Induction Of Metallothionein-like Proteins Research Articles

Effects of waterborne exposure to cadmium on biochemical responses in the freshwater gastropod, Bellamya aeruginosa.

The biochemical responses of Bellamya aeruginosa as a dominant and widespread freshwater gastropod throughout China to waterborne cadmium (Cd) were investigated to explore the impacts of exposure concentration and duration in this potential sentinel species. After the 7 days' test of dosage-mortality relationship, gastropods were exposed for either 7 days at the LC50 (1.7mg/L), the LC10 (0.7mg/L) and 0.02mg/L Cd, or 28 days at 0.02mg/L Cd. A suite of biochemical indicators including metallothionein-like protein (MTLP), reduced glutathione (GSH), catalase (CAT), contents of tissue metal (Cd, Fe, Mn, Cu, Zn), and the compartments of these metals bound to MTLP were examined. The treatment of 0.02mg/L Cd led to the increase of Cd bound to MTLP (Cd-MTLP) levels, the decrease of GSH content, and the upregulation of CAT activity, but no induction of MTLP, indicating that the intrinsic MTLP and GSH worked together for the detoxification of Cd at the low exposure. When the exposure concentration increased, GSH was depleted severely and synthesis of MTLP was triggered, leading to a strong and significant relationship between MTLP level and Cd accumulation. At the lethal concentrations (1.7mg/L), both MTLP induction and CAT activity were inhibited while the proportion of Cd-MTLP to total Cd were increased, suggesting more intrinsic MTLP were utilized to sequester free Cd ions. Therefore, the content of Cd-MTLP in digestive glands of B. aeruginosa was recommended as a reliable biomarker for Cd contamination.

Implications on the Pb bioaccumulation and metallothionein levels due to dietary and waterborne exposures: The Callinectes danae case

This study aimed to assess the bioaccumulation of Pb and induction of metallothionein-like proteins (MT) in Callinectes danae through single and combined dietary and waterborne exposures. Male C. danae individuals were collected in the south area of the Cananéia-Iguape-Peruíbe Protected Area (APA-CIP), in São Paulo State, Brazil. After an acclimatization period, exposure assays were performed during 7 and 14 days, at two Pb concentrations (0.5 e 2.0 µg/g) in 4 treatments: 1) control; 2) contaminated water only; 3) contaminated food only; 4) contaminated water and food. The results indicate that C. danae is highly tolerant to Pb exposure at the evaluated concentrations. In gills, Pb bioaccumulation is more dependent of water efflux and time of exposure (higher Pb values). However, pathways act simultaneously in the induction of MT expression in this tissue. The decreases in Pb accumulation in the combined treatments and MT increases after 14 days in gills suggests that these proteins play a detoxification function in the presence of Pb. In hepatopancreas, depending on the predominance of a certain pathway or combined pathways, accumulation occured at different times. For muscle tissue, bioaccumulation was observed due to contaminated water exposure, but not dietary exposure, probably because Pb concentrations were low.

The Effect of Zinc and Selenium Supplementation Mode on Their Bioavailability in the Rat Prostate. Should Administration Be Joint or Separate?

It is thought that zinc and selenium deficiency may play a significant role in the etiology of prostate cancer. Although joint zinc and selenium supplementation is frequently applied in the prevention of prostate diseases, the bioavailability of these elements in the prostate after co-administration is still unknown. The study examines the effect of subchronic supplementation of zinc gluconate and selenium compounds (sodium selenite or selenomethionine), administered together or separately, on their bioavailability in the prostate, as well as the induction of metallothionein-like proteins (MTs) bound to zinc in the prostate and liver. Zinc concentration in the dorso-lateral lobe of the prostate was significantly elevated already after the first month of supplementation of zinc alone. In the supplementation period, the MTs level increased together with zinc concentration. In contrast, the ventral lobe of the prostate did not demonstrate significantly higher levels of zinc until after three months of supplementation, despite the MTs induction noted after one-month supplementation. Increased selenium levels in the dorsolateral lobe were observed throughout the administration and post-administration periods, regardless of the selenium compound used or whether zinc was co-administered. The results of our studies suggested for the first time that these elements should not be administered jointly in supplementation.

Accumulation and detoxification of copper and zinc by the decapod crustacean Palaemonetes varians from diets of field-contaminated polychaetes Nereis diversicolor

In an investigation of the trophic transfer of ecotoxicologically significant amounts of trace metals from prey to predator, Palaemonetes varians were fed with diets of Nereis diversicolor from two metal-contaminated estuaries, the Tavy and Restronguet Creek in SW England, and a control estuary, the Blackwater in SE England. The decapods accumulated increased concentrations of copper (but not zinc) from the metal-contaminated worms above an initial apparently regulated body copper concentration, in response to a high dietary copper input over a period up to 30days. High dietary challenges of copper caused the induction of MTLP (metallothionein-like proteins) in the hepatopancreas of the feeding decapods as a detoxification response, although the non-detoxified subcellular component of hepatopancreas accumulated copper also increased over time, probably in association with sublethal toxic effects.

Ecotoxicological Heterogeneity in Transitional Coastal Habitats Assessed Through the Integration of Biomarkers and Sediment-Contamination Profiles: A Case Study Using a Commercial Clam

Transitional waterbodies, such as estuaries, are highly diversified environments with respect to ecology, geophysics, and nature of anthropogenic impacts. This spatial heterogeneity may pose important constraints when developing monitoring programmes for aquatic pollution. The present study compared three distinct coastal ecosystems located in Southern Portugal (subjected to different anthropogenic stressors), namely, two estuaries and a coastal lagoon, through the characterisation of sediment contamination and a biomarker approach to an important commercial clam (Ruditapes decussatus) obtained from local fishing grounds. The results showed high heterogeneity of sediment contamination for both estuaries and a marked distinction between industrially and agriculturally influenced areas as well as between natural and artificialized sites. Hydrodynamics and oceanic influence (in essence dictating sediment type) play a major role in environmental quality. Environmental heterogeneity constituted an important confounding factor for biomarker analysis in the clams' digestive glands since the animals appeared to respond to their immediate surroundings' characteristics rather than the geographical area where they were collected from, despite the relative distance to pollution hot spots. Still, oxidative stress biomarkers (lipid peroxidation and catalase activity) could correlate with each other and to both organic and metallic contamination, whereas metallothionein-like protein induction failed to correlate to any class of sediment toxicants (albeit metals being the most representative pollutants) and appeared to be strongly affected (unlike the previous) by clam size and probably other unknown internal and external variables, among which contaminant interactions should play a major role.

The impact of increased oxygen conditions on metal-contaminated sediments part II: Effects on metal accumulation and toxicity in aquatic invertebrates

The present study evaluated the effect of increasing oxygen concentrations in overlying surface water on the accumulation and toxicity of sediment-bound metals in the aquatic invertebrates Lumbriculus variegatus, Asellus aquaticus and Daphnia magna. A 54 days experiment using three experimental treatments (90% O2 in overlying surface water, 40% O2 and a non-polluted control) was conducted. At 6 different time points (after 0, 2, 5, 12, 32 and 54 days) acid volatile sulfides (AVS), simultaneously extracted metals (SEM) and total organic carbon (TOC) were measured in the superficial sediment layer (0–1 cm). At each time point, accumulated metal levels as well as the available energy stores were measured in L. variegatus and A. aquaticus and each time D. magna was exposed to surface water in a 24 h toxicity test. Additionally metallothionein-like protein (MTLP) induction was quantified in L. variegatus. Oxygen induced changes in sediment AVS resulted in faster accumulation of metals from contaminated sediments in A. aquaticus, while no differences in toxicity in this species were observed. Ag, Cr, As and Co accumulation as well as toxicity in water exposed D. magna were clearly enhanced after 54 days, caused by oxidation of metal-sulfide complexes. Due to their feeding and burrowing behaviour, metal accumulation and toxicity in L. variegatus was not influenced by geochemical characteristics. Nevertheless, a rapid induction of MTLP was observed in both the 90% O2 and the 40% O2 treatment. The present study showed that elevated oxygen concentrations in overlying surface water can directly enhance metal accumulation and toxicity in aquatic invertebrates, however this is highly dependent on the organisms ecology and most dominant metal exposure route (water vs. sediment).

Metallothionein-like proteins turnover, Cd and Zn biokinetics in the dietary Cd-exposed scallop Chlamys nobilis

In this study, we tested whether metallothionein-like proteins (MTLPs) affected the biokinetics of Cd and Zn in the scallops Chlamys nobilis following dietary Cd exposure. The scallops were fed Cd-contaminated diatoms for 8 or 40 days, and then the tissue Cd concentrations, MTLP turnover and their standing stocks, Cd and Zn kinetics were monitored. After 8 days of dietary Cd exposure (at 98 or 196 μg Cd g −1), their Cd levels were increased by 7.4–12.5 times, newly synthesized MLTPs by 1.7–2.1 times, and their MLTP stocks doubled. However, after 40 days of dietary Cd exposure (at 58 or 115 μg Cd g −1), MTLP synthesis and MTLP stocks did not change despite the fact that Cd bioaccumulation increased by 2.7–4.2 times. MTLPs played little role in the overall Cd and Zn uptake from either food or water, since enhanced MTLP induction did not improve the sequestration of newly incoming Cd or Zn. As MTLP-metal complexes degraded, the released Cd was immediately sequestered by newly synthesized MTLPs while most Zn was depurated. This may explain why scallops eliminated Cd more slowly than Zn. Induced proteins were shown to play a minor role in the detoxification of dietary Cd in C. nobilis, but the species can modify its Cd assimilation to reduce the impact of high dietary Cd levels. MTLPs are probably unsuitable as biomarkers for environmental monitoring involving C. nobilis.

Bioaccumulation and Effects of Metals Bound to Sediments Collected from Gulf of Cádiz (SW Spain) Using the Polychaete Arenicola marina

A short-term whole-sediment test using the polychaete Arenicola marina was conducted under laboratory conditions to assess the bioavailability of metals bound to sediments collected from 12 sites of the Gulf of Cádiz. To achieve this objective, the rate of increase of metal bioaccumulation and the induction of a typical biomarker, metallothioneinlike proteins (MTLPs) were determined. Results of the multivariate analysis showed associated metal-rich sediments, increased rate of Cu and Zn accumulations, but lower toxicity with an increased MTLP induction, whereas sedimentary Ni and Co concentrations were related to higher toxicity to lugworms, although it might be caused by other contaminants present in these sediments. The linear kinetic approach was shown to be valid in certain circumstances, but more validation studies of this parameter are required before it can be recommended for use in evaluating metal bioavailability in sediments.

Are Accumulated Sulfide-Bound Metals Metabolically Available in the Benthic OligochaeteTubifex tubifex?

The present study evaluates the relationship between metal-binding sediment characteristics like acid volatile sulfides (AVS), metal accumulation, and internal metal distribution in the benthic oligochaete Tubifex tubifex and relates this accumulation to the induction of metallothionein-like proteins (MTLPs). In total, 15 Flemish lowland rivers were sampled. Cd, Cu, Zn, Pb, Ni, As, Cr, Co, and Ag concentrations were measured in environmental fractions (water and sediment) and worm tissue (both total and subcellular fractions). Furthermore, total cytosolic MTLP concentrations were measured in the worm tissue. Our results showed that Cd, Pb, Ni, and Cr were mainly stored as biological detoxified metal (BDM) while Cu, Zn, As, and Ag were mostly available in the metal sensitive fraction (MSF). A remarkable difference in the subcellular distribution of accumulated Cd, Ni, and Co between anoxic (SEMMe-AVS<0; mostly stored as BDM) and oxic (SEMMe-AVS>0; mostly stored in the MSF) sediments was noticed. Moreover, a rapid increase in MTLP induction was found when SEMTot-AVS>0. Our results indicate that the accumulated sulfide-bound metals were detoxified and little available to the metabolism of T. tubifex under anoxic conditions.

Short-term metal accumulation and MTLP induction in the digestive glands of Perna virdis exposed to Zn and Cd

Time-dependent Zn and Cd accumulation and metallothionein like protein (MTLP) induction in the digestive glands of mussels, Perna virdis, were measured under different exposure conditions. The initial uptake rate at start of chase (ρ 0) and mean residence time (τ) were calculated to determine the physiological response of organisms and their potential detoxification mechanisms. It was found that in digestive glands, Zn had obviously higher ρ 0 and shorter mean residence time than Cd, indicating that these two metals had different accumulation dynamics even though they were very close in the periodic element table. MTLP levels in digestive glands varied from 0.51 to 1.05 μg/g ww (wet weight). The MTLP level increased continuously when mussels were exposed to low and middle levels of Zn and Cd media, and reached maximal levels at day 4, then decreased when they were exposed to high level Zn and Cd solutions. With regard to the fraction of Zn and Cd accumulated in the digestive glands, the ratios of soluble metal to total metal decreased continuously after exposure in low and middle levels of Zn and Cd media, and decreased continuously in the first 4 days and then to level off when mussels were exposed to media with high concentration of Zn and Cd. Results suggested that both MTLP induction and metal insolubilization were detoxification processes in digestive glands of mussels.

Comparative toxicity of cadmium in the commercial fish species Sparus aurata and Solea senegalensis

The induction of metallothionein-like proteins (MTLPs) as well as cadmium levels (Cd) was studied in tissues of gilthead sea bream Sparus aurata and flatfish Solea senegalensis after an intraperitoneal (i.p.) injection of cadmium (2.5 mg kg −1 body weight). The liver, gills, intestine and blood of S. aurata as well as the liver and intestine of S. senegalensis were collected for analysis at 0, 3 and 6 days after the injection. Cd levels significantly increased in all tissues of the treated animals, and the highest accumulation was found in the liver in both species ( p<0.05). Cadmium administration provoked significant inductions of MTLP synthesis in the liver and intestine of S. aurata ( p<0.05), while no increase in these protein levels was recorded in tissues of S. senegalensis. Our findings imply that MTLP synthesis in S. aurata probably led to an enhanced ability to cope with the toxicant, whereas the MTLP induction in S. senegalensis was inhibited by the accumulated cadmium, as reflected by the mortality rate of this species. Under the experimental conditions, hepatic MTLPs appear to be a good indicator of the Cd level in S. aurata; however, no link was found between the cadmium concentration and MTLP induction in S. senegalensis, which suggests that MTLPs did not play a main role in detoxification by metal sequestration.

In vivo variation of some biomarkers with time and cadmium concentration in the cockle Cerastoderma glaucum

The aim of this work was to detect in vivo effects of cadmium on marine organisms Cerastoderma glaucum and to determine the optimal conditions for the metallothionein-like protein (MTLP) induction in this species under laboratory conditions. The cockles Cerastoderma glaucum were exposed in vivo to different cadmium concentrations (50, 75, 100 and 150 µg L−1). Stress on stress test, cadmium, metallothionein-like protein (MTLP), malondialdehyde (MDA) and protein levels were measured after 5, 10, 15 and 20 days of metal exposure. The results showed that the CdCl2 concentrations of 50 µg L−1 for 15 days as well as 150 µg L−1 for 10 days provoked the highest MTLP induction. Nevertheless, the results obtained by the MDA, protein quantification and the stress on stress test showed that animals exposed to 50 µg L−1 for 15 days were in a better health status than those exposed to 150 µg L−1 CdCl2 for 10 days.

Evaluation of induction of metallothionein-like proteins (MTLPs) in the polychaetes for biomonitoring of heavy metal pollution in marine sediments

Polychaetes are suitable organisms for evaluation of impact of sediment pollution. We evaluated toxicity of cadmium and copper and measured metallothionein-like proteins (MTLPs) in the polychaete Perinereis nuntia. At the same concentration ranges copper was unexpectedly more toxic than cadmium. Copper also caused no significant increase in MTLPs in the polychaetes. When P. nuntia and another polychaete species, Cirratulus cirratus were cultured in the contaminated sediments collected from Lake Sihwa (Korea), a high mortality of 80% was observed on day 6 in P. nuntia in the sediment with the highest metal concentration. However, no mortality was observed up to 35 days in C. cirratus in any sediment. MTLP contents between two species also varied. These findings suggest that MTLP induction response in the polychaetes varies with the metal type and species and it may be used as a biomarker of sediment pollution in the polychaetes after further validation and field trials.

Field validation of a battery of biomarkers to assess sediment quality in Spanish ports

Two marine invertebrates, the crab Carcinus maenas and the clam Ruditapes philippinarum, were used as bioindicator species to assess contamination when exposed in situ to sediment from different sites from four Spanish ports Cadiz (SW Spain), Huelva (SW Spain), Bilbao (NE Spain) and Pasajes (NE Spain). In an attempt to determine sediments toxicity, a combination of exposure biomarkers was analyzed in both species: metallothionein-like-proteins (MTLPs), ethoxyresorufin O-deethylase (EROD), glutathione S-transferase activity (GST), glutathione peroxidase (GPX) and glutathione reductase (GR). In parallel, physical and chemical characterization of the different sediments was performed and biological responses related to the contaminants. Significant induction of MTLPs was observed when organisms were exposed to metal contaminated sediments (port of Huelva), and EROD and GPX activities after exposure to sediments containing organic compounds (port of Bilbao and Pasajes). No significant interspecies differences were observed in biomarker responses except for the GST and GR.

Cadmium and heat response of the fungus Heliscus lugdunensis isolated from highly polluted and unpolluted areas

Induction of heat shock protein (Hsp) 70 and distinct metallothionein-like proteins (MTLPs) in response to Cd and heat treatment were studied in two strains of the aquatic hyphomycete Heliscus lugdunensis: Hl-H4, isolated from a heavy metal polluted site, and Hl-BB taken from an unpolluted area. Upon Cd-exposure, Hsp70 was actively synthesized in the strain Hl-H4, and to a much lower degree in the strain Hl-BB. The Hsp70-expression was time- and dose-dependent, reaching a maximum after 24 h incubation with 80 microM Cd. Upon heat-stress, a similar response was observed: a strong Hsp70-expression in Hl-H4, and only a marginal one in Hl-BB. The strains reacted to Cd-exposure by a specific, environmentally related induction of MTLPs, as shown by the highly sensitive bimane derivatisation method of SH-rich proteins. In Hl-H4, a strong expression of 11 kDa MTLP was registered, which followed strictly the induction pattern of Hsp70. This suggests interdependence of the induction mechanisms and roles of these stress proteins in metal resistance. On the contrary, in Hl-BB a weak expression of MTLP of about 20 kDa was observed, exhibiting completely different induction pattern. The results suggest that the specific induction of Hsp70 and/or distinct MTLPs in the range of 11 kDa in H. lugdunensis strain Hl-H4 are essential adaptive mechanisms to continuous heavy metal exposure.

Induction of Metallothionein-like Proteins by Mercury and Distribution of Mercury and Selenium in the Cells of Hepatopancreas and Gill Tissues in Mussel Mytilus galloprovincialis

The binding of mercury (Hg) to metallothioneins (MTs) and the relation between Hg and selenium in supernatants of hepatopancreas and gill tissues of the common mussel Mytilus galloprovincialis (Lamarck, 1819) was investigated. The mussels were exposed to different Hg concentrations in laboratory conditions: 2.5 microgHg/L, 4 d exposure (short term) and 60 microgHg/L, 33 d exposure (long term). In addition, the results were compared to those found for mussels from nature (polluted and unpolluted region). In control and short-term-exposed mussels, the level of Hg extraction (cytosol) from hepatopancreas and gill cells was very low with respect to the total Hg concentrations in the corresponding tissues, around 10% in control and around 20% after exposure. As expected, Hg exposure was followed by Se increase. For Se, the levels of extraction were higher, around 20% in control and up to 50% (heaptopacrease) of 70% (gills) after exposure. In order to study the distribution of Hg and Se in the cells of these organs, the total Hg and Se concentrations were analyzed in the subcellular fractions obtained after differential centrifugation. Although after exposure the concentrations of both element increased in all subcellular fractions, their percentages in particular fractions were lower or higher. In this study, the convincing binding of Hg to metallothionein-like proteins was perceived after long-term laboratory exposure (gills, heapatopancreas) and in wild mussels collected near industrial port (hepatopancreas). In latter case, we also detected the traces of Se bound to the MT fractions after size-exclusion chromatography.

Influences of maternal exposure on the tolerance and physiological performance of Daphnia magna under mercury stress

We examined the tolerance development to mercury (Hg) by a population of freshwater zooplankton (Daphnia magna) with different pre-exposure histories to Hg. The growth and reproductive performance of the F1 offspring as affected by the maternal (F0) and offspring (F1) exposures was quantified. The F0 daphnids exposed to 2.5 and 25 nM of Hg for 4 d and followed by 4 d of depuration had elevated levels of Hg and metallothionein-like proteins (MTLPs), as well as higher tolerance to Hg toxicity than the control daphnids. The higher Hg tolerance may be attributed to the higher proportion of Hg partitioned to the MTLPs. Moreover, significant enhancement of Hg tolerance also was found in the F1 offspring originating from the F0 mothers exposed to 25 nM of Hg, but there was no significant induction of MTLPs in these F1 offspring when compared to the offspring from the control mothers. The Hg tissue concentrations in the F1 neonates were approximately 25% of those in the F0 adults. However, there was similar Hg tolerance in the F2 offspring originating from both the control and Hg-exposed F0 mothers, indicating that the Hg tolerance in the daphnids disappeared two generations after Hg contamination. Further exposure of the F1 offspring to different Hg concentrations (1.5 and 15 nM for 28 d) indicated that maternal exposure history did not affect their growth and reproductive performance, which solely were influenced by the offspring exposure. Unexpectedly, the F1 offspring exposed to Hg had significantly higher final wet weights and reproductive rates than the control groups, suggesting the possibility of Hg hormesis. Furthermore, the maternal exposure had no effect on the Hg accumulation and the MTLP concentrations in the F1 offspring. Therefore, we concluded that the Hg tolerance might disappear quickly once the Hg contamination was removed and the maternal exposure history was not important in determining the physiological performance and Hg accumulation of the subsequent generations.

Tissue-specific cadmium accumulation and metallothionein-like protein levels during acclimation process in the Chinese crab Eriocheir sinensis

Aquatic organisms chronically exposed to cadmium can increase their resistance to a subsequent elevated exposure. In order to investigate mechanisms involved in acclimation process in the Chinese crab Eriocheir sinensis, we compared Cd level as well as metallothionein-like protein (MTLP) content in different tissues after direct acute exposure (i.e. 500 μg Cd L −1 for 3 days), and after acute following chronic (i.e. 10 or 50 μg Cd L −1 for 30 days) exposure. Cadmium accumulation occurred in the following order: anterior gill>hepatopancreas>posterior gill>carapace>hemolymph>muscle. As high concentrations as 188 μg Cd g −1 w.w. were reported in anterior gills and seem to reach a saturation level. In these gills, the highest MTLP induction was observed after a direct acute exposure, for which a correlation with Cd content occurred. However, the Cd-binding potential by MTLPs was exceeded for any exposure condition. In hepatopancreas, the highest Cd level was reported for crabs acclimated during 30 days to 50 μg Cd L −1 before challenging with an acute exposure. Moreover, we showed that MTLPs were induced during the acclimation process. In this organ, MTLPs are theoretically sufficient to bind all Cd. These results suggest that during a chronic exposure to 50 μg Cd L −1, Chinese crabs acquire the capacity to hold more cadmium in hepatopancreas where it can be sequestrated by MTLPs. On the contrary, MTLP induction seems to be a rapid response to acute exposure in anterior gill, but is not sufficient to sequester all Cd. Other sequestration and/or detoxification mechanisms must take place in anterior gill to cope with high Cd levels.

Uptake and depuration of cadmium, nickel, and lead in laboratory-exposed Tubifex tubifex and corresponding changes in the concentration of a metallothionein-like protein.

Based on weight loss in water, 24 h is recommended for Tubifex tubifex gut clearance. Biota-to-sediment accumulation factors (BSAFs) in gut-cleared T. tubifex following six weeks of exposure to Cd-, Ni-, and Pb-spiked sediment were 12.4, 3.0, and 19.0, respectively. Tissue Ni concentrations peaked after 12 h, whereas Cd and Pb were accumulated for the duration of the exposure. Tubifex tubifex were transferred to either water (24 h) or sediment (10 weeks) to monitor changes in internal metal concentrations. After 24 h in water, only Ni concentration had declined significantly (p < 0.05), suggesting that the majority of Ni was associated with the gut content, while Cd and Pb were accumulated in the tissues. Metal depuration in sediment was described with two-compartment, first-order kinetic models (r2 = 0.7-0.8; p < 0.001), indicating that T. tubifex has both a quickly depurated and a more tightly bound pool of accumulated metal. Tubifex tubifex were also exposed to sediment spiked with just Cd (3.66 micromol/g). Cadmium uptake and induction of metallothionein-like protein (MTLP) were rapid; both parameters were significantly elevated within 24 h of exposure. Metallothionein-like protein (8.7 +/- 1.8 nmol/g) and Cd (60.8 +/- 11.0 micromol/g) reached maximum concentrations after 96 h and four weeks, respectively.

Partitioning of accumulated trace metals in the talitrid amphipod crustacean Orchestiagammarellus: a cautionary tale on the use of metallothionein-like proteins as biomarkers

The induction of metallothionein-like proteins (MTLP) as biomarkers of trace metal pollution has been investigated in the talitrid amphipod Orchestia gammarellus, an established biomonitor of trace metal availabilities in coastal areas. Sensitivity to metal exposure, MTLP and copper and zinc concentrations have been examined in amphipods from a metal-contaminated site (Dulas Bay, Wales) and two clean sites (Millport, Scotland and Bradwell, England). Groups of 10 amphipods from Dulas Bay and Millport were exposed separately to one of a concentration series (1, 3.16, 10, 31.6, 100 μM) of Cu or Zn for 17 days at 10 °C. Specimens from Bradwell were exposed under similar conditions but the experiment was extended to 25 days with intermediate samplings at 5, 10 and 17 days and additional groups were exposed to a mixture of Cu+Zn (7:10 molar ratio). In addition, specimens from Millport were exposed to 1 or 31.6 μM Cd for 17 days. Amphipods from Dulas Bay, which had been chronically exposed to metals in their natural environment had not acquired any tolerance to Cu and Zn since the lowest LC 50s were registered in these samples. Whatever the origin of the amphipods experimentally exposed to metals and whatever the dose of exposure, both Cu and Zn remained approximately equally distributed between cytosolic and insoluble fractions in the amphipods, suggesting that mechanisms of metal storage were identical over the whole range of conditions. Concentrations of MTLP were higher in O. gammarellus from Dulas Bay than in those from Millport analysed directly after collection, although laboratory exposures to dissolved Cd, Cu or Zn have failed to demonstrate differences in the induction of MTLP between amphipods from the clean or contaminated sites. A potential role for metallothionein-like proteins as biomarkers is thus unlikely although it remains plausible that turnover of these proteins does increase in response to increased metal challenges, enabling MTLP to play a role in metal detoxification.