Metals in coastal mollusks of Costa Rica

The input of contaminants to coastal ecosystems is a global problem on the rise. Data on metal pollution from tropical sites is often lacking and pilot surveys are urgently needed to provide a general framework to estimate the relative impact of these and other pollutants. The objective of this study was to make accessible data on trace metals collected during pilot surveys (2000-2006) at four coastal embayments of Costa Rica. Cu, Fe, Mn and Zn were analyzed by Flame Atomic Absorption Spectrometry (FAAS) and Al, Cd, Ni, Pb and Sn by Graphite Furnace Atomic Absorption Spectrometry (GFAAS). Concentrations of Fe, Pb, Zn, Mn, and Ni were determined by FAAS or GFAAS, in tissues of the rock snail Acanthais brevidentata, the infaunal bivalves Anadara tuberculosa and Tagelus affinis (from three Pacific sites: Culebra Bay, Gulf of Nicoya, and Golfito Bay) and in the clam Polymesoda arctata from the Caribbean coast (Moín Bay). In addition, Sn, Cd and Cu were evaluated in tissues of T. affinis. A group of specimens of T. affinis was depurated for 72 hours in filtered seawater from the collection site. Concentrations varied between and within sites and also between parts of the same organisms. Maximum concentrations (μg/g dry weight) were: Fe 2 230 (P. arctata-tissues, Moín Bay), Pb 195 (P. arctata-tissues, Moín Bay), Zn 961 (A. brevidentata-tissues, Golfito Bay), Mn 921 (P. arctata-tissues, Moín Bay), and Ni 10.5 (A. tuberculosa-shells, Golfito Bay). Minimum concentrations (μg/g dry weight) were: Fe 5.36 (P. arctata-tissues, Moín Bay), Pb < 0.20 (P. arctata-foot, Moín Bay), Zn 2.75 (P. arctata-shell, Moín Bay), Mn 5.5 (A. tuberculosa-foot, Gulf of Nicoya) and Ni 0.83 (A. tuberculosa-foot, Golfito Bay). Tissues of nondepurated T. affinis had maximum-minimum concentrations (μg/g dry weight) of Sn (3.74-2.73), Cd (0.69-0.43) and Cu (21.6-14.8). The concentrations (except for Pb and Mn) were within values reported in recent literature. The relatively high concentration of Pb was probably related to the use at the time of sampling of leaded fuel in small boat operations at the site, while no evident cause was found for the high Mn values. Depuration was partially effective in lowering the metal burden in T. affinis. Data gathered during the pilot survey were indicative of relatively clean conditions of Culebra Bay, while the other three sites have important concentrations of certain contaminants, including metals. In spite of the fact that this data was collected more than a decade ago, it remains as the most recent available on trace metals from coastal mollusks of Costa Rica. Rev. Biol. Trop. 63 (4): 1007-1019. Epub 2015 December 01.

There are numerous scientific reports focusing on spatial and temporal variability of trace metal concentrations in organisms from temperate latitudes, particularly for clams and snails, groups commonly used in pollution monitoring (Bayen, 2012;Menzel, 1979).The situation is different for many tropical sites, where data on metal pollution is often lacking and pilot surveys are urgently needed to provide a general framework to estimate the relative impact of these and other pollutants.These surveys are the first step towards the design of studies aimed at identifying spatial and temporal variability of concentrations.Data generated during pilot surveys are often scattered in space and time, difficult to analyze statistically and usually only available in reports of limited circulation.However, due to the high cost of performing trace metal analyses, more elaborated studies do not often follow up pilot surveys and thus scattered data become the only available information for future reference.Thus, present and future easy access to this type of information might be critical in spite of its inherent limitations.
On the Pacific coast of Costa Rica, Dean, Maurer, Vargas, and Tinsman (1986) published the first report on trace metals in marine organisms from the Gulf of Nicoya estuary.The concentrations found were those expected for non-industrialized estuaries.Fuller et al. (1990) studied metal concentrations at the mouth and upstream the Tárcoles River which discharges untreated waters into this Gulf and chromiun was found to be an important metal contaminant.Guzmán and Jiménez (1992) evaluated the presence of twelve metals in the stony coral Siderastrea siderea on the Caribbean coasts of Costa Rica and Panama.They found that metal levels were generally higher than the ranges reported from other coral reefs worldwide.Also on the Caribbean coast of Costa Rica, Rojas, Acuña, and Rodríguez (1998) studied the concentrations of metals in tissues of the sea cucumber Holothuria mexicana and reported maximum values (mg/kg dw) of Fe (1 044), Zn (174), Cu (69), Mn (46), and Cd (2.5).
Between the years 2000 and 2006 data on pollutants were collected as part of pilot surveys at four coastal embayments, three located on the Pacific coast and one on the Caribbean coast of Costa Rica.Data on the ecology of the organisms was published by Rojas-Figueroa and Vargas-Zamora (2008), Sibaja-Cordero and Vargas-Zamora (2006), and Tarrant et al. (2008).The surveys also evaluated the use of polychaete worms as indicators of pollution in tropical sites (Dean, 2008), and updated the lists of polychaete, sipunculan, and echiuran worms (Dean, 2009;Vargas & Dean, 2009).
Published results obtained during the surveys emphasized the usefulness of the multiparameter approach in the identification of species, sites, and concentrations of concern.
For instance, García-Céspedes, Vargas-Zamora and Acuña-González (2004) collected sediment samples between 2000 and 2002 at various locations within these four embayments.They found maximun concentrations of Fe (73 000 μg/g, Culebra Bay), Zn (127 μg/g, Moin Bay), Cu (128 μg/g, Golfito Bay), and Pb (8.2 μg/g, Gulf of Nicoya and Golfito).Lower concentrations of metals were found in sands from Culebra Bay and they considered this site as relatively unpolluted, while Golfito Bay appeared as the most impacted of the four sites.The evaluation of polychlorinated biphenyls (PCBs) in sediments from the four sites was conducted by Spongberg (2004 a, b, c), who found that concentrations were relatively low but highest near the port of Golfito.The presence of PCBs in intertidal sipunculan worms was also evaluated by Spongberg (2006) who found that concentrations ranged from as low as 0.01 ng/g dw in Antillesona antillarum from Culebra Bay to a maximum of 67 ng/g dw in Phascolosoma perlucens from near the Rincón river mouth close to the entrance of Golfito Bay.Moreover, the cosmpolitan peanut worm Sipunculus nudus from the Cocorocas sand flat (Gulf of Nicoya) had relatively high concentrations of PCBs ranging from 17.7 to 41.3 ng/g dw, whether the worms were depurated or not.Spongberg & Witter (2006) reviewed published reports on PCBs from tropical sites including those collected during the pilot study.They concluded that these values are low when compared to data from temperate latitudes.García, Acuña-González, Vargas-Zamora and García-Céspedes (2006) conducted an evaluation of coliform bacteria and beach litter at the four sites.The least polluted location was Culebra Bay with counts for total and fecal coliform bacteria of around 2 MPN (Most Probable Number)/100 mL.The port city of Golfito was the most impacted, with total and fecal coliform bacteria over 50 000 MPN / 100 mL.Plastic material was the most common item found on all beaches at the four sites.Acuña-González Vargas-Zamora, Gómez-Ramírez, and García-Céspedes (2004) studied by molecular florescence the concentrations of dissolved/dispersed petroleum hydrocarbons at the four sites.No fluorescence signal was detected in samples from Culebra Bay indicating no oil contamination at the time.Concentrations at the other three sites were below 2 μg/L and considered as indicative of very low contamination with oil.
The importance of low concentrations of other contaminants was not disregarded and the pilot studies also addressed this topic.For instance, no evidence of endocrine disruption was detected in marine turtles (Lepidochelys olivacea) nesting near Culebra Bay (Valverde, Selcer, Lara, & Sibaja-Cordero, 2008).Tin compounds like tributyltin oxide (TBT) from antifouling paints, found in extremely low concentrations in seawater, have been linked worldwide to the development of imposex in gastropods.Imposex was detected in the snail Acanthais brevidentata from the port of Caldera in the Gulf of Nicoya, but it was absent in Culebra Bay (Gravel, Johanning, McLachlan, Vargas, & Oberdörster, 2006).The presence of other organic residues is also important in this context as emphasized by Cheek (2006).As a follow up of the surveys Spongberg et al., (2011) have reported on the presence of Pharmaceutical and Personal Care Products (PPCPs) in 86 sampling locations in rivers draining into Pacific and Caribbean coastal areas of Costa Rica.The identification of 34 compounds including many antibiotics and caffeine, is noteworthy.
In addition to A. brevidentata, three other species of coastal mollusks, including two commercially harvested clams, were also chosen during the pilot studies to evaluate the presence of metals in tissues with the expectation of identifying concentrations of concern.Thus, the objective of this report is to make available these latter data on metals in tissues from mollusks, and to compare the maximum concentrations found to selected examples from the literature.

MATERIALS AND METHODS
Study sites and organisms: Three of the chosen sites (Culebra Bay, Gulf of Nicoya and Golfito Bay) are on the Pacific coast of Costa Rica, where mean tidal range is approximately 3 m.The fourth site, Moín Bay, is on the mid Caribbean coast (Fig. 1), where mean tidal range is less than 0.5 m.The presence of the four species of mollusks collected in this study (Fig. 2) was confirmed for Costa Rica by Wehrtmann and Cortés (2009) who listed them as: Acanthais brevidentata (Wood, 1828), Anadara tuberculosa (Sowerby, 1833), Tagelus affinis (C.B.Adams, 1852) and Polymesoda arctata (Deshayes, 1854).
Culebra Bay is an embayment on the upper Pacific coast and its mouth opens into the Gulf of Papagayo, an upwelling region.The bay is surrounded by tourism facilities that make use of the numerous white sand beaches.The published records on the marine biodiversity of Culebra Bay have been compiled by Cortés, Vargas-Castillo, and Nivia-Ruiz (2012) but no information on trace metals in organisms was located.The spatial variability of the rocky intertidal fauna, including the snail A. brevidentata, has been described by Sibaja-Cordero and García-Méndez (2014).
The Gulf of Nicoya is an estuary on the central Pacific coast of Costa Rica.Since the Mid-20 th Century the Gulf has been the main fishing ground of the country for finfish and shellfish.Research on the Gulf conducted mainly after 1980 makes it one of the best known tropical estuaries worldwide (Vargas, 1995, Vargas & Mata, 2004).The ark clam Anadara tuberculosa (known locally as "piangua") continues to be extracted for human consumption after the decline of the bigger Grandiarca grandis (known locally as "chucheca") due to overfishing (Stern-Pirlot & Wolff, 2006).This overfishing prompted the harvesting of smaller bivalves, like the white razor clam Tagelus affinis.The snail A. brevidentata is also a common gastropod on the rocky shores of the Gulf, particularly near the port of Caldera (Sibaja-Cordero & Vargas-Zamora, 2006).Golfo Dulce is an estuarine embayment on the South Pacific coast of Costa Rica.Its deep bathymetry, similar to a fjord, has encouraged comparative research leading to significant discoveries, such as the scarcity of infaunal invertebrates below 50 m (León-Morales & Vargas, 1998), the anammox reaction in the anoxic water column, and the presence of bacterially-derived bacteriohopanepolyols (Dalsgaard, Canfield, Petersen, Thamdrup, and Acuña-González 2003;Rush et al., 2014).Golfito Bay is a small and shallow embayment on the Northern shore of the gulf.It houses the main port for the region and the city of Golfito is located along its shores.The Purruja mangrove forest is on the South-Eastern shore of the bay.There, A. tuberculosa is extracted for local consumption (Silva-Benavides & Bonilla, 2001).So far the only report on pollutants from Golfo Dulce is by Spongberg and Davies (1998) who studied the concentrations of organochlorinated pesticides in sediments.
Moín Bay is on the central Caribbean coast of Costa Rica.It houses the main sea port facilities for container ships and is under the process of further development.Part of the Moín River runs parallel to the shore and is bordered by mangroves.An oil refinery is located several kilometers upstream.Brackish waters are found at the river mouth and a port facility for tourism river boats operates there.
Here the clam Polymesoda arctata was found to be abundant and is occasionally harvested.
The four species of molluks were chosen for several reasons: the snail A. brevidentata is a top predator in the rocky intertidal zone, where specimens are abundant and easy to identify in the field.The snails cluster together at low tide in shaded spaces which facilitates obtaining enough biomass for study.The three filter feeding infaunal bivalves are also easy to identify and to collect from intertidal sediments, and are haversted for local consumption.Moreover, the four species are easy to transport and to maintain in the laboratory.These characteristics are among those cited by Cunningham (1979) for mollusks to be used as bioindicator organisms.A. brevidentata snails were hand collected from rocks exposed at low tide at Culebra Bay (in July 2003 and October 2004), the port of Caldera (Gulf of Nicoya, in October 2004) and at the Golfito bay mouth (in October 2004).Specimens of T. affinis were purchased on site from a fisherwoman working at the Cocorocas sand flat in February 2006.Ark clams (A.tuberculosa) were hand collected from muddy sediments at the edges of mangrove stands in the Gulf of Nicoya (Cocorocas mangrove, in October 2004) and Golfito Bay (Purruja mangrove, in April 2006).P. arctata was also hand collected from muddy sediments between the roots of mangrove trees at the Moín river mouth, in July and September 2006.

Metal analyses:
After collection the mollusks were washed with sea water from the sites to remove dirt from the shells and transported to the laboratory in acid-washed polyester bags.At the laboratory the organisms were counted, rinsed with filtered sea water, blotted dry on a paper towel, weighed, separated each into equal-numbered groups of individuals, and stored frozen (-18 ºC) in a heat sealable, acid-washed polyester bags until analyses.
Prior to analyses individuals of A. brevidentata were placed in acid-washed polyester bags and the shells crushed on a vise.Shell fragments were removed by hand and tissues saved for analyses.Bivalve shells were opened with a plastic knife and tissues (gills, digestive tract, muscles, and mantle) removed from the shell.In specimens of A. tuberculosa and P. arctata the foot was cut off and analyzed separately and shell fragments for analyses included the periostracum.A group of T. affinis was kept in a plastic container filled with filtered sea water from the site (Fig. 2) and aerated with plastic aquarium pumps to explore if organisms were able to survive and remove sediments and other matter from their digestive tracts.Sea water, feces, and dead organisms, were discarded every 12 hours and the containers filled with fresh filtered sea water from the site.The procedure was repeated six times for a total of 72 hours.Surviving organisms (more than 90 %) were then stored frozen as above until trace metal analyses.
All metal determinations were performed at the Centro de Contaminación Ambiental (CICA) at the University of Costa Rica, under validated and accredited protocols by the Costa Rican Accrediting Agency (ECA).Groups of 10-30 specimens were randomly subsampled from each batch and weighed after drying in an oven at 65 ºC for 48 hours.Dried tissues were finely ground to a homogeneous powder and samples of 1.00 g were placed in Teflon containers with 6 mL of high purity nitric acid, 2 mL of high purity hydrochloric acid, and pressure-digested in a microwave oven (CEM ® MARSx).After digestion, the containers were cooled at room temperature and the liquid was transferred to 100 mL volumetric flasks and diluted with deionized water.Certified reference materials and blank tests were also run.Cu, Fe, Mn and Zn were analyzed by Flame Atomic Absorption Spectrometry (Perkin Elmer ® 3300) and Al, Cd, Ni, Pb and Sn by Graphite Furnace Atomic Absorption Spectrometry (Perkin Elmer ® HGA 600).Calibration curves were produced against certified standards for each element, according to the CICA accredited protocols.All the results were expressed in μg/g dry weight (dw).Concentrations of Fe, Pb, Zn, Mn, and Ni were determined in mollusks collected at the four sites (Culebra, Gulf of Nicoya, Golfito, Moín Bay), while Cd, Cu, and Sn were determined only in non-depurated T. affinis clams from the Gulf of Nicoya.Detection limits typical values were 0.40 μg/g for Al, 0.0020 μg/g for Cd, 1.6 μg/g for Cu, 0.10 μg/g for Sn, 1.9 μg/g for Fe, 0.91 μg/g for Mn, 0.022 μg/g for Ni, 0.027 μg/g for Pb, and 18 μg/g for Zn.

RESULTS
The concentrations of metals found in tissues of the snail A. brevidentata from Culebra Bay are shown in Table 1, and those found in A. brevidentata, the clams T. affinis and A. tuberculosa from the Gulf of Nicoya are listed in Table 2.The metal concentrations in A. brevidentata and A. tuberculosa from Golfito Bay are included in Table 3 and those for the clam P. arctata from Moín Bay are listed in Table 4.The maximum and minimum concentrations are summarized in Table 5.The uncertainty (±) corresponds to the expanded uncertainty using a coverage factor k = 2.The uncertainty (±) corresponds to the expanded uncertainty using a coverage factor k = 2.
Among the five determined metals at the four sites (Fe, Pb, Zn, Mn, and Ni), concentrations of iron above 1 400 μg/g-dw were found in tissues of non-depurated T. affinis from the Gulf of Nicoya (Cocorocas flat, Table 2).
On the Caribbean coast, iron was found to be greater than 1 400 μg/g-dw in tissues of P. arctata (Table 5).Concentrations of lead in T. brevidentata were around 22 μg/g-dw in Culebra, Gulf of Nicoya and Golfito (Table 1, The uncertainty (±) corresponds to the expanded uncertainty using a coverage factor k = 2.The uncertainty (±) corresponds to the expanded uncertainty using a coverage factor k = 2.
Table 2 and Table 3), while the highest concentration (195 μg/g-dw, Table 5) was found in P. arctata at the Caribbean port.Zinc above 300 μg/g-dw was found in the snail T. brevidentata at the three Pacific sites, with a maximum of 961 μg/g-dw (Golfito Bay-tissues, Table 5).
Maximum concentration for the Caribbean coast was 485 μg/g-dw in tissues of P. arctata (Table 4).Manganese above 100 μg/g-dw was found in non-depurated T. affinis clams from the Cocorocas sand flat (Table 2), with a maximum of 255 μg/g-dw (Table 5), while much higher values were detected in tissues of P. arctata, with a maximum of 921 μg/g-dw (Table 5).Nickel in concentrations above 2 μg/g-dw was present in tissues of the snail A. brevidentata from Culebra Bay, Gulf of Nicoya, and Golfito Bay (Tables 1, 2, 3), with the highest concentration (10.5 μg/g-dw) in the shell of the ark clam A. tuberculosa from Golfito (Table 5).Concentrations of nickel were above 2 μg/g-dw in tissues and shell of P. arctata from the Caribbean coast, with a maximum of 8.8 μg/g-dw (Table 5).Maximum concentrations of tin, cadmium and copper were respectively 3.74 μg/g-dw, 0.69 μg/g-dw, and 21.6 μg/g-dw, all in nondepurated tissues of the clam T. affinis (Table 2).The 72 hour depuration for T. affinis yielded interesting results as concentrations of Fe, Zn, Mn, and Ni were lower in depurated as compared with non-depurated tissues (Table 2).However, Pb concentrations were higher in depurated tissues.
Table 6 includes examples of maximum concentrations of metals in other species of molluks from selected locations around the world.When maximum concentrations in Table 5 are compared with those in Table 6, lead (195 μg/g-dw) and manganese (921 μg/g-dw) both in P. arctata, were found much higher in this study.

DISCUSSION
The generation and interpretation of data on trace metals in marine molluks are difficult for several reasons.Metal concentrations vary in individuals of the same species depending on their sex, age, and on the physiological and environmental conditions.In addition, parts (foot, tissues, gills, shell) of the same organism may accumulate metals in different concentrations (Cunningham, 1979;Zuykov, Pelletier, & Harper, 2013).The differences in accumulation of trace metals in parts of the same organism, is well known.The early report by Brooks and Rumsby (1965) on the scallop Pecten novaezelandiae illustrates this fact: Concentrations (μg/g) of metals in soft tissues were: Fe (6 900), Pb (23), Zn (368), Mn (306), Cd (299) and Cu (14), while for instance the gills had Fe up to 21 600 μg/g, the kidney had 137 μg/g of Pb, 2 630 μg/g of Zn, 2 660 μg/g of Mn, and 106 μg/g of Ni.The intestine had 131 μg/g of Cu, and the visceral mass had 2 000 μg/g of Cd.
Samples collected relatively close to each other may also show differences due to point sources of different magnitudes.Data presented in this study include examples of this variability and point out to the need for more detailed studies in space and time.Some metals are essential in the physiology of molluks.For instance, iron is found in hemoglobin, a respiratory red pigment that gives the name "blood cockles" to Anadara ark clams, while other mollusks may contain hemocyanin, a green pigment with copper.Clark (2001) classifies metals in two groups: transitional (like Fe, Cu, and Mn), which are essential at low concentrations, and metalloids (like Pb and Sn) which are not essential but are toxic at low concentrations.Cadmiun is also not essential but bivalve molluks tend to accumulate it.Many enzymes contain zinc and other metals (Lane et al., 2005).Clark ( 2001) also indicates that in some marine ecosystems the natural amount of a substance might be altered due to anthropogenic actions, and this can be considered as contamination.But at the same time if there is a bigger natural input of such a substance in other areas, its concentration there could be higher than that caused by human intervention.
With the above facts in mind the four coastal sites evaluated for trace metals in this study differ in the nature and magnitude of the inputs from natural sources and human activities.For instance, the Gulf of Nicoya is under the influence of estuarine water mass circulation patterns (Voorhis, Epifanio, Maurer, Dittel, & Vargas, 1983) that may bring contaminants from the Tempisque (agricultural) and Tárcoles (agricultural, industrial, sewage) rivers in contact with mollusks located far apart from the point sources.On the other hand, no major river discharges wastes into Culebra Bay where sewage and runoff inputs are also minimal in this the driest region of Costa Rica.Thus, the expected low concentrations of metals in Culebra Bay were so for Fe, Mn, and Ni, but Pb and Zn were similar to values found in the other Pacific sites (Table 5).Golfito Bay is under the direct influence of runoff (heavy rains are present year-round) and untreated sewage from the city of Golfito settled along its shores.Concentrations of Fe and Pb were similar in the Gulf of Nicoya and in Golfito, but Zn and Ni in Golfito had the highest values of the four embayments (961 and 10.5 μg/gdw, respectively, Table 5).Concentrations of Pb were around 22 μg/g in tissues of A. tuberculosa from the Gulf of Nicoya and Golfito (Table 5).
On the Caribbean coast Moín Bay receives inputs (agricultural, industrial) from the Moín River and from the operation of many small river boats equipped with outboard fuel-leaded engines.The highest concentration of Pb found in P. arctata from Moín Bay (195 μg/g-dw) is probably related to this activity.The highest mean concentration of Mn (921 μg/g-dw) also found in P. arctata is difficult to identify as to its possible source.Nonetheless, according to Simkiss & Mason (as cited in Páez-Osuna, 2005), it is usual to observe high concentrations of metals in some tissues of mollusks as part of a detoxification process related to proteins, and also it has been found that some female mollusks have higher Mn and Zn concentrations than males (Marina & Enzo, 1983).In our study we did not discriminate between males and females.The nature of the substrate is also important in studies of trace metals.Finer sediments usually contain higher concentrations of contaminants than coarser sediments (Ujevic, Odzak, & Baric, 2000).In this context clams like A. tuberculosa (Gulf of Nicoya and Golfito) and P. arctata (Moín Bay) are found in fine grained sediments, while T. affinis lives in an extensive coarse sand flat at the mouth of the Lagarto River, which drains mostly grass lands for cattle raising.The snail A. brevidentata lives on sedimentary and igneous rocks in Culebra Bay, Gulf of Nicoya, and Bay.Recent evidence also suggests that the nature (type and species of microalgae) of phytoplankton ingested by bivalves influences the assimilation of certain metals, Hédouin et al. (2010).
In Costa Rica the clams A. tuberculosa, T. affinis and P. arctata are under different fishing pressures as food items.However, research has been focused mainly on the heavily harvested A. tuberculosa (Stern-Pirlot & Wolff, 2006).Hossen, Hamdan and Rahman (2014) found concentrations of Pb of 2.65 to 4.36 μg/g-dw in Anadara granosa in Malaysia.They point out that the maximum permissible limit of Pb by the Malaysian Food Authority is 2.00 μg/gdw.According to this reference level, the A. tuberculosa specimens from Costa Rica analyzed in this study were not suitable for human consumption, nor were specimens of the other bivalves analyzed.The results obtained with the 72 hour depuration experiment of T. affinis clams are important in the context of lowering the content of certain metals enough to make clams suitable for human consumption.Median concentrations of Fe, Zn, Mn, and Ni were lower in depurated tissues when compared to non-depurated.However, the increase of Pb in depurated tissues is interesting and deserves further study.
We have also included some reports on trace metal concentrations from tropical molluks.The maximum values (μg/g-dw) reported are: Fe 3400, Pb 46.34, Zn 4 380, Mn 45.9, Ni 54, Cd 374.1, and Cu 866.In this context the relatively high concentrations of Pb (195) and Mn (921) found in this study are noteworthy.Depuration could be applied also to the widely used food item A. tuberculosa from the Gulf of Nicoya and Golfo Dulce.However, depuration seems to produce unexpected results.For instance, Yap Muhamad, Cheng and Tan (2011) found no significant difference in the depuration levels between Cu and Zn in soft tissues of A. granosa from Malaysia.Their study also evaluated the concentration of metals in the shells and concluded that in the long term shells may be able to accumulate higher concentrations of metals than the soft tissues.The relatively high concentrations of Ni and Mn found in shells of A. tuberculosa may be indicative of similar mechanisms in this species.The concentrations of trace metals found in this pilot survey are generally high for mollusks that continue to be harvested for human consumption.
It must be taken into account that the data on metals presented herein was collected more than a decade ago but remains as the most recent reference for trace metal concentrations in coastal mollusks from Costa Rica.Input sources of contaminants to the four embayments are expected to have increased over the past decade due to local coastal development and to water and sediment loads carried by the rivers.A new detailed spatial and temporal evaluation of metals in mollusks and other invertebrates is advised.
Rica-United States of America Foundation for Cooperation (CR-USA), and the University of Costa Rica (UCR).We thank Davis Morera, Eleazar Ruiz, and Jeffrey Sibaja for their help in the field and the laboratory.Karina Rodríguez collected the specimens of Polymesoda arctata.The preparation of this paper was part of project 808-B0-007 supported by UCR.We thank Harlan K. Dean for the review of an earlier draft.

Fig. 1 .
Fig. 1.Study sites on the Pacific and Caribbean coasts of Costa Rica.

Fig. 2 .
Fig. 2. A. Acanthais brevidentata snails from the rocky intertidal zone at the port of Caldera, Gulf of Nicoya (Reproduced from Gravel et al. 2006).B. Tagelus affinis clams during the depuration period in clean sea water (Note long siphons), Cocorocas sand flat, Gulf of Nicoya.C. Shell of Anadara tuberculosa from the edge of the Purruja mangrove forest, inner Golfito Bay, Golfo Dulce.Botton right: eroded shell.D. Polymesoda arctata clams from the Moin River mouth, Moin Bay,Caribbean coast.

TABLE 4
Trace metals (Fe, Pb, Zn, Mn, Ni) concentrations (µg/g dw) in tissues and shells of the clam Polymesoda arctata from the Moin river mouth, Caribbean, Costa Rica.July and September 2006.