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Distribution of Zn, Pb, Cd and Cu between seawater and transplanted mussels (Mytilus galloprovincialis)
The Science of the Total Environment, 119 (1992) 211-230 Elsevier Science Publishers B.V., Amsterdam
211
Distribution of Zn, Pb, Cd and Cu between seawater and transplanted mussels (Mytilus galloprovincialis) D. Martin~i6 a, Z. Kwokal a, 7,. Peharec a, D. Margug a and M. Branica ~'b aCenter for Marine Research Zagreb, 'Ruder Bogkovik' Institute, POB 1016, 41001 Zagreb, Croatia bInstitute of Applied Physical Chemistry, ICH-4, KFA, Juelich, POB 1913, D-5170 Juelich, Germany (Received June 28th, 1991; accepted July 22nd, 1991)
ABSTRACT Cultured one-year old mussels, Mytilus galloprovincialis, (Lmk.) were transplanted to four sites (120 individuals per site) throughout the Krka River Estuary, and nearby Adriatic coastal area. Concentrations of zinc, cadmium, lead and copper, and their content in mussels were analysed four times during 1988/89 (270 days). The body weight of mussels increased during the period studied and a discernible effect of the size of mussels on metal concentration was detected. The concentrations of the metals studied, expressed on a per unit dry weight, generally decreased with the body size and the age of mussels, but no seasonal trends could be established. The mussels transplanted to sites situated in the vicinity of the Sibenik harbour accumulated zinc and lead. This could be attributed to elevated concentrations of zinc and lead in water (higher than 500 ng Zn 1-~ and 14 ng Pb 1-I of 'bioavailable' forms). Significant correlation between body lead concentrations and lead amount in the surrounding water was obtained, while for zinc, no statistically significant relationship exists between body concentrations and the water phase. The fluctuations of cadmium and copper in the adjacent water have no significant effect on the changes of their concentrations in the transplanted mussels. The data obtained indicate that because of biological similarities and better reproducibility of the results the transplanted mussels are more suitable for monitoring of trace metals than indigenous mussel populations as originally proposed by the 'Mussel Watch' programme.
Key words: sea water; mussels; transplantation; trace metal concentration INTRODUCTION
Mussels have been utilized as valuable sentinel organisms for indicating levels of trace metal concentrations in marine environment because of their widespread distribution along coastal zones (Schulz-Baldes, 1973; Godlberg et al., 1978; Majori et al., 1978). Trace metal concentrations in mussels can also be affected both by biological and physical factors, such as: reproductive 004~-9697/92/$05.00
© 1992 Elsevier Science Publishers B.V. All rights reserved
212
o. MARTIN~I(~ETAL
state, sex, and biological condition of an individual mussel, season of sampling, temperature, salinity and a depth of organisms habitat. The effect of these variables on the metal content in tissues has not been completely clarified. Their effect is evident from a large variation in metal concentrations described by Martin~i6 et al. (1987c). The objectives of the long-term study undertaken (from 1983 to 1988) by these authors were to recognize and quantify influence of slightly polluted environment, with regard to trace metals present in the water phase (Branica et al., 1985), on metal concentration in indigenous mussel populations from various parts of the Krka River Estuary, Yugoslavia and nearby coastal area (Martin~i6 et al. (1985; 1986; 1987c). However, they found that natural populations of mussels, Mytilus galloprovincialis, often exhibit considerable natural variability in terms of trace metal levels which complicate attempts to detect changes resulting from anthropogenic sources. Mussels were sampled according to monitoring programme formulated by the UNEP Coordinated Mediterranean Pollution Monitoring and Research Programme (MED POL I, 1980). Moreover, it has been observed that, among all causative biological factors, ages, reproductive states, and habitats of these populations were different. For example, natural habitats of mussels differed significantly throughout the Krka River Estuary, depending on salinity and water depth. It is desirable and often possible, to eliminate factors that may confuse the interpretation of a monitoring survey. Therefore, the objective of this work is to reduce the causative biological factors which have been studied so far for indigenous mussel populations, and improve the homogeneity of the response in the test environments by transplanting mussels from the shell farm to selected sites. It has already been observed that the transplantation of mussels or similar organisms to various areas may provide an useful approach to marine environmental monitoring (Widdows et al., 1980-81; Widdows et al., 1984). Bioaccumulation of dissolved trace metals in the organisms is usually observed when they are moved from an uncontaminated to a contaminated area (Simpson, 1979; Stephensen et al., 1979; Martin et al., 1984). SAMPLING AND SAMPLE PRETREATMENT
Sampling area The Krka River Estuary situated in the eastern Adriatic coast is characterized by a low terrigenuous input of trace metals owing to a karstic drainage area and a series of travertine barriers which precede the estuary. A mean freshwater inflow of the Krka River is approximately 55 m 3 s -1 with considerable monthly variations ranging from 5 to 410 m 3 s -I. Apart from that, torrents and submarine springs occasionally contribute minor inflows. The Krka River Estuary is a stratified estuary characterized by a low
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
213
tidal range (20-50 cm) and a sheltered position. The brackish surface water current is directed seawards, whereas bottom seawater counter-current can be traced up to the Skradinski Buk waterfalls (Fig. 1). The area investigated is confined to the part of the ancient river valley between the last active calc tufa barrier, through a central enlarged part, and the Sibenik Channel for a total length of 22 km. The estuary gradually deepens from 2 m (at the waterfalls), to 42 m on the seaward side (the Sibenik Channel). The city of Sibenik (40 000 inhabitants) situated in the lower part of the estuary, with its shipping and metal industry, represents a hazard to aquatic life by directly discharging wastes into the estuary. Mussels can only survive in the water of salinity of 10%o or higher (Margug, 1983), which means at a depth of 5-6 m (station E-2 in the vicinity of Skradin, Fig. 1). In the Sibenik harbour (the lower part of the estuary) the mussels natural habitats are closer to the water surface and under possible tidal influence.
Hydrologic and chemical features of the estuary Multidisciplinary investigations of the Krka River Estuary have been carried out by the Centre for Marine Research since 1980, and form part of the long-term study MED POL Phase II (Annual reports, 1983-1989). During the spring of 1988 a mean freshwater inflow of the Krka River was approximately 58 m a s -l. It was greatly reduced to only 5 m 3 s -~ in the dry period (summer and autumn). Water temperature was measured in situ at a depth of 6 m and gradually increased from May (17°C) to August (22°C). In December water cooled down to 15°C. Salinity, measured conductometrically, varied between S = 37 and 39%0. Total suspended matter (SM), particulate (POC) and dissolved (DOC) organic carbon were measured in water samples collected at a depth of 6 m on May 15th, August 5th, September 4th and December 4th, 1988. Prior to measurements filtration of samples was performed on Whatman GF/F glass-fibre filters. Most of the time the river water is an optically very clear organic-poor medium (Cauwet, 1989). During the summer and autumn of 1988 concentrations of dissolved organic carbon (DOC) were generally < 0.9 mg 1-~ at all sampling sites with the exception of site E-2 where values exceeding 1 mg 1-I were obtained. This may be ascribed to resuspension since the sampling was performed close to the sediment surface. A significant difference in the concentrations of DOC was noted between water samples collected from the coastal site C-I (0.55-0.65 mg C Iq) and those from estuarine sites. High planktonic productivity which occurred in May 1988 was responsible for elevated concentrations of DOC at all sampling sites including the site C-1 (1.20 mg C 1-~). Particulate organic carbon (POC) concentrations varied between 0.168
214
D. MARTINI~I~ El" AL,
and 0.334 mg C 1-1 at all sampling sites. No significant difference in POC values was observed between spring and summer, 1988. Data obtained during the period 1988-1989 indicate that concentrations of suspended matter in the Krka River Estuary are lower than in bther estuaries (Martinic, 1988). The concentrations of suspended matter varied between 0.70 and 1.87 mg 1-1 Higher amount (5.43 mg 1-1) of suspended matter was only observed in May at the site E-4. Oxygen concentrations in the saline water layer were constant (5-6 ml 1-1) during the entire period with the exception of site E-2 in December when a minimum oxygen concentration of 3.7 ml 1-1 was obtained.
Transplanted mussels One-year old cultured mussels Mytilus galloprovincialis (Lmk.), (480 individuals), with well-defined biometric features, cultivated in the fishery and shell farm Sarina Draga (central part of the Krka River Estuary), were used as the starting stock. Average shell-lengths varied slightly (between 60.2 and 63.4 mm), with variation coefficients between 8 and 15%, (Table 1). The mussels were placed into oval plastic cages and moored in water of practically constant salinity (37-38% o), at a depth of 6 m, at four different locations (120 individuals each) (denoted E-2, E-4, E-5 and C-I, on Fig. 1). Since the depth of the estuary gradually increases from 8 m (at station E-2) to 42 m (stations E-5 and C-I), living depths for indigenous mussels differ along the estuary. The mussels were kept at four sites from April 24, 1988 to January 14, 1989 (266 days). The results published by Widdows et al. (1980-81) suggest that one month of exposure to different environmental conditions is sufficient to allow the equilibration with new environmental concentrations within the animals tissues, as well as for physiological adjustment to new living conditions. A month of acclimatization was allowed prior to sampling. Sampling and measurements of 25 individuals separated into four groups were performed periodically: June 1st, July 20th and September 30th, 1988 and January 14th, 1989. Sampling procedure was carefully observed in order to avoid contamination problems (Martin~i6 et al., 1980; 1984). Prior to analysis, no gutcleansing of the collected specimens was performed. Ten individuals from each group were used for biometric characterization, and only soft tissues of the remaining 15 specimens were homogenized and wet-digested prior to analyses for metal concentrations using ETAAS and FLAME-AAS (Martin~i6 et al., 1987c). The bivalve condition index was determined according to the Hopkins method (Margug, 1985), (the ratio between dry flesh and total weight of mussels).
0.102 0.151 0.178
E.4 b
stock stock stock stock
0.077 0.090 0.107 0.137
C.1 d
aThe starting bThe starting CThe starting dThe starting
0.095 0.125 0.164 0.158
E.5 c
of of of of
0.138 0.176 0.234 0.146
E_2 a
0.234
Dry/wet factor
Location
(1.2) (l.0) (1.3) (1.3)
(1.2) (2.0) (2.1) (2.1)
(1.5) (1.9) (1.5) (3.6)
(1.6) (3.1) (2.2) (2.7)
121 individuals 120 individuals 120 individuals 120 individuals
3.8 4.3 4.9 6.0
4.8 5.6 7.1 6.8
4.8 6.6 7.5 9.7
6.2 7.6 9.4 6.4
Wet weight in g (S.D.)
of of of of
(9) (6) (8) (7)
(8) (6) (7) (4)
(6) (8) (7) (6)
(5) (6) (6) (5)
(6) (8) (7) (7)
60 63 63 62
62 65 65 69
67 66 70 72
66 69 69 75
67 71 70 66
mm mm mm mm
Shell length in mm (S.D.)
in in in in
length length length length
was was was was
June July September January
June July September January
June July September January
June July September January
Month of collection
settled settled settled settled
14.6 15.0 19.2 21.8
22.0 29.7 23.7 30.4
17.8 20.2 26.0 29.1
23.0 23.0 27.0 39.3
in in in in
April. April April. April.
(0.3) (0.2) (1.0) (1.2)
(0.4) (1.3) (2.0) (1.0)
(0.4) (1.4) (0.3) (0.1)
(0.1) (0.8) (0.1) (0.1)
Zn (S.D.)
0.17 0.17 0.16 0.32
0.17 0.16 0.14 0.30
0.17 0.15 0.14 0.18
0.15 0.14 0.12 0.29
(0.01) (0.01) (0.01) (0.02)
(0.01) (0.01) (0.01) (0.01)
(0.01) (0.01) (0.01) (0.01)
(0.01) (0.01) (0.01) (0.01)
Cd (S.D.)
Me #g g-l wet weight
Data for transplanted mussels collected from various locations in the Krka River Estuary
TABLE 1
0.24 0.23 0.22 0.27
0.34 0.38 0.24 0.36
0.42 0.38 0.33 0.30
0.16 0.12 0.11 0.17
(0.01) (0.02) (0.02) (0.03)
(0.01) (0.03) (0.01) (0.02)
(0.01) (0.03) (0.03) (0.01)
(0.01) (0.01) (0.01) (0.01)
Pb (S.D.)
0.72 0.76 0.84 0.87
0.73 0.81 0.92 0.89
0.80 0.87 1.02 1.11
0.88 0.98 1.03 0.93
(0.06) (0.01) (0.01) (0.03)
(0.06) (0.01) (0.02) (0.03)
(0.04) (0.01) (0.02) (0.03)
(0.02) (0.06) (0.04) (0.03)
Cu (S.D.)
t,~
7~
Z ~7
7
216
D. MARTIN(~I(~ET AL,
Ambient water
Water samples for metal analyses were collected from a depth of 6 m by divers (Kniewald et al., 1987) on May 14th, July 29th, 1988 and January 15th, 1989. Prior to sampling pre-cleaned 1 1 Nalgene sampling bottles were thoroughly washed with diluted Suprapure HNO3. They were also preconditioned in the sea for 7 days. The bottles were protected against fouling by enclosing them in a plankton net (150/~m pore size) and fixed to the seafloor at a depth of 40 m. The top of the plankton net was fixed to an air buoy, so that the entire system floated - 2 0 m above the bottom. The bottles were left unstoppered so that seawater could rinse them. Prior to sampling, the bottles were again treated with acid (hot 10% Suprapure HNO3 in order to remove organic films formed on the bottles. Finally, the bottles were washed with tetra-distilled water. Electrochemical characterization of Zn, Cd, Pb and Cu was performed within 12 h of sampling. Differential pulse anodic stripping voltammetry (glassy carbon and hanging mercury drop electrodes) was used for determination of 'total' and 'free' dissolved metal concentrations in unfiltered water samples at pH 2 and 8 (4.7 and 8 for zinc). The instrumentation used was a PAR Polarographic Analyzer, Model 174A with an X-Y recorder, as described in detail elsewhere (M. Branica et al., 1985). RESULTS Dissolved and particulate metal concentration in water
Our results show (Table 2) that trace metal concentrations in the Krka River Estuary are significantly lower than the values obtained for estuaries throughout the world (Martinrir, MED POL Phase II, Annual Report, 1983-1987). There appears to be a number of reasons for this. Firstly, a relatively unpolluted catchment area provides effective baseline trace metal values for the Krka River feeding the Adriatic Sea. Secondly, a solid-liquid exchanges are limited by a low particulate loading (0.7-5.43 mg 1-Z) in the river water. Water samples collected from sampling sites E-4 and E-5 (in the vicinity of an industrial waste outflow of the city of Sibenik), were found to be enriched with zinc, lead and copper. However, the enrichment through anthropogenic sources is limited, because average 'total' metal concentration values decreased seawards reaching lower average values at the coastal site C-1 (520 ng Zn 1-1, 11 ng Cd -z, 34 ng Pb 1-Z, and 180 ng Cu 1-1). Uniform distribution of lead was observed in the entire water column at sites E-4 and E-5, while distribution of Zn, Cd and Cu differed significantly. Surface layer
May July January
May July January
May July January
May July January
E-2
E-4
E-5
C-I
Location Month of collection
420 530 670
490 820
690 350 --
1100 340 2110
.
.
320 260 --
720 150 300
210 430 440
240 620
.
370 90 --
380 190 1810
210 100 230
250 200
.
19 8 5
8 10 5
11 10 26
8 40
.
16 5 2
6 2 3
5 6 11
3 17
.
pH8
pH2
pH 4.8-8
pH4.8
pH8
Cd (S.D. < 2%)
Zn (S.D. < 5%)
3 3 3
4 8 2
6 4 15
5 23
.
pH2-8
.
65 22 15
61 49 35
89 53 27
23 11
pH2
.
51 10 4
18 13 7
11 13 6
5 3
14 12 11
43 36 28
78 40 21
. 18 8
pH8 pH2-8
Pb (S.D. < 3%)
.
220 120 190
160 290 160
596 483 260
340 230
pH2
.
60 20 60
30 30 50
70 97 57
12 37
pH8
Cu (S.D. < 2%)
160 100 130
130 260 110
526 386 203
328 173
pH2-8
Metal concentration (ng 1-i) in the unfiltered seawater samples collected at a depth of 6 m in the surrounding water of transplants
TABLE 2
-Zt r~
z
..-I >
> Z
-I
>.
> Z
218
o. MARTINI~I~ ET AL.
(0.5 m) at sites E-4 and E-5 contained higher concentrations of zinc and cadmium than deeper water (6 m), while copper concentrations at site E-4 were inverse. However, an average 'total' metal distribution in the water column at site E-2 differed from sites E-4, E-5 and C-1. The water samples collected at a depth of 6 m contained about two-fold higher average zinc concentration (660 ng Zn 1-1) than the surface water samples (310 ng Zn l-l). Concentrations of copper at a depth of 6 m (205 ng Cu -1) and at the surface (212 ng Cu l -l) are practically the same. This could be explained by the fact that a water mass at this depth is normally exposed to mineralized organic material remobilizing metals from adjacent sediments. The bottom depth of water at this sampling site is only 7-8 m. The data obtained through annual monitoring of the Krka River Estuary support this opinion. The water at a depth of 6 m contained significantly higher quantities of suspended matter than the surface water layer with the exception of sites E-4, E-5 and C-1 in December 1988. Dissolved and particulate metal concentrations varied through the year, mainly depending on the freshwater inflow. For example, in May 1988 a normal plankton bloom, caused by higher nutrients was accompanied by higher POC values, particularly at site E-4. This resulted in maximum concentration of zinc (850 ng Zn 1-1) and cadmium (30 ng Cd 1-1) in the surface water. It seems that lead and copper exhibited seasonal variations, reaching maximal values in August. Surface water from sites E-4 and E-5 (vicinity of the city of Sibenik) was found to be more enriched with lead and copper than deeper water indicating enrichment through anthropogenic sources (MED POL PHASE II, Annual Report, 1988).
Transplanted mussel population Mussels condition Cultured one-year old mussels were transplanted from the shellfish farm Sarina Draga (Fig. 1) to the Krka River Estuary. Prior to transplantation, 80% of individuals had already spawned, with an average condition index of 5.9. The somatic growth and induction of reproductive cycle (gametogenesis) are evident from the condition indexes, calculated by the Hopkins method and illustrated in Fig. 2. Mussels transplanted to site E-2 reached the highest condition indexes as compared with the mussels transplanted to other sites. Condition indexes of the mussels from four stations are illustrated in Fig. 2.
Biometric analysis The biometric data are summarized in Table 1 and illustrated in Fig. 3. A
219
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
15°45
-~(f 16°00'
Skrad Prokljan take nski Buk r falls)
arina Draoa.
.SIBENIK , Sibenik bay
~b
o2
4
Fig. 1. Map of the investigated area.
I0-
• E-2
9-
o E-4
.o
o 6-
initial musse[s
~ ^
~m,~
x~
~32
monfh M year days of exposure
' A ' M ' J' 1988 J' A' S' O' N' D' J'I~89 0
3B
8B
160
266
Fig. 2. Index of conditions (calculated by the Hopkins method) for transplanted mussels during 270 days of exposure at different sites in the Krka River Estuary.
220
D. MARTIN(~I(~ ET A L
b
108-
10-
-,-
i6:
4-
/ "
b=1,29 df= 1/, ' ~ '~'lb
b=0,96 df=l/, '
2,
;
' &'~'16
shelf weight(g)
meat v01ume(ml)
20
~lOt=n
-
oJ
6-
r =0,999 b =0,/,1 df=lt,
3=
I
I
0,2
l
d
20
t E°J =~ o .~, 10
O,I/+
I
I
I
I
I
I
I
0,6 0,8 1,0
2,0 dry weight(g)
r
4,0
• June o July z~ September ~ a n n u a r y
~ . .
~1 ~ +-~
'-t
:il ,=0,,+
\ ^ z.~
b= - 0,62 df = 1L,
I
0,2
i
I
0,/*
I
I
I
I
'
I
0,6 0,8 1,0
21
,0 dry weight(g)
I
4,0
Fig. 3. The biometric features of transplanted mussel populations collected in June, July, September 1988, and January 1989 at different sites in the Krka River Estuary (df, degree of freedom).
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
221
satisfactory correlation between shell weight and wet weight of the mussels was obtained (r = 0.944, d f = 14). This indicates that during the period investigated the changes in mussels body weight are mainly the result of their somatic growth (Fig. 3a). Flesh volume of all mussels studied varied proportionally (b = 0.96) with their wet weights (Fig. 3b) owing to the constant salinity (37%0) in the environment. A satisfactory correlation between dry and wet weights of mussels was also obtained (Fig. 3c). The ratio between the flesh volume and dry weight decreased with the increase of mussel dry weights (Fig. 3d) indicating different influence of the somatic growth of the mussels on the body increment expressed on the wet or dry units basis. In the case when the somatic growth has a priority significance over the reproductive cycle, the wet body increment is less pronounced than the increment of dry organic substances. Trace metal concentration in mussels Metal concentrations in mussels flesh varied significantly, depending on their living conditions, induced bio-rhythm and physico-chemical properties of surrounding waters. The data are summarized in Table 1. The metal concentrations in mussels are satisfactorily comparable with already published values for indigenous mussels from the same area (Martin6i6 et al., 1987). The zinc concentrations varied significantly between 14.6 and 39.3 #g Zn g-l wet weight. Mussels from the coastal site (C-l) show significantly lower zinc and copper concentrations (17.7 #g g-i and 0.80 tag g-i, respectively) than mussels from site E-2, where the highest values were obtained (28 tag Zn g-l and 0.95 tag Cu g-1 wet weight). The values obtained for cadmium are practically the same (0.18 and 0.20 tag Cd g-l). Concentrations of lead in mussels is reversed at these locations. The lead concentrations in mussels from site E-2 are lower (0.14 #g Pb g-l) than in mussels from site C-1 (0.24 tag Pb g-l). Average metal concentrations obtained suggest that the mussel body condition factors had pronounced effect on metal concentrations in mussels. Expressing the values on the wet body weight basis it becomes obvious that mussels accumulated zinc and copper during the tissue growth, while at the same time, average cadmium and lead concentrations decreased. The average values obtained for zinc and lead concentrations in mussels at sites near areas of a pollutant discharge (E-4 and E-5) are significantly higher than the concentrations obtained at other sites. In August 1988, the highest zinc concentrations (29/~g g-l) were found in the mussels from site E-5. The mussels from sites E-4 and E-5 were found to be enriched with lead of most probably anthropogenic origin, and contained two or three-fold
222
D. MARTIN~I~ ET AL
higher lead concentrations than mussels from sites E-2 (0.14 #g Pb g-l) and C-1 (0.24 #g Pb g-l). No enrichment in Cu and Cd which could exceed the calculated 'background' levels in mussels was observed. DISCUSSION Since metal concentrations in mussels from unpolluted environments vary widely (Martin~i6 et al., 1987c), only differences several orders of magnitude larger are considered to be significant (Goldberg et al., 1978). It appears that applicability of mussels as indicators of trace metal concentration levels refers only to significantly contaminated environments. Our previous studies show that the variations in trace metal concentrations of mussels are generally related to local living conditions and seasonal weight fluctuations (MartinEi6 et al., 1980; 1984; 1986; 1987a,b,c,). The results of a systematic study of the Krka River Estuary and the Kornati Islands carried out from 1983 to 1988, show almost uniform average zinc concentration values, while cadmium, lead and copper distributions differ (Martin~i6 et al., 1987c). Moreover, the mussels from the Kornati Islands (the reference area) contained significantly higher cadmium concentrations than might be expected from the area free from anthropogenic influence (Branica et al., 1985). It has been observed that the mussels from the areas of pollutant discharge (vicinity of the Sibenik harbour) contained higher lead and copper concentrations than the mussels from the Kornati Islands owing to the presence of higher metal concentrations in the water phase. However, these relations were not statistically proved since differences in metal concentrations as well as various biological factors, such as the mussels size, living conditions, season of sampling, etc., influence the uptake of trace metals in indigenous mussels in such a way that the metal concentrations in mussels from the slightly polluted site can be similar to those in specimens from an unpolluted site. Therefore, it was not possible to distinguish between slightly polluted and unpolluted areas. Indigenous populations of mussels sampled from different parts of the Krka River Estuary and the Kornati Islands could not be biologically compared. The average values of the biometric measurements of mussels transplanted from the shellfish farm Sarina Draga to the Krka River Estuary are summarized in Table 1. The transplants were distributed throughout the area at a depth of 6 m which guarantees marine living conditions. Condition indexes of the transplanted mussels depend on their local living conditions and season of sample collection, which is satisfactorily comparable with findings of Margug (1983). The mussels from the site E-2 in the upper part of the estuary, living near
Zn, Pb, Cd AND Cu 1N SEAWATER AND TRANSPLANTED MUSSELS
223
sediment surface, tended to have the highest nutritional state (Fig. 2) owing to the presence of larger quantities of suspended material, rich in POC. The average indexes of condition of the mussels from other sites gradually decreased in the seaward direction. The transplants living in the coastal environment, relatively poor in suspended matter, had the lowest indexes of condition. A maximum observed in June (Fig. 2) in the mussels from the site E-4 coincided with the phytoplankton bloom which usually occurs during this period of the year. The mussels from all studied sites reached the lowest indexes of condition in January 1989 owing to the spawning in December 1988. It has also become evident that the shell lengths of the mussels studied did not necessarily increase with age which can successfully be compared with earlier observations (Martin~i6 et al., 1986; 1987a,b). However, shell weights changed proportionally with the aging of mussels (Fischer, 1983). A significant (P < 0.01) regression line with a slope ofb = 1.29 (Fig. 3a) was obtained by relating log average wet weight to log average shell weight of transplants suggesting that the mussels somatic growth was primarily responsible for the body weight fluctuations during the entire period. Wet weights of mussels from all locations are directly proportional to their flesh volumes (Fig. 3b) (b = 0.96 with 1% probability level). Furthermore, the ratio between wet weights of mussels and water content in their bodies was fairly constant over the entire period owing to stable salinity values in the aquatic environment. Relation between wet and dry weights, shows different ratios for different mussel sizes (Table 1, Fig. 3d). The water concentration in the mussels decreases thereby increasing their dry weight, size and age. Concentrations of all metals in all samples collected from four experimental sites were fitted with regressions of log weight-log concentration of significant non-zero slopes. Our results indicate that larger mussels contained higher concentrations of zinc and copper than smaller ones (metal concentrations are expressed on a per unit wet weight). Accumulation of these metals depends on the mussels somatic growth, thereafter on the water content (Figs. 4a, d). Furthermore, different slopes of regression lines obtained for zinc and copper concentrations indicate that mussels metabolize these metals at different rates (Figs. 4a,b,d,e). On the other hand, zinc and copper concentrations expressed as a per unit dry weight exhibited the reverse association in comparison with concentrations based on a per unit wet weight. Their concentrations decrease as the body water content increases. The slopes obtained (Figs. 4c, f) indicate that larger mussels contain lower metal concentrations than smaller ones, which is in a good agreement with the results obtained by Cossa et al. (1980). Moreover, from the above relations it is evident that some metal concen-
224
D. MARTIN(~IC El" AL.
60a
Zn
~0-
2,0.
aE-2 /
d
Cu
3~ c
N 20
= 0,8-
r=0,872 b= 0,93 df=11 '
~
~ 'lb
~0,6-
b=0,58 df= I~,
0,4-
2b
~, ' b ' l b
'
shell weight(g )
shell weight(g)
6O
b
Zn
"G ~0-
2,0
oE-2
4--:
e
Cu
:" 30-
%
1,0-
~= o,a-
o, 20-
jo
;@3
~.o,6-
df=lO
10
b=0,4/+ dr= 11+
O,t,-
'
wet weight(g)
4
' ~'~'Ib
wet weight(g)
I+00
c
Zn oE-5
201
oE-2
o ~ ' ~ t _ ~
2O
hE'5
f
Cu
r =-0,9B7 b=- 0,63
10-
8Z
-.: 80-
20
0,2
16-
r= - 0,937 b=- 0,23 df= 10
-o 60-
i
i
0,~
f
°s, -
i
i
i
v 1,0
0,6 0,8 1
i
2,0 dry weighttg)
i
2't 0,2
i
i
0,~
i
t
i
i
i
i
0,6 0,8 1,0
i
2.0
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Fig. 4. Zn and Cu concentrations in the transplanted mussel populations collected in June, July, September 1988, and January 1989 at different sites related to the mussel biometric features.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
225
trations deviate from the calculated ideal line, i.e. baseline concentrations of the mussels. This suggests that besides tissue weights or shell sizes of the mussels there are other factors influencing the metal body concentrations. For example, the results obtained indicate that zinc concentration in mussels (E-5 in June, July, 1988 and January, 1989) does not overlap calculated regression lines (Figs. 4a,b,c). Comparing the zinc concentrations in the mussels from these periods with the zinc concentrations in the water phase, it becomes obvious that such deviations could only be attributed to the elevated metal concentrations in the adjacent water (Table 2), although no significant correlation was obtained when relating body zinc concentrations to the zinc concentrations in the water phase. Figure 4c and Table 2 show that amounts of zinc in mussels from site E-5 (June, July 1988 and January 1989) coincide with particulate and strongly complexed zinc concentrations (pH 4.8-8) (portion of 'bioavailable' metal) which exceed 500 ng Zn 1-1 in water phase. In the study carried out by Olafsson (1986), the metal concentrations of the juvenile mussels which had colonized the mooring structures were similar to the post-adaptation concentrations of the caged mussels, except for Hg and Zn. The concentrations of these metals were significantly lower in the juvenile mussels than the mean post-adaptation concentrations in the caged mussels. He suggests that a proportion of this initial tissue content of these metals is firmly bound in the caged mussels and therefore he proposes that mussels grown from a juvenile stage at monitoring sites could be more sensitive indicators of seawater metal concentrations than mussels transferred from a coastal site. Copper concentrations in mussels depend on body sizes only, regardless of the fact that the animals were exposed to significantly higher quantities of particulate and strongly complexed copper (pH 2 - p H 8) in surrounding waters. The highest copper concentrations in water were found in the vicinity of the Sibenik harbour (sites E-4 and E-5) (Table 2), but in spite of that no enrichment in copper which could exceed the baseline metal concentrations of mussels was obtained. Phillips (1976) found that the net uptake of copper by mussels was extremely erratic and suggested that the mussels are not suitable indicators of copper pollution in the marine environment. The same conclusion was drawn by Lyngby and Brix (1987). The ability of mussels to maintain their internal copper and zinc compositions at a steady level when variations in the metal composition of the external medium occur, was observed by ArmiardTriquet et al. (1986). Therefore, they concluded that the use of mussels as bioindicators of pollution was doubtful for essential metals, particularly as regards short-term pollution, since the levels of these trace elements in the organisms were largely independent of their concentration in the ambient seawater.
D. MARTIN(~IC ET AL
226
The concentrations of lead in larger mussels are lower than in smaller ones. Lead concentrations in mussel tissues decreased proportionally with an increase of tissue, water content, size and age (Figs. 5b,c). Most of the values of the Pb tissue concentrations fell above the regression line when related to dry tissue weight. This is most probably the result of anthropogenic input. 0,6-
Pb
a
_ E-t~o E~E-5
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\
.
?~E~ E-Z, mE-Z, xx~'1"E -5
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% g_ 0,2 r:-0,951 b: - lJ+
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•
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131o o [] E-2 E'5
i
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~o
shell weiohtlg)
0,~
b
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e
~: 0,4-
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O (~]A
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~0,,
8. o,:
r:-0,986
~N.
b:-0.96
"~
df:5
- N~,
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I
' -
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"~ N o
I
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,
,,I
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~
'
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~
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Fig. 5. P b and Cd concentrations in the transplanted mussel populations collected in June, J u l y , S e p t e m b e r 1988, and January 1989 at different sites related to the mussel biometric
features.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
227
Indeed, particulate and complexed lead concentrations which exceed 14 ng Pb 1-~ of seawater provoke proportional increase in metal concentrations in tissues (Fig. 5c). Mobilization of Pb by human activities, particularly the combustion of leaded petrol, has caused significant increase of Pb concentrations in the upper layers of the estuary. An unknown proportion of this lead has been deposited on the streets and roads and is washed away into the estuarine water. Lead of this origin is most probably the primary cause of the frequently encountered elevated Pb concentrations in mussels from sites E-4 and E-5. Moreover, this has been confirmed by the fact that lead concentrations in the mussels which fell above the regression line are well correlated with total (r = 0.912, P < 0.001) and 'bioavailable' lead (r = 0.905, P < 0.05) in the surrounding water. The same relation was obtained by Popham and D'Auria (1982). Satisfactory correlation was found between cadmium concentrations in the soft part of tissues and shell weights, the Cd concentrations changing inversely with respect to shell weight. Cadmium concentrations in the soft parts decreased when the water content-increased, i.e., size of mussels, what is in a good agreement with the findings by Phillips (1976). A seasonal dependence was not observed. All transplanted mussels collected in January 1989 contained cadmium concentrations which exceeded baseline cadmium values. During this period, cadmium concentrations in the water phase were similar to concentrations found during other sampling periods, with the exception of site E-2. Particulate and strongly complexed cadmium was rather high at this site, so that the concentrations obtained in the mussels from this site could be attributed to this event, but not in the case of other sites. A possible explanation of this event can be found in the data on surfactant activities obtained by a routine monitoring of the area studied (MED POL PHASE II, Annual Reports, 1988 (1989)). Surfactant activities measured both directly and in the filtrated water samples collected throughout the area at the end of December 1988, showed that water was enriched with dissolved organic material to the extent which exceeds the concentrations found during the period between June and December 1988. Such a water component of a nutritive nature could stimulate the mussels to filter larger volumes of water, thus accumulating higher quantities of cadmium naturally present in the water phase. At any rate, the event observed in the case of cadmium and mercury (unpublished data) confuses the interpretation of all other values fitted by log/log regression of weight-concentration, and needs further field investigations. CONCLUSIONS
Concentrations of all metals studied changed significantly and inversely with body sizes of transplanted mussels of well-defined biometric features.
228
D. MARTIN~I(~ ET AL.
Trace metals such as zinc, cadmium, lead and copper were metabolized with different rates during the period of 270 days. Mussels from the sites located in the vicinity of the Sibenik harbour tended to accumulate significantly high quantities of zinc and lead from water. Water concentrations of bioavailable metals which exceeded 500 ng Zn 1-1 and 20 ng l -~ Pb provoked a significant increase in the metal burdens of mussels. A significant correlation between body lead concentrations and lead present in the surrounding water was obtained, while no significant relationship was observed for zinc. The metal fluctuation caused by the presence of cadmium and copper in the water phase had no significant influence on the baseline metal concentrations in the flesh of transplanted mussels. Based on biological similarities between the individuals of the transplanted population, their higher degree of bio-homogeneity, and better reproducibility of the results of metal accumulation, one can conclude that utilizing transplants instead of indigenous mussels, is more useful for monitoring of dissolved metals in t h e water phase. ACKNOWLEDGEMENTS Financial support by the International Bureau of the K F A Juelich, within the framework of the bilateral agreement between Germany and SFR Yugoslavia on the joint project 'Environmental Research in Aquatic Systems' is gratefully acknowledged. In addition, this study was partly supported by the U.S. National Bureau of Standards and the Research Council of SR Croatia, through funds made available to the U.S.-Yugoslav Joint Board on Scientific and Technical Cooperation under the project JFP (NSB)/649 as well as E U R E K A No. 493 E U R O M A R - E L A N I Project 'Development of Electroanalytical Instrumentation for Physico-Chemical Characterization of Trace Metals in the Marine Environment'. We thank Moira Spanovi~ for correcting the English and for typing the manuscript. REFERENCES Amiard-Triquet, C., B. Berthet, C. Metayer and J.C. Amiard, 1986. Contribution to the ecotoxicologicalstudy of cadmium,copper and zinc in the musselMytilus edulis. II. Experimental study. Mar. Biol., 92: 7-13. Branica, M., 7.. Peharec, 7.. Kwokal and S. Kozar, 1985. Trace metals in the Sibenik aquatorium: P-1 Concentrations of Zn, Cd, Pb and Cu analysed in the 1983/84 period. Rap. Comm. Int. Met Medit., 29: lll-ll3. Cauwet, G., 1989. Distribution and fate of organic carbon in a stratified estuary (Krka, Yugoslavia). Presented at First International Symposiumof Small Estuaries, May 21-27, 1989, Primogten,Yugoslavia, p. 40.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
229
Cossa, D., E. Bourget, D. Pouliot, J. Piuze and J.P. Chanut, 1980. Geographical and seasonal variations in the relationship between trace metal content and body weight in Mytilus edulis. Mar. Biol., 58: 7-14. Fischer, H., 1983. Shell weight as an independent variable in relation to cadmium content of molluscs. Mar. Ecol. Prog. Ser., 12: 59-73. Goldberg, E.D., V.T. Bowen, J.W. Farrington, G. Harvey, J.H. Martin, P.L. Parker, R.W. Riseborough, W. Robertson, E. Schneider and E. Gamble, 1978. The mussel watch. Environ. Conserv., 5: 101-125. Kniewald, G., Z. Kwokal and M. Branica, 1987. Marine sampling by scuba diving. 3. Sampling procedures for measurements of mercury concentrations in estuarine waters and seawater. Mar. Chem., 22: 343-352. Lyngby, J.E. and H. Brix, 1987. Monitoring of heavy metal contamination in the Lim Fjord, Denmark, using biological indicators and sediment. Sci. Total Environ., 64: 239-252. Majori, L., G. Nedoclan, G.B. Modunutti and F. Daris, 1978. Study of the seasonal variations of some trace metals in the tissues of Mytilus galloprovincialis. Rev. Int. Oceanogr. Med., 49" 37-40. Margu~, D., 1983. Possibility of culturing the mussel Mytilus galloprovincialis Lmk., in the Krka River Estuary. M. Sc. Thesis, University of Zagreb, p. 81 (in Croatian). Margug, D., 1985. Comparative methods for calculation of index of condition in mussels (Mytilus galloprovincialis, Link.). Ichthyologia, 17: 59-67. Martin, N., G. Ichikawa, J. Goetzl, M. de los Reyes and M.D. Stephenson, 1984. Relationship between physiological stress and trace toxic substances in the mussels Mytilus edulis from San Francisco Bay, California. Mar. Environ. Res., l 1:91-110. Martin~i~, D., H.W. Niirnberg, M. Stoeppler and M. Branica, 1980. Toxic metal levels in bivalves and their ambient water from the Lim Channel. Thalassia Jugosl., 16: 297-315. Martin~i~, D., H.W. Niirnberg, M. Stoeppler and M. Branica, 1984. Bioaccumulation of heavy metals by bivalves from Lim Fjord (North Adriatic Sea). Mar. Biol., 81: 177-188. Martin~i~, D., H.W. Niirnberg and M. Branica, 1986. Bioaccumulation of heavy metals by bivalves from Limski Kanal (North Adriatic Sea). II. Cu distribution between oysters, Ostrea edulis and ambient water. Mar. Chem., 18: 299-319. Martin~ig, D., H.W. Niirnberg and M. Branica, 1987a. Bioaccumulation of heavy metals by bivalves from Limski Kanal (North Adriatic Sea). III. Cu distribution between mussels, Mytilus galloprovincialis, and ambient water. Sci. Total Environ., 60: 121-142. Martin~ig, D., M. Stoeppler and M. Branica, 1987b. Bioaccumulation of metals by bivalves from the Limski Kanal (North Adriatic Sea). IV. Zinc distribution between Mytilus galloprovincialis, Ostrea edulis, and ambient water. Sci. Total Environ., 60: 143-172. Martin~ig, D., M. Stoeppler, Z. Kwokal and M. Branica, 1987c. Trace metals in selected organisms from the Adriatic Sea. Mar. Chem., 22: 207-220. Martin~ig, D., M. Branica, Z. Kwokal and Z. Peharec, 1988. In: Long-term Research and Pollution Monitoring of the Krka Estuary and Kornati Archipelago (Adriatic Sea), MED POL PHASE II (Center for Marine Research, 'Rudjer Bo~ovi~' Institute, Zagreb), Annual Reports, 1983-1987. MED POL PHASE II, 1989. Long-term Research and Pollution Monitoring of the Krka Estuary and Kornati Archipelago (Adriatic Sea), Center for Marine Research, 'Rudjer Bogkovig' Institute, Zagreb, Annual Reports 1988. Popham, J.D. and J.H. D'Auria, 1982. Effects of season and seawater concentrations on trace metal concentrations in organs of Mytilus edulis. Arch. Environ. Contain. Toxicol., 1l" 273-282. Phillips, D.J.H., 1976. The common mussel Mytilus edulis as an indicator of pollution by zinc,
230
D. MARTIN(~I(~ ET AL.
cadmium, lead and copper. I. Effects of environmental variables on uptake of metals. Mar. Biol., 38: 59-69. Olafsson, J., 1979. Trace metals in mussels (Mytilus edulis) from southwest Iceland. Mar. Biol., 90: 74-78. Schulz-Baldes, M., 1973. Die Miesmuchel Mytilus edulis als Indicator fur die Bleikonzentration in Weserastuar und in der deutschen Bucht. Mar. Biol., 21: 98-102. Simpson, R.D., 1979. Uptake and loss of zinc and lead by mussels (Mytilus edulis) and relationship with body weight and reproductive cycle. Mar. Pollut. Bull., 10: 74-78. Stephensen, M.D., R.M. Gordon, and J.H. Martin, 1979. Biological monitoring of trace metals in the marine environment with transplanted oysters and mussels. In: J.H. Martin (Ed.), Bioaccumulation of Heavy Metals by Littoral and Pelagic Marine Organisms, EPA-600/3-79-038, U.S. Department of Commerce, National Technical Information Service, PB-279-493, pp. 12-24. Widdows, J., D.K. Phelps and W. Galloway, 1980-81. Measurements of physiological condition of mussels transplanted along a pollution gradient in Narragansett Bay. Mar. Environ. Res., 4: 181-194. Widdows, J., P. Donkin, P.N. Salkeld, J.J. Cleary, D.M. Lowe, S.V. Evans and P.E. Thomson, 1984. Relative importance of environmental factors in determining physiological differences between two populations of mussels (Mytilus edulis). Mar. Ecol. Prog. Ser., 17: 33-47.
211
Distribution of Zn, Pb, Cd and Cu between seawater and transplanted mussels (Mytilus galloprovincialis) D. Martin~i6 a, Z. Kwokal a, 7,. Peharec a, D. Margug a and M. Branica ~'b aCenter for Marine Research Zagreb, 'Ruder Bogkovik' Institute, POB 1016, 41001 Zagreb, Croatia bInstitute of Applied Physical Chemistry, ICH-4, KFA, Juelich, POB 1913, D-5170 Juelich, Germany (Received June 28th, 1991; accepted July 22nd, 1991)
ABSTRACT Cultured one-year old mussels, Mytilus galloprovincialis, (Lmk.) were transplanted to four sites (120 individuals per site) throughout the Krka River Estuary, and nearby Adriatic coastal area. Concentrations of zinc, cadmium, lead and copper, and their content in mussels were analysed four times during 1988/89 (270 days). The body weight of mussels increased during the period studied and a discernible effect of the size of mussels on metal concentration was detected. The concentrations of the metals studied, expressed on a per unit dry weight, generally decreased with the body size and the age of mussels, but no seasonal trends could be established. The mussels transplanted to sites situated in the vicinity of the Sibenik harbour accumulated zinc and lead. This could be attributed to elevated concentrations of zinc and lead in water (higher than 500 ng Zn 1-~ and 14 ng Pb 1-I of 'bioavailable' forms). Significant correlation between body lead concentrations and lead amount in the surrounding water was obtained, while for zinc, no statistically significant relationship exists between body concentrations and the water phase. The fluctuations of cadmium and copper in the adjacent water have no significant effect on the changes of their concentrations in the transplanted mussels. The data obtained indicate that because of biological similarities and better reproducibility of the results the transplanted mussels are more suitable for monitoring of trace metals than indigenous mussel populations as originally proposed by the 'Mussel Watch' programme.
Key words: sea water; mussels; transplantation; trace metal concentration INTRODUCTION
Mussels have been utilized as valuable sentinel organisms for indicating levels of trace metal concentrations in marine environment because of their widespread distribution along coastal zones (Schulz-Baldes, 1973; Godlberg et al., 1978; Majori et al., 1978). Trace metal concentrations in mussels can also be affected both by biological and physical factors, such as: reproductive 004~-9697/92/$05.00
© 1992 Elsevier Science Publishers B.V. All rights reserved
212
o. MARTIN~I(~ETAL
state, sex, and biological condition of an individual mussel, season of sampling, temperature, salinity and a depth of organisms habitat. The effect of these variables on the metal content in tissues has not been completely clarified. Their effect is evident from a large variation in metal concentrations described by Martin~i6 et al. (1987c). The objectives of the long-term study undertaken (from 1983 to 1988) by these authors were to recognize and quantify influence of slightly polluted environment, with regard to trace metals present in the water phase (Branica et al., 1985), on metal concentration in indigenous mussel populations from various parts of the Krka River Estuary, Yugoslavia and nearby coastal area (Martin~i6 et al. (1985; 1986; 1987c). However, they found that natural populations of mussels, Mytilus galloprovincialis, often exhibit considerable natural variability in terms of trace metal levels which complicate attempts to detect changes resulting from anthropogenic sources. Mussels were sampled according to monitoring programme formulated by the UNEP Coordinated Mediterranean Pollution Monitoring and Research Programme (MED POL I, 1980). Moreover, it has been observed that, among all causative biological factors, ages, reproductive states, and habitats of these populations were different. For example, natural habitats of mussels differed significantly throughout the Krka River Estuary, depending on salinity and water depth. It is desirable and often possible, to eliminate factors that may confuse the interpretation of a monitoring survey. Therefore, the objective of this work is to reduce the causative biological factors which have been studied so far for indigenous mussel populations, and improve the homogeneity of the response in the test environments by transplanting mussels from the shell farm to selected sites. It has already been observed that the transplantation of mussels or similar organisms to various areas may provide an useful approach to marine environmental monitoring (Widdows et al., 1980-81; Widdows et al., 1984). Bioaccumulation of dissolved trace metals in the organisms is usually observed when they are moved from an uncontaminated to a contaminated area (Simpson, 1979; Stephensen et al., 1979; Martin et al., 1984). SAMPLING AND SAMPLE PRETREATMENT
Sampling area The Krka River Estuary situated in the eastern Adriatic coast is characterized by a low terrigenuous input of trace metals owing to a karstic drainage area and a series of travertine barriers which precede the estuary. A mean freshwater inflow of the Krka River is approximately 55 m 3 s -1 with considerable monthly variations ranging from 5 to 410 m 3 s -I. Apart from that, torrents and submarine springs occasionally contribute minor inflows. The Krka River Estuary is a stratified estuary characterized by a low
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
213
tidal range (20-50 cm) and a sheltered position. The brackish surface water current is directed seawards, whereas bottom seawater counter-current can be traced up to the Skradinski Buk waterfalls (Fig. 1). The area investigated is confined to the part of the ancient river valley between the last active calc tufa barrier, through a central enlarged part, and the Sibenik Channel for a total length of 22 km. The estuary gradually deepens from 2 m (at the waterfalls), to 42 m on the seaward side (the Sibenik Channel). The city of Sibenik (40 000 inhabitants) situated in the lower part of the estuary, with its shipping and metal industry, represents a hazard to aquatic life by directly discharging wastes into the estuary. Mussels can only survive in the water of salinity of 10%o or higher (Margug, 1983), which means at a depth of 5-6 m (station E-2 in the vicinity of Skradin, Fig. 1). In the Sibenik harbour (the lower part of the estuary) the mussels natural habitats are closer to the water surface and under possible tidal influence.
Hydrologic and chemical features of the estuary Multidisciplinary investigations of the Krka River Estuary have been carried out by the Centre for Marine Research since 1980, and form part of the long-term study MED POL Phase II (Annual reports, 1983-1989). During the spring of 1988 a mean freshwater inflow of the Krka River was approximately 58 m a s -l. It was greatly reduced to only 5 m 3 s -~ in the dry period (summer and autumn). Water temperature was measured in situ at a depth of 6 m and gradually increased from May (17°C) to August (22°C). In December water cooled down to 15°C. Salinity, measured conductometrically, varied between S = 37 and 39%0. Total suspended matter (SM), particulate (POC) and dissolved (DOC) organic carbon were measured in water samples collected at a depth of 6 m on May 15th, August 5th, September 4th and December 4th, 1988. Prior to measurements filtration of samples was performed on Whatman GF/F glass-fibre filters. Most of the time the river water is an optically very clear organic-poor medium (Cauwet, 1989). During the summer and autumn of 1988 concentrations of dissolved organic carbon (DOC) were generally < 0.9 mg 1-~ at all sampling sites with the exception of site E-2 where values exceeding 1 mg 1-I were obtained. This may be ascribed to resuspension since the sampling was performed close to the sediment surface. A significant difference in the concentrations of DOC was noted between water samples collected from the coastal site C-I (0.55-0.65 mg C Iq) and those from estuarine sites. High planktonic productivity which occurred in May 1988 was responsible for elevated concentrations of DOC at all sampling sites including the site C-1 (1.20 mg C 1-~). Particulate organic carbon (POC) concentrations varied between 0.168
214
D. MARTINI~I~ El" AL,
and 0.334 mg C 1-1 at all sampling sites. No significant difference in POC values was observed between spring and summer, 1988. Data obtained during the period 1988-1989 indicate that concentrations of suspended matter in the Krka River Estuary are lower than in bther estuaries (Martinic, 1988). The concentrations of suspended matter varied between 0.70 and 1.87 mg 1-1 Higher amount (5.43 mg 1-1) of suspended matter was only observed in May at the site E-4. Oxygen concentrations in the saline water layer were constant (5-6 ml 1-1) during the entire period with the exception of site E-2 in December when a minimum oxygen concentration of 3.7 ml 1-1 was obtained.
Transplanted mussels One-year old cultured mussels Mytilus galloprovincialis (Lmk.), (480 individuals), with well-defined biometric features, cultivated in the fishery and shell farm Sarina Draga (central part of the Krka River Estuary), were used as the starting stock. Average shell-lengths varied slightly (between 60.2 and 63.4 mm), with variation coefficients between 8 and 15%, (Table 1). The mussels were placed into oval plastic cages and moored in water of practically constant salinity (37-38% o), at a depth of 6 m, at four different locations (120 individuals each) (denoted E-2, E-4, E-5 and C-I, on Fig. 1). Since the depth of the estuary gradually increases from 8 m (at station E-2) to 42 m (stations E-5 and C-I), living depths for indigenous mussels differ along the estuary. The mussels were kept at four sites from April 24, 1988 to January 14, 1989 (266 days). The results published by Widdows et al. (1980-81) suggest that one month of exposure to different environmental conditions is sufficient to allow the equilibration with new environmental concentrations within the animals tissues, as well as for physiological adjustment to new living conditions. A month of acclimatization was allowed prior to sampling. Sampling and measurements of 25 individuals separated into four groups were performed periodically: June 1st, July 20th and September 30th, 1988 and January 14th, 1989. Sampling procedure was carefully observed in order to avoid contamination problems (Martin~i6 et al., 1980; 1984). Prior to analysis, no gutcleansing of the collected specimens was performed. Ten individuals from each group were used for biometric characterization, and only soft tissues of the remaining 15 specimens were homogenized and wet-digested prior to analyses for metal concentrations using ETAAS and FLAME-AAS (Martin~i6 et al., 1987c). The bivalve condition index was determined according to the Hopkins method (Margug, 1985), (the ratio between dry flesh and total weight of mussels).
0.102 0.151 0.178
E.4 b
stock stock stock stock
0.077 0.090 0.107 0.137
C.1 d
aThe starting bThe starting CThe starting dThe starting
0.095 0.125 0.164 0.158
E.5 c
of of of of
0.138 0.176 0.234 0.146
E_2 a
0.234
Dry/wet factor
Location
(1.2) (l.0) (1.3) (1.3)
(1.2) (2.0) (2.1) (2.1)
(1.5) (1.9) (1.5) (3.6)
(1.6) (3.1) (2.2) (2.7)
121 individuals 120 individuals 120 individuals 120 individuals
3.8 4.3 4.9 6.0
4.8 5.6 7.1 6.8
4.8 6.6 7.5 9.7
6.2 7.6 9.4 6.4
Wet weight in g (S.D.)
of of of of
(9) (6) (8) (7)
(8) (6) (7) (4)
(6) (8) (7) (6)
(5) (6) (6) (5)
(6) (8) (7) (7)
60 63 63 62
62 65 65 69
67 66 70 72
66 69 69 75
67 71 70 66
mm mm mm mm
Shell length in mm (S.D.)
in in in in
length length length length
was was was was
June July September January
June July September January
June July September January
June July September January
Month of collection
settled settled settled settled
14.6 15.0 19.2 21.8
22.0 29.7 23.7 30.4
17.8 20.2 26.0 29.1
23.0 23.0 27.0 39.3
in in in in
April. April April. April.
(0.3) (0.2) (1.0) (1.2)
(0.4) (1.3) (2.0) (1.0)
(0.4) (1.4) (0.3) (0.1)
(0.1) (0.8) (0.1) (0.1)
Zn (S.D.)
0.17 0.17 0.16 0.32
0.17 0.16 0.14 0.30
0.17 0.15 0.14 0.18
0.15 0.14 0.12 0.29
(0.01) (0.01) (0.01) (0.02)
(0.01) (0.01) (0.01) (0.01)
(0.01) (0.01) (0.01) (0.01)
(0.01) (0.01) (0.01) (0.01)
Cd (S.D.)
Me #g g-l wet weight
Data for transplanted mussels collected from various locations in the Krka River Estuary
TABLE 1
0.24 0.23 0.22 0.27
0.34 0.38 0.24 0.36
0.42 0.38 0.33 0.30
0.16 0.12 0.11 0.17
(0.01) (0.02) (0.02) (0.03)
(0.01) (0.03) (0.01) (0.02)
(0.01) (0.03) (0.03) (0.01)
(0.01) (0.01) (0.01) (0.01)
Pb (S.D.)
0.72 0.76 0.84 0.87
0.73 0.81 0.92 0.89
0.80 0.87 1.02 1.11
0.88 0.98 1.03 0.93
(0.06) (0.01) (0.01) (0.03)
(0.06) (0.01) (0.02) (0.03)
(0.04) (0.01) (0.02) (0.03)
(0.02) (0.06) (0.04) (0.03)
Cu (S.D.)
t,~
7~
Z ~7
7
216
D. MARTIN(~I(~ET AL,
Ambient water
Water samples for metal analyses were collected from a depth of 6 m by divers (Kniewald et al., 1987) on May 14th, July 29th, 1988 and January 15th, 1989. Prior to sampling pre-cleaned 1 1 Nalgene sampling bottles were thoroughly washed with diluted Suprapure HNO3. They were also preconditioned in the sea for 7 days. The bottles were protected against fouling by enclosing them in a plankton net (150/~m pore size) and fixed to the seafloor at a depth of 40 m. The top of the plankton net was fixed to an air buoy, so that the entire system floated - 2 0 m above the bottom. The bottles were left unstoppered so that seawater could rinse them. Prior to sampling, the bottles were again treated with acid (hot 10% Suprapure HNO3 in order to remove organic films formed on the bottles. Finally, the bottles were washed with tetra-distilled water. Electrochemical characterization of Zn, Cd, Pb and Cu was performed within 12 h of sampling. Differential pulse anodic stripping voltammetry (glassy carbon and hanging mercury drop electrodes) was used for determination of 'total' and 'free' dissolved metal concentrations in unfiltered water samples at pH 2 and 8 (4.7 and 8 for zinc). The instrumentation used was a PAR Polarographic Analyzer, Model 174A with an X-Y recorder, as described in detail elsewhere (M. Branica et al., 1985). RESULTS Dissolved and particulate metal concentration in water
Our results show (Table 2) that trace metal concentrations in the Krka River Estuary are significantly lower than the values obtained for estuaries throughout the world (Martinrir, MED POL Phase II, Annual Report, 1983-1987). There appears to be a number of reasons for this. Firstly, a relatively unpolluted catchment area provides effective baseline trace metal values for the Krka River feeding the Adriatic Sea. Secondly, a solid-liquid exchanges are limited by a low particulate loading (0.7-5.43 mg 1-Z) in the river water. Water samples collected from sampling sites E-4 and E-5 (in the vicinity of an industrial waste outflow of the city of Sibenik), were found to be enriched with zinc, lead and copper. However, the enrichment through anthropogenic sources is limited, because average 'total' metal concentration values decreased seawards reaching lower average values at the coastal site C-1 (520 ng Zn 1-1, 11 ng Cd -z, 34 ng Pb 1-Z, and 180 ng Cu 1-1). Uniform distribution of lead was observed in the entire water column at sites E-4 and E-5, while distribution of Zn, Cd and Cu differed significantly. Surface layer
May July January
May July January
May July January
May July January
E-2
E-4
E-5
C-I
Location Month of collection
420 530 670
490 820
690 350 --
1100 340 2110
.
.
320 260 --
720 150 300
210 430 440
240 620
.
370 90 --
380 190 1810
210 100 230
250 200
.
19 8 5
8 10 5
11 10 26
8 40
.
16 5 2
6 2 3
5 6 11
3 17
.
pH8
pH2
pH 4.8-8
pH4.8
pH8
Cd (S.D. < 2%)
Zn (S.D. < 5%)
3 3 3
4 8 2
6 4 15
5 23
.
pH2-8
.
65 22 15
61 49 35
89 53 27
23 11
pH2
.
51 10 4
18 13 7
11 13 6
5 3
14 12 11
43 36 28
78 40 21
. 18 8
pH8 pH2-8
Pb (S.D. < 3%)
.
220 120 190
160 290 160
596 483 260
340 230
pH2
.
60 20 60
30 30 50
70 97 57
12 37
pH8
Cu (S.D. < 2%)
160 100 130
130 260 110
526 386 203
328 173
pH2-8
Metal concentration (ng 1-i) in the unfiltered seawater samples collected at a depth of 6 m in the surrounding water of transplants
TABLE 2
-Zt r~
z
..-I >
> Z
-I
>.
> Z
218
o. MARTINI~I~ ET AL.
(0.5 m) at sites E-4 and E-5 contained higher concentrations of zinc and cadmium than deeper water (6 m), while copper concentrations at site E-4 were inverse. However, an average 'total' metal distribution in the water column at site E-2 differed from sites E-4, E-5 and C-1. The water samples collected at a depth of 6 m contained about two-fold higher average zinc concentration (660 ng Zn 1-1) than the surface water samples (310 ng Zn l-l). Concentrations of copper at a depth of 6 m (205 ng Cu -1) and at the surface (212 ng Cu l -l) are practically the same. This could be explained by the fact that a water mass at this depth is normally exposed to mineralized organic material remobilizing metals from adjacent sediments. The bottom depth of water at this sampling site is only 7-8 m. The data obtained through annual monitoring of the Krka River Estuary support this opinion. The water at a depth of 6 m contained significantly higher quantities of suspended matter than the surface water layer with the exception of sites E-4, E-5 and C-1 in December 1988. Dissolved and particulate metal concentrations varied through the year, mainly depending on the freshwater inflow. For example, in May 1988 a normal plankton bloom, caused by higher nutrients was accompanied by higher POC values, particularly at site E-4. This resulted in maximum concentration of zinc (850 ng Zn 1-1) and cadmium (30 ng Cd 1-1) in the surface water. It seems that lead and copper exhibited seasonal variations, reaching maximal values in August. Surface water from sites E-4 and E-5 (vicinity of the city of Sibenik) was found to be more enriched with lead and copper than deeper water indicating enrichment through anthropogenic sources (MED POL PHASE II, Annual Report, 1988).
Transplanted mussel population Mussels condition Cultured one-year old mussels were transplanted from the shellfish farm Sarina Draga (Fig. 1) to the Krka River Estuary. Prior to transplantation, 80% of individuals had already spawned, with an average condition index of 5.9. The somatic growth and induction of reproductive cycle (gametogenesis) are evident from the condition indexes, calculated by the Hopkins method and illustrated in Fig. 2. Mussels transplanted to site E-2 reached the highest condition indexes as compared with the mussels transplanted to other sites. Condition indexes of the mussels from four stations are illustrated in Fig. 2.
Biometric analysis The biometric data are summarized in Table 1 and illustrated in Fig. 3. A
219
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
15°45
-~(f 16°00'
Skrad Prokljan take nski Buk r falls)
arina Draoa.
.SIBENIK , Sibenik bay
~b
o2
4
Fig. 1. Map of the investigated area.
I0-
• E-2
9-
o E-4
.o
o 6-
initial musse[s
~ ^
~m,~
x~
~32
monfh M year days of exposure
' A ' M ' J' 1988 J' A' S' O' N' D' J'I~89 0
3B
8B
160
266
Fig. 2. Index of conditions (calculated by the Hopkins method) for transplanted mussels during 270 days of exposure at different sites in the Krka River Estuary.
220
D. MARTIN(~I(~ ET A L
b
108-
10-
-,-
i6:
4-
/ "
b=1,29 df= 1/, ' ~ '~'lb
b=0,96 df=l/, '
2,
;
' &'~'16
shelf weight(g)
meat v01ume(ml)
20
~lOt=n
-
oJ
6-
r =0,999 b =0,/,1 df=lt,
3=
I
I
0,2
l
d
20
t E°J =~ o .~, 10
O,I/+
I
I
I
I
I
I
I
0,6 0,8 1,0
2,0 dry weight(g)
r
4,0
• June o July z~ September ~ a n n u a r y
~ . .
~1 ~ +-~
'-t
:il ,=0,,+
\ ^ z.~
b= - 0,62 df = 1L,
I
0,2
i
I
0,/*
I
I
I
I
'
I
0,6 0,8 1,0
21
,0 dry weight(g)
I
4,0
Fig. 3. The biometric features of transplanted mussel populations collected in June, July, September 1988, and January 1989 at different sites in the Krka River Estuary (df, degree of freedom).
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
221
satisfactory correlation between shell weight and wet weight of the mussels was obtained (r = 0.944, d f = 14). This indicates that during the period investigated the changes in mussels body weight are mainly the result of their somatic growth (Fig. 3a). Flesh volume of all mussels studied varied proportionally (b = 0.96) with their wet weights (Fig. 3b) owing to the constant salinity (37%0) in the environment. A satisfactory correlation between dry and wet weights of mussels was also obtained (Fig. 3c). The ratio between the flesh volume and dry weight decreased with the increase of mussel dry weights (Fig. 3d) indicating different influence of the somatic growth of the mussels on the body increment expressed on the wet or dry units basis. In the case when the somatic growth has a priority significance over the reproductive cycle, the wet body increment is less pronounced than the increment of dry organic substances. Trace metal concentration in mussels Metal concentrations in mussels flesh varied significantly, depending on their living conditions, induced bio-rhythm and physico-chemical properties of surrounding waters. The data are summarized in Table 1. The metal concentrations in mussels are satisfactorily comparable with already published values for indigenous mussels from the same area (Martin6i6 et al., 1987). The zinc concentrations varied significantly between 14.6 and 39.3 #g Zn g-l wet weight. Mussels from the coastal site (C-l) show significantly lower zinc and copper concentrations (17.7 #g g-i and 0.80 tag g-i, respectively) than mussels from site E-2, where the highest values were obtained (28 tag Zn g-l and 0.95 tag Cu g-1 wet weight). The values obtained for cadmium are practically the same (0.18 and 0.20 tag Cd g-l). Concentrations of lead in mussels is reversed at these locations. The lead concentrations in mussels from site E-2 are lower (0.14 #g Pb g-l) than in mussels from site C-1 (0.24 tag Pb g-l). Average metal concentrations obtained suggest that the mussel body condition factors had pronounced effect on metal concentrations in mussels. Expressing the values on the wet body weight basis it becomes obvious that mussels accumulated zinc and copper during the tissue growth, while at the same time, average cadmium and lead concentrations decreased. The average values obtained for zinc and lead concentrations in mussels at sites near areas of a pollutant discharge (E-4 and E-5) are significantly higher than the concentrations obtained at other sites. In August 1988, the highest zinc concentrations (29/~g g-l) were found in the mussels from site E-5. The mussels from sites E-4 and E-5 were found to be enriched with lead of most probably anthropogenic origin, and contained two or three-fold
222
D. MARTIN~I~ ET AL
higher lead concentrations than mussels from sites E-2 (0.14 #g Pb g-l) and C-1 (0.24 #g Pb g-l). No enrichment in Cu and Cd which could exceed the calculated 'background' levels in mussels was observed. DISCUSSION Since metal concentrations in mussels from unpolluted environments vary widely (Martin~i6 et al., 1987c), only differences several orders of magnitude larger are considered to be significant (Goldberg et al., 1978). It appears that applicability of mussels as indicators of trace metal concentration levels refers only to significantly contaminated environments. Our previous studies show that the variations in trace metal concentrations of mussels are generally related to local living conditions and seasonal weight fluctuations (MartinEi6 et al., 1980; 1984; 1986; 1987a,b,c,). The results of a systematic study of the Krka River Estuary and the Kornati Islands carried out from 1983 to 1988, show almost uniform average zinc concentration values, while cadmium, lead and copper distributions differ (Martin~i6 et al., 1987c). Moreover, the mussels from the Kornati Islands (the reference area) contained significantly higher cadmium concentrations than might be expected from the area free from anthropogenic influence (Branica et al., 1985). It has been observed that the mussels from the areas of pollutant discharge (vicinity of the Sibenik harbour) contained higher lead and copper concentrations than the mussels from the Kornati Islands owing to the presence of higher metal concentrations in the water phase. However, these relations were not statistically proved since differences in metal concentrations as well as various biological factors, such as the mussels size, living conditions, season of sampling, etc., influence the uptake of trace metals in indigenous mussels in such a way that the metal concentrations in mussels from the slightly polluted site can be similar to those in specimens from an unpolluted site. Therefore, it was not possible to distinguish between slightly polluted and unpolluted areas. Indigenous populations of mussels sampled from different parts of the Krka River Estuary and the Kornati Islands could not be biologically compared. The average values of the biometric measurements of mussels transplanted from the shellfish farm Sarina Draga to the Krka River Estuary are summarized in Table 1. The transplants were distributed throughout the area at a depth of 6 m which guarantees marine living conditions. Condition indexes of the transplanted mussels depend on their local living conditions and season of sample collection, which is satisfactorily comparable with findings of Margug (1983). The mussels from the site E-2 in the upper part of the estuary, living near
Zn, Pb, Cd AND Cu 1N SEAWATER AND TRANSPLANTED MUSSELS
223
sediment surface, tended to have the highest nutritional state (Fig. 2) owing to the presence of larger quantities of suspended material, rich in POC. The average indexes of condition of the mussels from other sites gradually decreased in the seaward direction. The transplants living in the coastal environment, relatively poor in suspended matter, had the lowest indexes of condition. A maximum observed in June (Fig. 2) in the mussels from the site E-4 coincided with the phytoplankton bloom which usually occurs during this period of the year. The mussels from all studied sites reached the lowest indexes of condition in January 1989 owing to the spawning in December 1988. It has also become evident that the shell lengths of the mussels studied did not necessarily increase with age which can successfully be compared with earlier observations (Martin~i6 et al., 1986; 1987a,b). However, shell weights changed proportionally with the aging of mussels (Fischer, 1983). A significant (P < 0.01) regression line with a slope ofb = 1.29 (Fig. 3a) was obtained by relating log average wet weight to log average shell weight of transplants suggesting that the mussels somatic growth was primarily responsible for the body weight fluctuations during the entire period. Wet weights of mussels from all locations are directly proportional to their flesh volumes (Fig. 3b) (b = 0.96 with 1% probability level). Furthermore, the ratio between wet weights of mussels and water content in their bodies was fairly constant over the entire period owing to stable salinity values in the aquatic environment. Relation between wet and dry weights, shows different ratios for different mussel sizes (Table 1, Fig. 3d). The water concentration in the mussels decreases thereby increasing their dry weight, size and age. Concentrations of all metals in all samples collected from four experimental sites were fitted with regressions of log weight-log concentration of significant non-zero slopes. Our results indicate that larger mussels contained higher concentrations of zinc and copper than smaller ones (metal concentrations are expressed on a per unit wet weight). Accumulation of these metals depends on the mussels somatic growth, thereafter on the water content (Figs. 4a, d). Furthermore, different slopes of regression lines obtained for zinc and copper concentrations indicate that mussels metabolize these metals at different rates (Figs. 4a,b,d,e). On the other hand, zinc and copper concentrations expressed as a per unit dry weight exhibited the reverse association in comparison with concentrations based on a per unit wet weight. Their concentrations decrease as the body water content increases. The slopes obtained (Figs. 4c, f) indicate that larger mussels contain lower metal concentrations than smaller ones, which is in a good agreement with the results obtained by Cossa et al. (1980). Moreover, from the above relations it is evident that some metal concen-
224
D. MARTIN(~IC El" AL.
60a
Zn
~0-
2,0.
aE-2 /
d
Cu
3~ c
N 20
= 0,8-
r=0,872 b= 0,93 df=11 '
~
~ 'lb
~0,6-
b=0,58 df= I~,
0,4-
2b
~, ' b ' l b
'
shell weight(g )
shell weight(g)
6O
b
Zn
"G ~0-
2,0
oE-2
4--:
e
Cu
:" 30-
%
1,0-
~= o,a-
o, 20-
jo
;@3
~.o,6-
df=lO
10
b=0,4/+ dr= 11+
O,t,-
'
wet weight(g)
4
' ~'~'Ib
wet weight(g)
I+00
c
Zn oE-5
201
oE-2
o ~ ' ~ t _ ~
2O
hE'5
f
Cu
r =-0,9B7 b=- 0,63
10-
8Z
-.: 80-
20
0,2
16-
r= - 0,937 b=- 0,23 df= 10
-o 60-
i
i
0,~
f
°s, -
i
i
i
v 1,0
0,6 0,8 1
i
2,0 dry weighttg)
i
2't 0,2
i
i
0,~
i
t
i
i
i
i
0,6 0,8 1,0
i
2.0
dry weight(g)
Fig. 4. Zn and Cu concentrations in the transplanted mussel populations collected in June, July, September 1988, and January 1989 at different sites related to the mussel biometric features.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
225
trations deviate from the calculated ideal line, i.e. baseline concentrations of the mussels. This suggests that besides tissue weights or shell sizes of the mussels there are other factors influencing the metal body concentrations. For example, the results obtained indicate that zinc concentration in mussels (E-5 in June, July, 1988 and January, 1989) does not overlap calculated regression lines (Figs. 4a,b,c). Comparing the zinc concentrations in the mussels from these periods with the zinc concentrations in the water phase, it becomes obvious that such deviations could only be attributed to the elevated metal concentrations in the adjacent water (Table 2), although no significant correlation was obtained when relating body zinc concentrations to the zinc concentrations in the water phase. Figure 4c and Table 2 show that amounts of zinc in mussels from site E-5 (June, July 1988 and January 1989) coincide with particulate and strongly complexed zinc concentrations (pH 4.8-8) (portion of 'bioavailable' metal) which exceed 500 ng Zn 1-1 in water phase. In the study carried out by Olafsson (1986), the metal concentrations of the juvenile mussels which had colonized the mooring structures were similar to the post-adaptation concentrations of the caged mussels, except for Hg and Zn. The concentrations of these metals were significantly lower in the juvenile mussels than the mean post-adaptation concentrations in the caged mussels. He suggests that a proportion of this initial tissue content of these metals is firmly bound in the caged mussels and therefore he proposes that mussels grown from a juvenile stage at monitoring sites could be more sensitive indicators of seawater metal concentrations than mussels transferred from a coastal site. Copper concentrations in mussels depend on body sizes only, regardless of the fact that the animals were exposed to significantly higher quantities of particulate and strongly complexed copper (pH 2 - p H 8) in surrounding waters. The highest copper concentrations in water were found in the vicinity of the Sibenik harbour (sites E-4 and E-5) (Table 2), but in spite of that no enrichment in copper which could exceed the baseline metal concentrations of mussels was obtained. Phillips (1976) found that the net uptake of copper by mussels was extremely erratic and suggested that the mussels are not suitable indicators of copper pollution in the marine environment. The same conclusion was drawn by Lyngby and Brix (1987). The ability of mussels to maintain their internal copper and zinc compositions at a steady level when variations in the metal composition of the external medium occur, was observed by ArmiardTriquet et al. (1986). Therefore, they concluded that the use of mussels as bioindicators of pollution was doubtful for essential metals, particularly as regards short-term pollution, since the levels of these trace elements in the organisms were largely independent of their concentration in the ambient seawater.
D. MARTIN(~IC ET AL
226
The concentrations of lead in larger mussels are lower than in smaller ones. Lead concentrations in mussel tissues decreased proportionally with an increase of tissue, water content, size and age (Figs. 5b,c). Most of the values of the Pb tissue concentrations fell above the regression line when related to dry tissue weight. This is most probably the result of anthropogenic input. 0,6-
Pb
a
_ E-t~o E~E-5
-y 0,~-
\
.
?~E~ E-Z, mE-Z, xx~'1"E -5
\
% g_ 0,2 r:-0,951 b: - lJ+
r:-O,93t+ b=-0,51 df:lO
•
f:s-
O,T
131o o [] E-2 E'5
i
°X,,
o,1~ , ~,~,~b
shell weight(g)
~o
shell weiohtlg)
0,~
b
Pb
e
~: 0,4-
O,L,-
•
O (~]A
Cd n I:]Q
~0,,
8. o,:
r:-0,986
~N.
b:-0.96
"~
df:5
- N~,
O,'L
I
' -
I
oE-/, eE-5 oE-5
"~ N o
I
r:o0.996
0,2
,
0,4
,
,,I
o~ 0.8 1'o
f
2,0
E-Z,
~ 0,6" I
wet weight(g)
oC-1 _
0,~
0,21
L ' ~'~'lb
Cd
zxE-5
~
'
C
oFZ, oE5 qC-1 zxE-z,
~.
0,1
I
) B1 L 'w6te weioh~(g
Pb
Z,_ \
~
I
l
zo
dry weighf(g)
0,2
r :-0,998 b:- 0.79
"-~
E~Z, ~.
df:lO
'
0',z, ' 0',6' 0'8'{0,
2',0 dry weight(O)
Fig. 5. P b and Cd concentrations in the transplanted mussel populations collected in June, J u l y , S e p t e m b e r 1988, and January 1989 at different sites related to the mussel biometric
features.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
227
Indeed, particulate and complexed lead concentrations which exceed 14 ng Pb 1-~ of seawater provoke proportional increase in metal concentrations in tissues (Fig. 5c). Mobilization of Pb by human activities, particularly the combustion of leaded petrol, has caused significant increase of Pb concentrations in the upper layers of the estuary. An unknown proportion of this lead has been deposited on the streets and roads and is washed away into the estuarine water. Lead of this origin is most probably the primary cause of the frequently encountered elevated Pb concentrations in mussels from sites E-4 and E-5. Moreover, this has been confirmed by the fact that lead concentrations in the mussels which fell above the regression line are well correlated with total (r = 0.912, P < 0.001) and 'bioavailable' lead (r = 0.905, P < 0.05) in the surrounding water. The same relation was obtained by Popham and D'Auria (1982). Satisfactory correlation was found between cadmium concentrations in the soft part of tissues and shell weights, the Cd concentrations changing inversely with respect to shell weight. Cadmium concentrations in the soft parts decreased when the water content-increased, i.e., size of mussels, what is in a good agreement with the findings by Phillips (1976). A seasonal dependence was not observed. All transplanted mussels collected in January 1989 contained cadmium concentrations which exceeded baseline cadmium values. During this period, cadmium concentrations in the water phase were similar to concentrations found during other sampling periods, with the exception of site E-2. Particulate and strongly complexed cadmium was rather high at this site, so that the concentrations obtained in the mussels from this site could be attributed to this event, but not in the case of other sites. A possible explanation of this event can be found in the data on surfactant activities obtained by a routine monitoring of the area studied (MED POL PHASE II, Annual Reports, 1988 (1989)). Surfactant activities measured both directly and in the filtrated water samples collected throughout the area at the end of December 1988, showed that water was enriched with dissolved organic material to the extent which exceeds the concentrations found during the period between June and December 1988. Such a water component of a nutritive nature could stimulate the mussels to filter larger volumes of water, thus accumulating higher quantities of cadmium naturally present in the water phase. At any rate, the event observed in the case of cadmium and mercury (unpublished data) confuses the interpretation of all other values fitted by log/log regression of weight-concentration, and needs further field investigations. CONCLUSIONS
Concentrations of all metals studied changed significantly and inversely with body sizes of transplanted mussels of well-defined biometric features.
228
D. MARTIN~I(~ ET AL.
Trace metals such as zinc, cadmium, lead and copper were metabolized with different rates during the period of 270 days. Mussels from the sites located in the vicinity of the Sibenik harbour tended to accumulate significantly high quantities of zinc and lead from water. Water concentrations of bioavailable metals which exceeded 500 ng Zn 1-1 and 20 ng l -~ Pb provoked a significant increase in the metal burdens of mussels. A significant correlation between body lead concentrations and lead present in the surrounding water was obtained, while no significant relationship was observed for zinc. The metal fluctuation caused by the presence of cadmium and copper in the water phase had no significant influence on the baseline metal concentrations in the flesh of transplanted mussels. Based on biological similarities between the individuals of the transplanted population, their higher degree of bio-homogeneity, and better reproducibility of the results of metal accumulation, one can conclude that utilizing transplants instead of indigenous mussels, is more useful for monitoring of dissolved metals in t h e water phase. ACKNOWLEDGEMENTS Financial support by the International Bureau of the K F A Juelich, within the framework of the bilateral agreement between Germany and SFR Yugoslavia on the joint project 'Environmental Research in Aquatic Systems' is gratefully acknowledged. In addition, this study was partly supported by the U.S. National Bureau of Standards and the Research Council of SR Croatia, through funds made available to the U.S.-Yugoslav Joint Board on Scientific and Technical Cooperation under the project JFP (NSB)/649 as well as E U R E K A No. 493 E U R O M A R - E L A N I Project 'Development of Electroanalytical Instrumentation for Physico-Chemical Characterization of Trace Metals in the Marine Environment'. We thank Moira Spanovi~ for correcting the English and for typing the manuscript. REFERENCES Amiard-Triquet, C., B. Berthet, C. Metayer and J.C. Amiard, 1986. Contribution to the ecotoxicologicalstudy of cadmium,copper and zinc in the musselMytilus edulis. II. Experimental study. Mar. Biol., 92: 7-13. Branica, M., 7.. Peharec, 7.. Kwokal and S. Kozar, 1985. Trace metals in the Sibenik aquatorium: P-1 Concentrations of Zn, Cd, Pb and Cu analysed in the 1983/84 period. Rap. Comm. Int. Met Medit., 29: lll-ll3. Cauwet, G., 1989. Distribution and fate of organic carbon in a stratified estuary (Krka, Yugoslavia). Presented at First International Symposiumof Small Estuaries, May 21-27, 1989, Primogten,Yugoslavia, p. 40.
Zn, Pb, Cd AND Cu IN SEAWATER AND TRANSPLANTED MUSSELS
229
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