Toxicity of analytically cleaned pentabromodiphenylether after prolonged exposure in estuarine European flounder (Platichthys flesus), and partial life-cycle exposure in fresh water zebrafish (Danio rerio)

Residues of polybrominated diphenylethers (PBDEs), extensively applied as flame retardants, are widely spread in the aquatic environment and biota. The present study investigates effects of the environmentally relevant lower brominated diphenylethers in two fish species in vivo under controlled laboratory conditions. Euryhaline flounder (Platichthys flesus) and freshwater zebrafish (Danio rerio) were exposed to a range of concentrations of a commercial pentabromodiphenylether mixture, DE-71. Chemical analysis of exposed fish showed a pattern of PBDE congeners that was very similar to that in wild fish. The resulting range included environmentally relevant, as well as higher levels. Animals were investigated histopathologically with emphasis on endocrine and reproductive organs. In zebrafish, hatching of embryos and larval development were assessed. Biochemical parameters were investigated in flounder as markers for suggested dioxin-like activity (ethoxyresorufin-O-deethylase=EROD), and activation of endogenous estrogen synthesis (gonad aromatase activity). Thyroid hormones were analyzed in plasma in both species. Benchmark analysis using internal PBDE concentrations showed a mild dose-dependent decrease of hepatic EROD and ovarian aromatase activities, and plasma thyroxin levels in flounder, and an increase of plasma thyroid hormone levels in zebrafish. These trends did not result in statistically significant differences from control fish, and major histopathological changes were not observed. Reproduction in zebrafish appeared to be the most sensitive parameter with statistically significantly reduced larval survival and non-significant indications for decreased egg production at internal levels that were more than 55 times the highest environmental recordings. The present results indicate limited risk for endocrine or reproductive effects of current environmental PBDE contamination in fish.     

Accumulation pattern and biotransformation enzyme induction in rainbow trout embryos exposed to sublethal aqueous concentrations of 3,3',4,4'-tetrachlorobiphenyl

Accumulation pattern of 3,3',4,4'-tetrachlorobiphenyl (PCB 77) and the exposure time needed to activate the monooxygenase (EROD) and conjugation (GST) enzyme systems of fish at the advanced embryonic stage were studied. Eyed stage embryos of rainbow trout (Oncorhynchus mykiss) were exposed to sublethal doses (0, 1, 10, and 100 micrograms/l) of PCB 77. Results indicated direct accumulation of the chemical into the eggs, but the exposure time was not long enough for PCB 77 to reach constant steady state. However, at the two lowest test concentrations (1 microgram/l and 10 micrograms/l) a temporary plateau at chemical accumulation was reached at the third exposure day (185 and 1221 ng PCB/g egg w.w). At the highest concentration (100 micrograms/l) the decrease in the accumulation rate was already evident after the first day (2182 ng PCB/g egg w.w). The chemical uptake increased again at day 7 in all the exposure groups. That event could have been caused by the increased metabolic rate of the embryos in preparation for the upcoming hatching event. The microsomal CYP1A monooxygenase system (EROD) was shown to be a sensitive indicator of embryonic exposure, being induced at the low (1 microgram/l, 182 ng/g egg) PCB concentration and after a short (3 day) exposure time. The conjugation enzyme system (GST) was shown to be functioning already at the advanced embryonic stage, although no response to the studied chemical stress was detected.     

Microsomal monooxygenase activity in Tilapia (Oreochromis mossambicus) exposed to a bleached kraft mill effluent using different exposure systems.

Bleached kraft pulp and paper mill effluents (BKMEs) are known to have adverse effects on aquatic organisms. One of the effects of BKMEs is its ability to induce cytochrome P4501A activity in exposed fish. 7-Ethoxyresorufin O-deethylase (EROD) activity is the most common biomarker used to measure the mixed-function monooxygenase activity. In this study, Tilapia were exposed to BKMEs using different exposure systems and their hepatic EROD activity, as well as liver/somatic index (LSI), were determined. In the Phase I study, Tilapia treated with betaNF and a whole (100%) BKME using a static, non-renewal system exhibited statistically significant EROD induction, but LSI values were not altered. In the Phase II study, fish were either caged in the mill's fishpond with the whole effluent passing through or cultured in tanks receiving 100% of the BKME continuously using a flow-through system in the laboratory. Their EROD activities were then compared with the non-exposed fish (control). The EROD activities in both groups of fish were elevated significantly with the greatest induction being observed in the field-exposed group. The LSI values in all of the field-exposed fish were significantly greater than the control Tilapia. The EROD assay was sensitive in detecting biological changes in fish exposed to the BKME. Further studies are warranted to better understand the impacts of BKMEs on aquatic organisms in Taiwan.     

Comparative embryotoxicity of a pentabrominated diphenyl ether mixture to common terns (Sterna hirundo) and American kestrels (Falco sparverius)

Concentrations of polybrominated diphenyl ethers (PBDEs) in Forster’s tern (Sterna forsteri) eggs from San Francisco Bay have been reported to range up to 63 μg g⁻¹ lipid weight. This value exceeds the lowest-observed-adverse-effect level (1.8 μg g⁻¹ egg wet weight; ∼32 μg⁻¹ lipid weight) reported in an embryotoxicity study with American kestrels (Falco sparverius). As a surrogate for Forster’s terns, common tern (Sterna hirundo) eggs were treated by air cell injection with corn oil vehicle (control) or a commercial penta-BDE formulation (DE-71) at nominal concentrations of 0.2, 2, and 20 μg g⁻¹ egg. As a positive control, kestrel eggs received vehicle or 20 μg DE-71 g⁻¹ egg. In terns, there were no effects of DE-71 on embryonic survival, and pipping or hatching success; however, treated eggs hatched later (0.44 d) than controls. Organ weights, organ-to-body weight ratios, and bone lengths did not differ, and histopathological observations were unremarkable. Several measures of hepatic oxidative stress in hatchling terns were not affected by DE-71, although there was some evidence of oxidative DNA damage (8-hydroxy-deoxyguanosine; 8-OH-dG). Although DE-71 did not impair pipping and hatching of kestrels, it did result in a delay in hatch, shorter humerus length, and reduced total thyroid weight. Concentrations of oxidized glutathione, reduced glutathione, thiobarbituric acid reactive substances, and 8-OH-dG in liver were greater in DE-71-treated kestrels compared to controls. Our findings suggest common tern embryos, and perhaps other tern species, are less sensitive to PBDEs than kestrel embryos.     

Latent effects of early life stage exposure to triclosan on survival in fathead minnows,Pimephales promelas

The objective of this study was to evaluate the effects of early life stage triclosan (5-chloro-2-(2,4, dichlorophenoxy)phenol, TCS) exposure on hatching, development, and survival in the fathead minnow, Pimephales promelas. Embryonic minnows were exposed to TCS (50 and 100 µg L^?1) for 10 days followed by 6 weeks depuration. Mortality and morphological deformities were recorded and quantified during exposure and at the end of depuration. No significant effects on embryonic survival, time to reach the eyed stage, or hatching were found. However, at the conclusion of the depuration period, survival was significantly reduced in TCS exposed fish depending on the concentration. Visual inspection of the exposed fish suggests that mortality is related to spinal deformities, emaciation, and reduced foraging ability. Triclosan exhibits deleterious effects in fish at lower concentrations over longer durations than previously reported. Further, mortality in exposed fish 6 weeks after exposure demonstrates the need for various exposure assays to evaluate effects of TCS.     

Responses of juvenile sea bass, Dicentrarchus labrax, exposed to acute concentrations of crude oil, as assessed by molecular and physiological biomarkers

In the present study, juvenile sea bass were exposed for 48 and 96 h to an Arabian light crude oil and their responses were assessed at the molecular and physiological levels. The aim of the study was therefore to assess (i) the short term effects of crude oil exposure by the measurement of several molecular biomarkers, (ii) the consequences of this short term exposure on fish health by using growth and condition indices measured after a decontamination period of 28 and 26 d in seawater. Hydrocarbon petroleum concentrations was monitored during the 96 h experiments and an increase of PAH concentrations were found in fish following both exposure times. An 7-ethoxyresorufin-O-deethylase (EROD) induction was observed after 48 h of exposure, while a significant decrease in the sea bass specific growth rate in length and for the RNA:DNA ratio was observed 28 d after that exposure ceased. The EROD induction doubled after the 96 h exposure, and a significant increase in GST activities was observed. A significant decrease in the specific growth rates, the otolith recent growth, the RNA:DNA ratio and the Fulton’s K condition index were then observed in sea bass 26 d after the 96 h exposure to mechanically dispersed crude oil compared to the control. The present study shows that growth and condition indices can prove useful in assessing fish health status following an oil spill. Their complementary analysis with sensitive molecular biomarkers as EROD could improve the determination of oil spill impact on fish populations.     

A field study on EROD activity and quantitative hepatocytological changes in an immature demersal fish

Demersal fish, Solea ovata, were sampled from a reference site and a site where highly contaminated sediment is dumped. Sexually immature fish from the contaminated site exhibited significantly higher EROD activity compared with counterparts sampled from the reference site. No significant difference in EROD activity could be found for sexually mature males and females between sites. The relationship between EROD activity and quantitative changes in hepatic lipofuscin/ceroid, as well as peroxisome, was investigated for immature S. ovata. A significant correlation was found between EROD activity and volume density of lipofuscin/ceroid in fish hepatocyte (r = 0.750; P < 0.05), but no significant correlation was discernible between EROD activity and peroxisomes. Results from this field study corroborate our earlier laboratory findings, in which induction of EROD activity by intraperitoneal injection of benzo[a]pyrene was associated with increase in absolute volume and absolute number of lipofuscin/ceroid in hepatocytes. The present study provides further evidence that induction of EROD activity is associated with an increase in hepatic lipofuscin/ceroid and possibly cytological damages in immature S. ovata. This cytological change may serve as a potential marker for exposure to PAHs and PCBs.     

In vitro toxicity of selected pesticides on RTG-2 and RTL-W1 fish cell lines

The rainbow trout fish cell lines RTG-2 and RTL-W1 were used to determine the cytotoxic effects of the pesticides bifenthrin, cypermethrin, cyhalothrin, lambda-cyhalothrin, quinalphos and chlorpyrifos. Cytotoxicity was measured by EROD and beta-Gal enzymatic activities, the neutral red (NR) uptake assay, and the FRAME KB protein (KBP) assay. The beta-Gal activity was unaffected by the pesticide exposure. The EROD activity was induced by cyhalothrin and lambda-cyhalothrin (RTG-2 and RTL-W1) and by bifenthrin (RTL-W1). Dose dependent inhibition responses were observed for EROD activity in cells exposed to quinalphos (RTL-W1) and chlorpyrifos (RTG-2 and RTL-W1). RTL-W1 offered a better response for EROD induction. The EC50 values on EROD endpoint were more sensitive than NR and KBP. The acute fish toxicity of chlorpyrifos and quinalphos depends highly on the species; the species sensitivity distributions cover several orders of magnitude and the values obtained for EROS were within the lowest part of the reported ranges.     

EROD activity and biliary fluorescence in Schroederichthys chilensis (Guichenot 1848): biomarkers of PAH exposure in coastal environments of the South Pacific Ocean

Schroederichthys chilensis is a common shark that lives in Chilean coastal environments. In this work, the relationship between liver 7-ethoxyresorufin-O-deethylase dealkylation (EROD) activity and Fluorescent Aromatic Compounds (FAC) in bile of S. chilensis sampled in three bays with different degrees of pollution were performed including a reference area. Sixty individuals were collected, 20 for each site; (10 males and 10 females per site) livers and bile samples were obtained and immediately frozen. EROD activity and FAC were measured according to three standard methods. EROD activity and FAC were higher in polluted areas than in the reference area. Synchronous Fluorescence Spectra of the bile from the fish collected at the most polluted area showed a peak at 347nm representing a metabolite corresponding to 1-hydroxypyrene. The low EROD activity in the reference area is likely related to the low level of PAH in sediments. We propose that this species is a good indicator of exposure to FACs, since it presents a series of characteristics that make it suitable for monitoring PAH exposure in coastal zones.     

Vitamin and thyroid status in arctic grayling (Thymallus arcticus) exposed to doses of 3,3',4,4'-tetrachlorobiphenyl that induce the phase I enzyme system

Induction of phase I biotransformation enzymes is recognized as a hallmark response in fish exposed to coplanar PCBs. Depletions of vitamins A and E and disrupted thyroid hormone and glandular structure secondary to this induction have not yet been examined in an arctic fish species. Arctic grayling were exposed to a single oral dose of 0 (control), 10, 100 or 1000 ng 3,3',4,4'-tetrachlorobiphenyl (TCB) g(-1) bodyweight, a contaminant found in most arctic fish. After 30 and 90 days of exposure, TCB concentrations in tissues, hepatic phase I activity (as ethoxyresorufin-O-deethylase (EROD)), plasma and tissue vitamin A and E concentrations, plasma thyroid hormone levels and thyroid glandular structure were examined. Total plasma osmolality, as an indicator of overall fish health was also monitored. TCB recovery in tissues was low and extremely variable, making comparisons between intended dose groups inappropriate. Therefore, correlation analysis between actual recovered TCB concentrations and biochemical responses was employed. Hepatic EROD activity correlated strongly with liver TCB concentrations. Liver concentrations of vitamin A were altered as a function of TCB concentrations and EROD activity, but plasma vitamin A status was not affected. Vitamin E was depleted by TCB accumulation in blood and EROD induction in liver of males only at 90 days postexposure. Thyroid hormones status and glandular structure were not affected by the short duration TCB exposures used in this experiment. TCB concentrations were correlated with an elevation in plasma osmolality. Results from this experiment indicate that the vitamin status and osmoregulation of arctic grayling exposed to TCB can be compromised. Further studies of field populations exposed to this type of contaminant are warranted.     

Monitoring contaminants from oil production at sea by measuring gill EROD activity in Atlantic cod (Gadus morhua)

An ex vivo gill EROD assay was applied in Atlantic cod (Gadus morhua) as a biomarker for waterborne CYP1A-inducing compounds derived from oil production at sea. Exposure to nominal concentrations of 1 ppm or 10 ppm North Sea crude oil in a static water system for 24 h caused a concentration-dependent gill EROD induction. Further, exposure of cod for 14 days to environmentally relevant concentrations of produced water (PW, diluted 1:200 or 1:1000) from a platform in the North Sea using a flow-through system resulted in a concentration-dependent induction of gill EROD. Crude oil (0.2 ppm) from the same oil field also proved to induce EROD. Finally, gill EROD activity in cod caged for 6 weeks at 500-10 000 m from two platforms outside Norway was measured. The activities in these fish were very low and did not differ from those in fish caged at reference sites.     

EROD activity and stable isotopes in seabirds to disentangle marine food web contamination after the Prestige oil spill

In this study, we measured via surgical sampling hepatic EROD activity in yellow-legged gulls from oiled and unoiled colonies, 17 months after the Prestige oil spill. We also analyzed stable isotope composition in feathers of the biopsied gulls, in an attempt to monitor oil incorporation into marine food web. We found that yellow-legged gulls in oiled colonies were being exposed to remnant oil as shown by hepatic EROD activity levels. EROD activity was related to feeding habits of individual gulls with apparent consequences on delayed lethality. Capture-recapture analysis of biopsied gulls suggests that the surgery technique did not affect gull survival, giving support to this technique as a monitoring tool for oil exposure assessment. Our study highlights the combination of different veterinary, toxicological and ecological methodologies as a useful approach for the monitoring of exposure to remnant oil after a large oil spill.     

Monitoring PAHs in the Brisbane River and Moreton Bay, Australia, using semipermeable membrane devices and EROD activity in yellowfin bream, Acanthopagrus australis

Two water quality monitoring strategies designed to sample hydrophobic organic contaminants have been applied and evaluated across an expected concentration gradient in PAHs in the Moreton region. Semipermeable membrane devices (SPMDs) that sequester contaminants via passive diffusion across a membrane were used to evaluate the concentration of PAHs at four and five sites in spring and summer 2001/2002, respectively. In addition, induction of hepatic cytochrome P4501, EROD activity, in yellowfin bream, Acanthopagrus australis, captured in the vicinity of SPMD sampling sites following deployment in summer was used as a biomarker of exposure to PAHs and related chemicals. SPMDs identified a clear and reproducible gradient in PAH contamination with levels increasing from east to west in Moreton Bay and upstream in the Brisbane River. The highest PAH concentrations expressed as B(a)P-toxicity equivalents (TEQs) were found in urban areas, which were also furthest upstream and experienced the least flushing. Cytochrome P4501 induction in A. australis was similar at all sites. The absence of clear trends in EROD activity may be attributable to factors not measured in this study or variable residency time of A. australis in contaminated areas. It is also possible that fish in the Moreton region are displaying enzymatic adaptation, which has been reported previously for fish subjected to chronic exposure to organic contaminants. These potential interferences complicate interpretation of EROD activity from feral biota. It is, therefore, suggested that future monitoring combine the two methods by applying passive sampler extracts to in vitro EROD assays.     

Some factors influencing EROD activity in winter flounder (Pleuronectes americanus) exposed to effluent from a pulp and paper mill

EROD (ethoxyresorufin-O-deethylase) activity was determined in winter flounder, a sediment-inhabiting and non-migratory fish species, living near a pulp and paper mill in Newfoundland in relation to temperature, gender, sexual maturity, and lesions in the liver. Samples of liver were taken from fish captured by SCUBA divers at 0 degrees C, 5 degrees C and 16 degrees C. Enzymic activity was detected in fish living only above 0 degrees C. Adult males and juvenile fish had higher levels of EROD activity than prespawning females at 5 degrees C. Macrophage aggregates only or occurring simultaneously with bile ductule hyperplasia and clear cell foci in the liver, did not impair EROD activity but necrosis had a negative effect. Results from this study indicate the importance of water temperature, gender, sexual maturity and liver pathology in assessing EROD activity of fish in biomonitoring programs.     

Temporal induction pattern of hepatic cytochrome P450 1A in thermally acclimated dab (Limanda limanda) treated with 3,3′,4,4′-Tetrachlorobiphenyl (CB77)

Mature male dab (Limanda limanda) acclimated at 10° and 16°C were orally administered a single dose of 0.5 mg/kg 3,3′,4,4′-tetrachlorobiphenyl (CB77). At both temperatures, levels of cytochrome P450 1A (CYP1A) protein and 7-ethoxyresorufin O-deethylase (EROD) activity showed a two to six fold induction 40 days after CB77 treatment compared to control groups. Maximum responses of both EROD activity and CYP1A protein for the warm-acclimated fish were observed at 5 days after treatment. For the cold-acclimated fish a slow, progressive elevation for both EROD activity and CYP1A protein was observed and maximum responses were measured 40 days after treatment. Absolute EROD activity and CYPIA protein levels of fish from both temperatures were equally high at 40 days after treatment. Since in the control groups EROD activity and CYP1A protein levels were higher in the cold-acclimated fish, the magnitude of induction was higher in the warm acclimated ones. The highest concentrations of CB77 in muscle of fish from both temperatures were found at 5 and 10 days after treatment. The liver somatic index (LSI) showed 1.5 fold significantly higher values for the fish acclimated at 10°C.     

Comparing the response of biochemical indicators (biomarkers) and biological indices to diagnose the ecological impact of an oil spillage in a Mediterranean river (NE Catalunya, Spain)

Three biomarkers of hydrocarbon exposure, liver 7-ethoxyresourfin-O-deethylase activity (EROD), fluorescent hydrocarbon compounds (FACs) in bilis, and the liver antioxidant enzyme catalase (CAT) were examined in the autochthonous fish species Barbus meridionalis collected in the river Fluvià (NE Catalunya, Spain) after an oil spillage. Four different locations were sampled, including the impacted site, upstream and downstream sites and a reference site. Biomarker responses were compared with diatom and macroinvertebrate community assemblage metrics (Specific Pollution Sensitivity index - IPS, and Iberian Biological Monitoring Working Party - IBMWP, respectively). Chemical analyses denoted that polycyclic aromatic hydrocarbon levels in sediment were much higher at the impacted site than in downstream reaches. Four fold increase of EROD activity together with increased levels of biliary FACs in barbs collected at the spilled site indicated exposure of inhabiting fish to the oil. Additionally, CAT activity was significantly depressed (four fold) when compared to other stations, thus suggesting that fish collected from the impacted sites could be more susceptible to suffer oxidative stress. Biological indices (particularly that of the diatom community IPS) showed slight significant effects between control and impacted sites, indicating that more tolerant taxa were favoured because of the oil spillage. These results support the need to include biochemical responses measured in local species in monitoring programmes aimed to diagnose specific pollution effects in stressed river ecosystems.     

Biological effect monitoring in dab (<Emphasis Type="Italic">Limanda limanda</Emphasis>) using gene transcript of CYP1A1 or EROD—a comparison

BACKGROUND, AIM, AND SCOPE: Gene expression analyses with real-time (RT)-polymerase chain reaction (PCR) gains importance in marine monitoring. This new technique has to be compared to the classical approaches like the well known biomarker ethoxyresorufin-O-deethylase (EROD) to test their suitability for monitoring programmes. The goal of the present study is to compare EROD activity and CYP1A1 mRNA expression in the important monitoring fish species dab (Limanda limanda) and to answer the question of whether these parameters reflect the polycyclic aromatic hydrocarbon (PAH) contamination of the fish. Further on, glyceraldehyd-3-phosphate dehydrogenase (GAPDH) was investigated as a potential housekeeping gene. MATERIALS AND METHODS: Female dab were caught in the summer of 2004 in the North Sea and in the Baltic. EROD activity was determined in liver samples by a kinetic fluorimetric assay according to a standard protocol. The gene expression of CYP1A (cytochrome P450 1A) and GAPDH were determined by means of RT-PCR. Results were compared to gonado somatic index and to the concentration of PAH metabolite 1OHPyr (1-hydroxypyrene) analysed in the bile fluids of the fish, respectively. RESULTS: Dab from all stations showed a considerable individual variation in the levels of both CYP1A mRNA and EROD. Highest mean values for CYP1A mRNA and EROD were detected in the northern part of the sampling area. In contrast, the PAH metabolite 1OHPyr was found at the highest concentration in fish caught near the German coast. CYP1A mRNA and EROD showed only a minor but significant correlation (r = 0.32, p < 0.05, n = 123). 1OHPyr in bile correlated significantly (p < 0.05) with the amount of GAPDH mRNA content in the liver. DISCUSSION: The significant but low correlation of CYP1A mRNA and EROD activity on an individual basis illustrates that these two parameters are apparently not closely linked. However, maximum EROD values correspond with maximum CYP1A mRNA concentrations when station means are regarded. Because EROD and CYP1A mRNA in dab follow different physiological principles, their application will lead to related but not identical monitoring results. This should be taken into account when future marine monitoring programmes are designed. The results also indicate that PAH are not the crucial factor for CYP1A and EROD levels in dab from the off-shore areas in the North Sea. This is remarkable because the PAH metabolism is known to be CYP1A-dependent and the widely used biomarker EROD has been recommended for monitoring PAH-related effects in fish from the North Sea. Due to a correlation between GAPDH and 1OHPyr, GAPDH was not suitable as housekeeping gene for dab. CONCLUSIONS: Neither the results from EROD nor from CYP1A1 mRNA measurements in dab reflected their exposure to PAH as measured by the PAH metabolite 1OHPyr. Thus, the question arises of whether EROD or CYP1A mRNA is a suitable biomarker at all to indicate PAH exposure in dab from the open North Sea. RECOMMENDATIONS AND PERSPECTIVES: For future biological effect monitoring, it is advisable to measure more and predominately independent parameters by RT-PCR and to incorporate more components of the detoxification system.     

Inhibition of metamorphosis in tadpoles of Xenopus laevis exposed to polybrominated diphenyl ethers (PBDEs)

Tadpoles of the African clawed frog, Xenopus laevis were exposed, beginning at stage 50, to a commercial pentabromodiphenyl ether mixture (DE-71) through the diet. Subsequent experiments were conducted using a single intraperitoneal injection at stage 58 with limited quantities of two purified brominated diphenyl ether (BDE) congeners, BDE47 and BDE99 and DE-71 to determine the relative potency of these BDE congeners within the commercial mixture. Significant inhibition of tail resorption, delayed metamorphosis and impacts on skin pigmentation were observed in Xenopus exposed to DE-71 in the diet at nominal doses of 1000 and 5000 microgg(-1) of food. The estimated time required for 50% of the tadpoles to complete metamorphosis was significantly lengthened in Xenopus exposed to a dietary concentration of 1 microg DE-71 per gram of food. Analysis of PBDEs (sum of 32 congeners) in Xenopus from the treatment with 5000 microgg(-1) of DE-71 indicated that the frogs accumulated an average of 1030 microgg(-1) (wet weight) of PBDEs. In the intraperitoneal injection trials, similar inhibitory responses were observed in Xenopus injected with DE-71 at a nominal dose of 60 microg per tadpole, or injected with BDE47 at a nominal dose of 100 microg per tadpole. No responses were observed in Xenopus injected with BDE99 at doses up to 100 microg per tadpole. Complete inhibition of metamorphosis was observed only in the highest DE-71 dietary treatment. The results of this study are consistent with a mechanism of action of PBDEs involving competitive inhibition of binding of thyroid hormones to transporter proteins, although the mechanism cannot be definitively determined from this study. The observed effects may have occurred through other mechanisms, including sublethal toxicity. The doses used in this study are greater than the levels of PBDEs to which anurans are exposed in the environment, so further studies are required to determine whether exposure to PBDEs at environmentally relevant concentrations can affect frog metamorphosis.     

Tributyltin causes abnormal development in embryos of medaka, Oryzias latipes

We examined the effects of tributyltin (TBT) on embryonic development, hatching success and sexual differentiation in Japanese medaka (Oryzias latipes). Embryos (within 8h after fertilization) were exposed to TBT in ovo via nanoinjection at concentrations of 0 (control), 0.16, 0.80, 3.96, 19.2 and 82.1 ng/egg. Embryonic survival, development and hatching were observed. Hatched fry were reared until 60 days when they sexually matured, and sexual differentiation was also examined by accordance of genetic and phenotypic sex, based on existence of DMY (a male determining gene in medaka) and secondary sex characteristics. As results, TBT caused a concentration-dependent mortality and impaired the embryonic development. However, no masculinization was detected at 60 dph medaka adults. Lowest observed effective concentration for inducing abnormal embryonic development was estimated to 0.16 ng/egg (ca. 160 ng/g egg).     

Induced cytochrome P450 1A activity in cichlid fishes from Guandu River and Jacarepaguá Lake, Rio de Janeiro, Brazil

The induction of cytochrome P4501A-mediated activity (e.g. ethoxyresorufin-O-deethylation, EROD) has been used as a biomarker for monitoring fish exposure to AhR-receptor ligands such as polycyclic aromatic hydrocarbons (PAH), polychlorinated biphenyls (PCB) and polychlorinated dibenzo-dioxins/furans (PCDD/Fs). In this study we found that hepatic EROD is induced in fish ("Nile tilapia", Oreochromis niloticus and "acará", Geophagus brasiliensis) from the Guandu River (7-17-fold) and Jacarepaguá Lake (7-fold), Rio de Janeiro, Brazil. Since both cichlid fish are consumed by the local population and the Guandu River is the main source of the drinking water supply for the greater Rio de Janeiro metropolitan area, pollution by cytochrome P4501A-inducing chemicals is a cause for concern and should be further investigated in sediments, water and biota. We additionally showed that EROD activity in the fish liver post-mitochondrial supernatant-simpler, cheaper and less time consuming to prepare than the microsomal fraction-is sufficiently sensitive for monitoring purposes.