aquatic areas where both species are present.
Acknowledgements This study was partially supported by project HP 2005-0011 (ACIN) from the Secretaría de Estado de Educación, Universidades, Investigación y Desarrollo (SEID) and Conselho de reitores das Universidades Portuguesas (CRUP), and by fellowship SFRH/BPD/27014/2006 provided by the Fundação para a Ciência e Tecnologia of the Portuguese MCTES. Authors wish to thank all personnel at the “Serveis Científics i Tècnics” of the University of Barcelona (Spain).
REFERENCES Atli G, Canli M. Natural occurrence of metallothionein-like proteins in liver tissues of four fish species from the Northeast Mediterranean sea. Water Environ Res 2007;79(9):958–63. Belpaire C, Goemans G. Eels: contaminant cocktails pinpointing environmental contamination. ICES J Mar Sci 2007;64:1423–36. Bevacqua D, Melià P, Crivelli AJ, Gatto M, de Leo GA. Multi-objective assessment of conservation measures for the European eel (Anguilla anguilla): an application to the Camargue lagoons. ICES J Mar Sci 2007;64:1483–90. Bird D, Rotchell J, Hesp S, Newton L, Hall N, Potter I. To what extent are hepatic concentrations of heavy metals in Anguilla anguilla at a site in a contaminated estuary related to body size and age an reflected in the metallothionein concentrations? Environ Pollut 2008;151:641–51. Bordajandi LR, Gomez G, Fernandez MA, Abad E, River J, Gonzalez MJ. Study on PCBs, PCDD/Fs, organochlorine pesticides, heavy
Author's personal copy S CIE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
metals and arsenic content in freshwater species from the river Turia (Spain). Chemosphere 2003;53:163–71. Cabral MJ, Almeida J, Almeida PR, Dellinger T, Ferrand de Almeida N, Oliveira ME, et al. Livro vermelho dos vertebrados de Portugal, 2nd ed. Instituto da Conservação da Natureza/Assírio and Alvim. Lisboa, 2006. Collings SE, Johnson MS, Leah RT. Metal contamination of angler-caught fish from the Mersey Estuary. Mar Environ Res 1996;4:281–97. Dias JM, Lopes JF, Dekeyser I. Lagrangian transport of particles in Ria de Aveiro Lagoon, Portugal. Phys Chem Earth B 2001;26:721–7. EC 466/2001. Official Journal of the European Community; 2001. Annex I, JO L77 16.3. Edwards SC, MacLeod CL, Lester JN. Mercury contamination of the eel (Anguilla anguilla) and roach (Rutilus rutilus) in East Anglia, UK. Environ Monit Assess 1999;55:371–87. Fernandes D, Bebianno MJ, Porte C. Hepatic levels of metal and metallothioneins in two commercial fish species of the Northern Iberian shelf. Sci Total Environ 2008;391:159–67. Feunteun E. Restoration and management of the European eel: an impossible bargain? Ecol Eng 2002;18:575–91. Jirsa F, Leodolter-Dvorak M, Krachler R, Frank C. Heavy metals in the nase, Chondrostoma nasus (L. 1758), and its intestinal parasite Caryophyllaeus laticeps (Pallas 1781) from Austrian Rivers: Bioindicative Aspects. Arch Environ Contam Toxicol 2008;55:619–26. Kennedy C. The pathogenic helminth parasites of eels. J Fish Dis 2007;30:319–34. Langston WJ, Chesman BS, Burt GR, Pope ND, McEnvoy J. Metallothionein in liver of eels Anguilla anguilla from the Thames Estuary: an indicator of environmental quality? Mar Environ Res 2002;53:263–93. Maes J, Belpaire C, Goemans G. Spatial variations and temporal trends between 1994 and 2005 in polychlorinated biphenyls, organochlorine pesticides and heavy metals in European eel (Anguilla anguilla L.) in Flanders, Belgium. Environ Pollut 2008;153:223–37. Monterroso P, Abreu SN, Pereira E, Vale C, Duarte AC. Estimation of Cu, Cd and Hg transported by plankton from a contaminated area (Ria de Aveiro). Acta Oecol 2003;24:S351–7. Monterroso P, Pato P, Pereira E, Millward GE, Vale C, Duarte AC. Metal-contaminated sediments in a semi-closed basin: implications for recovery. Estuar Coast Shelf Sci 2007;71:148–58. Münderle M, Sures B, Taraschewski H. Influence of Anguillicola crassus (Nematoda) and Ichthyophthirius multifiliis (Ciliophora) on swimming activity of European eel Anguilla anguilla. Dis Aquat Organ 2004;60:133–9. Olsson PE, Kling P, Hogstrand C. Mechanisms of heavy metal accumulation and toxicity in fish. In: Langston WJ, Bebianno MJ, editors. Metal metabolism in Aquatic Environments. London: Chapman and Hall; 1998. p. 321–50. Pérez Cid B, Boia C, Pombo L, Rebelo E. Determination of trace metals in fish species of the Ria de Aveiro (Portugal) by electrothermal atomic absorption spectrometry. Food Chem 2001;75:93–100. Pierron F, Baudrimont M, Bossy A, Bourdineaud JP, Brèthes D, Elie P, et al. Impairment of lipid storage by cadmium in the European eel (Anguilla anguilla). Aquat Toxicol 2007;81:304–11. Ramalhosa E, Pato P, Monterroso P, Pereira E, Vale C, Duarte AC. Accumulation versus remobilization of mercury in sediments of a contaminated lagoon. Mar Pollut Bull 2006;52:332–56. Reynders H, Bervoets L, Gelders M, de Coen WM, Blust R. Accumulation and effects of metals in caged carp and resident roach along a metal pollution gradient. Sci Total Environ 2008;391:82–95. Riggs MR, Lemly AD, Esch GW. The growth, biomass and fecundity of Bothriocephalus acheilognathi in a North Carolina cooling reservoir. J Parasitol 1987;73:893–900.
997
Santillo D, Johston P, Labunska I, Brigden K. Swimming in Chemicals: widespread presence of brominated flame retardants and PCBs in eels (Anguilla anguilla) from rivers and lakes in 10 European countries. Greenpeace Technical Note, vol. 12.; 2005. p. 55. Scholz T, Hanzelová, Škeříková A, Shimazu T, Rolbiecki L. An annotated list of species of the Proteocephalus Weinland, 1858 aggregate sensu de Chambrier et al. (2004) (Cestoda: Proteocephalidea), parasites of fishes in the Palaearctic Region, their phylogenetic relationships and a key to their identification. Syst Parasitol 2007;67:139–56. Sures B. The use of parasites as bioindicators in aquatic ecosystems: a review. Aquat Ecol 2001;35:245–55. Sures B, Siddall R. Pomphorhynchus laevis: the intestinal acanthocephalan as a lead sink for its fish host, chub (Leuciscus cephalus). Exp Parasitol 1999;93:66–72. Sures B, Siddall R. Pomphorhynchus laevis (Palaeacanthocephala) in the intestine of chub (Leuciscus cephalus) as an indicator of metal pollution. Int J Parasitol 2003;33:65–70. Sures B, Taraschewski H. Cadmium concentrations of two adult acanthocephalans (Pomphorhynchus laevis, Acanthocephalus lucii) compared to their fish hosts and cadmium and lead levels in larvae of A. lucii compared to their crustacean host. Parasitol Res 1995;81:494–7. Sures B, Taraschewski H, Jackwerth E. Lead content of Paratenuisentis ambiguus (Acanthocephala), Anguillicola crassus (Nematodes) and their host Anguilla anguilla. Dis Aquat Organ 1994;19:105–7. Sures B, Taraschewski H, Rokicki J. Lead and cadmium content of two cestodes Monobothrium wageneri, and Bothriocephalus scorpii, and their fish hosts. Parasitol Res 1997;83:618–23. Sures B, Siddall R, Taraschewski H. Parasites as accumulation indicators of heavy metal pollution. Parasitol Today 1999;15:16–22. Sures B, Zimmermann S, Messerschmidt J, Bohlen A. Relevance and analysis of traffic related Platinum Group Metals (Pt, Pd, Rh) in the aquatic biosphere, with emphasis on palladium. Ecotoxicology 2002;11:385–92. Sures B, Zimmermann S, Messerschmidt J, Bohlen A, Alt F. First report on the uptake of automobile catalyst emitted palladium by European eels (Anguilla anguilla) following experimental exposure to road dust. Environ Pollut 2001;113:341–5. Sures B, Zimmermann S, Sonntag C, Stuben D, Taraschewski H. The acanthocephalan Paratenuisentis ambiguus as a sensitive indicator of the precious metals Pt and Rh from automobile catalytic converters. Environ Pollut 2003;122:401–5. Taraschewski H. Hosts and parasites as aliens. J Helminthol 2006;80:99–128. Tenora F, Baruš V, Kráčmar S, Dvořáček J, Srnková J. Parallel analysis of some heavy metals concentrations in metals concentrations in the Anguillicola crassus (Nematoda) and the European eel Anguilla anguilla (Osteichthyes). Helminthologia 1999;36:79–81. Torres J, Peig J, Eira C, Borrás M. Cadmium and lead concentrations in Skrjabinotaenia lobata (Cestoda: Catenotaeniidae) and in its host Apodemus sylvaticus (Rodentia: Muridae) in an urban dumping site in Spain. Environ Pollut 2006;143:4–8. Turčková L, Hanzelová V. Concentrations of Cd, As and Pb in non-infected and infected Perca fluviatilis with Proteocephalus percae. Helminthologia 1999;36(Suppl.):31. Turčková L, Hanzelová V, Špakulová M. Concentration of heavy metals in perch and its endoparasites in the polluted water reservoir in Eastern Slovakia. Helminthologia 2002;39:23–8. Ureña R, Peri S, del Ramo J, Torreblanca A. Metal and metallothionein content in tissues from wild and farmed Anguilla anguilla at commercial size. Environ Int 2007;33:532–9. Usero J, Izquierdo C, Morillo J, Gracia I. Heavy metals in fish (Solea vulgaris, Anguilla anguilla and Liza aurata) from salt marshes on the southern Atlantic coast of Spain. Environ Int 2003;29:949–56.
Author's personal copy 998
SC IE N CE OF T H E TOT AL E N V I RO N ME N T 4 0 7 ( 2 00 9 ) 9 9 1–9 98
Vinhas T, Shirley ML. Metais pesados em enguias e sedimentos colhidos na Ria de Aveiro. Instituto Hidrográfico, Divisão de Química e Poluição, Lisboa;1986. Yamaguchi N, Gazzard D, Scholey G, MacDonald DW. Concentrations and hazard assessment of PCBs, organochlorine pesticides and mercury in fish species from the upper Thames. River pollution and its potential influence on top predators. Chemosphere 2003;50:265–73. Zimmermann S, Sures B, Taraschewski H. Experimental studies on lead accumulation in the eel-specific endoparasites
Anguillicola crassus (Nematoda) and Paratenuisentis ambiguus (Acanthocephala) as compared with their host, Anguilla anguilla. Arch Environ Contam Toxicol 1999;37:190–5. Zimmermann S, Baumann U, Taraschewski H, Sures B. Accumulation and distribution of platinum and rhodium in the European eel Anguilla anguilla following aqueous exposure to metal salts. Environ Pollut 2004;127:195–202.
