Gut bacterial community structure (Porcellio scaber, Isopoda, Crustacea) as a measure of community level response to long-term and short-term metal pollution

Prokaryotes are of high importance in the assessment of environmental pollution effects. Due to fast responsiveness of bacterial communities to environmental physicochemical factors, it is difficult to compare results of bacterial community investigations on the temporal and spatial scale. To reduce the effects of variable physicochemical environmental conditions on bacterial microbiota when investigating the specific impact of contaminants on bacterial communities, we investigated the bacterial community in the gut of terrestrial isopods (Porcellio scaber, Isopoda, Crustacea) from clean and metal-polluted environments. Animals were collected from a chronically mercury-polluted site, a chronically multiple metal-(Cd, Pb, Zn) polluted site, and two reference sites. In addition, animals from an unpolluted site were laboratory exposed to 5 microg Hg/g food in order to compare the effect of acute and chronic Hg exposure. The bacterial gut microbiota was investigated by temporal temperature gradient gel electrophoresis (TTGE) and clone library construction based on polymerase chain reaction amplified 16S rRNA genes. The major bacterial representatives of the emptied gut microbiota in the animals from the chronically polluted environments seemed not affected when analyzed by TTGE. The detailed bacterial community structure investigated by 16S rRNA clone library construction, however, showed that the community from the Hg-polluted site also was affected severely (242.4 operational taxonomic units [OTU] in the polluted and 650.6 OTU in the unpolluted environment). When animals were acutely exposed to mercury, changes of bacterial community structures already were seen on TTGE profiles and no additional analysis was needed. We suggest the use of P. scaber gut bacterial community structure as a measure of effects caused by both long- and short-term exposure to pollution.     

Lysosomal membrane stability in laboratory- and field-exposed terrestrial isopods Porcellio scaber (Isopoda, Crustacea)

Two established methods for assessment of the cytotoxicity of contaminants, the lysosomal latency (LL) assay and the neutral red retention (NRR) assay, were successfully applied to in toto digestive gland tubes (hepatopancreas) of the terrestrial isopod Porcellio scaber (Isopoda, Crustacea). In vitro exposure of isolated gland tubes to copper was used as a positive control to determine the performance of the two methods. Lysosomal latency and the NRR assay were then used on in vivo (via food) laboratory-exposed animals and on field populations. Arbitrarily selected criteria for determination of the fitness of P. scaber were set on the basis of lysosomal membrane stability (LMS) as assessed with in toto digestive gland tubes. Decreased LMS was detected in animals from all polluted sites, but cytotoxicity data were not in agreement with concentrations of pollutants. Lysosomal membrane stability in the digestive gland tubes of animals from an environment in Idrija, Slovenia that was highly polluted with mercury (260 microg/g dry wt food and 1,600 microg/g dry wt soil) was less affected than LMS in laboratory animals fed with 5 and 50 microg Hg/g dry weight for 3 d. This probably indicates tolerance of P. scaber to mercury in the mercury-polluted environment and/or lower bioavailability of environmental mercury. In animals from the vicinity of a thermal power plant with environmental mercury concentrations three to four orders of magnitude lower than those in Idrija, LMS was severely affected. In general, the LL assay was more sensitive than the NRR assay. The LMS assay conducted on digestive gland tubes of terrestrial isopods is highly recommended for integrated biomarker studies.     

An iron-rich diet protects the liver and kidneys against cadmium-induced injury in the bank vole (Clethrionomys glareolus)

The objective of this study was to determine whether supplemental dietary iron (Fe) would protect against cadmium (Cd)-induced injury in the liver and kidneys of bank voles. The rodents were provided, for 6 weeks, Fe-adequate (60-80 microg/g) and Fe-enriched (250-270 microg/g) diets containing 0.05 (control), 40, and 80 microg Cd/g. Histological examinations and analyses of Cd, Cd bound and not bound to metallothionein (MT), Fe, and lipid peroxidation in liver and kidneys were carried out. The Fe-enriched diet prevented Cd-induced histopathological changes as well as deprivation of tissue Fe and lipid peroxidation. Also, supplemental Fe significantly decreased hepatic and renal Cd burden. However, in the Cd-80 bank voles fed the Fe-enriched diet, the non-MT-bound Cd, considered a toxic species, reached 4.7 microg/g liver and 13.7 microg/g kidney, these values being similar to those at which histopathological changes occurred in the voles fed Cd diets not supplemented with Fe. The data indicate that the protective effect of supplemental Fe in the bank vole may be due to the prevention of Cd-induced deprivation of tissue Fe and Fe-dependent oxidative processes rather than to reduction of cadmium accumulation.     

Precocious gut maturation and immune cell expansion by single dose feeding the lectin phytohaemagglutinin to suckling rats

The dietary lectin phytohaemagglutinin (PHA) induces gut growth and precocious maturation in suckling rats after mucosal binding. The present study investigated the dose range in which PHA provokes gut maturation and if it coincided with immune activation. Suckling rats, aged 14 d, were orogastrically fed a single increasing dose of PHA: 0 (control), 2, 10, 50 or 250 microg/g body weight (BW) in saline. The effect on gut, lymphoid organs and appearance of CD3+ (T-lymphocyte) and CD19+ (B-lymphocyte) cells in the small-intestinal mucosa was studied at 12 h (acute) and 3 d (late phase) after treatment. The low PHA doses (2 and 10 microg/g BW) induced intestinal hyperplasia without mucosal disarrangement but did not provoke gut maturation. Only the high PHA doses (50 and 250 microg/g BW) temporarily disturbed the intestinal mucosa with villi shortening and decrease in disaccharidase activities, and later after 3 d provoked precocious maturation, resulting in an increase in maltase and sucrase activities and decrease in lactase activity and disappearance of the fetal vacuolated enterocytes in the distal small intestine. Exposure to the high, but not to the low, PHA doses increased the number of mucosal CD19+ and CD3+ cells in the small intestine after 12 h, a finding also observed in untreated weaned rats aged 21-28 d. In conclusion, there was a dose-related effect of PHA on gastrointestinal growth and precocious maturation that coincided with a rapid expansion of mucosal B- and T-lymphocytes, indicating a possible involvement of the immune system in this process.     

Calcium/cadmium interactions at uptake surfaces in rainbow trout: waterborne versus dietary routes of exposure

Juvenile rainbow trout (Oncorhynchus mykiss) were exposed to control, 3 microg/L waterborne Cd, or 500 mg/kg dietary Cd in combination with either a control (20 mg/g Ca2+ as CaCO3) or elevated (60 mg/g Ca2+) Ca2+ diet for 28 d. No mortality or growth effects were observed in response to either route of Cd exposure, although fish fed Ca2+-supplemented diets exhibited minor reductions in growth within the first few days of feeding. Waterborne and dietary Cd resulted in significant Cd accumulation in most tissues, with dietary uptake being far in excess of waterborne under the exposure conditions used. The order of Cd accumulation strongly reflected the exposure pathway, being gill and kidney > liver > gut > carcass (waterborne Cd); gut > kidney > liver > gill > carcass > bone (dietary Cd). On a whole-body basis, the net retention of Cd from the diet was < 1%, indicating that the gut wall forms an important protective barrier reducing Cd accumulation into internal tissues. Dietary Ca2+ supplementation reduced short-term whole-body uptake rates of waterborne Ca2+ and Cd by >50% and resulted in much lower chronic accumulation of Cd (via the water and diet) in target tissues. Results suggest that Ca2+ and Cd share common pathway(s)/transport mechanism(s) in the gill and gut and that increased gastrointestinal Ca2+ uptake likely caused downregulation of branchial and gastrointestinal Ca2+ and therefore Cd uptake pathways. Because nutrient metals other than Ca2+ may also influence Cd (and other metal) uptake, new regulatory approaches to metal toxicity (e.g., biotic ligand model) require understanding of the influence of dietary status on metal accumulation.     

Marginal nutritional status of zinc, iron, and calcium increases cadmium retention in the duodenum and other organs of rats fed rice-based diets

Dietary minerals Zn, Fe, and Ca are antagonistic to Cd absorption. We showed earlier that rats fed a rice-based diet with a marginal content of these nutrients absorbed more Cd than rats fed adequate Zn-Fe-Ca (Environ. Sci. Technol., 36 (2002) 2684-2692). The present experiment was designed to determine the effects of marginal dietary Zn, Fe, and Ca on the uptake and turnover of Cd in the gastrointestinal tract. Two groups of weanling female rats (six per treatment) were fed a diet containing 40% cooked, dried rice containing 0.6 mg Cd/kg. The diet of one group contained adequate Zn (35 mg/kg), Fe (30 mg/kg), and Ca (5000 mg/kg), while that of the other group contained marginal Zn (6 mg/kg), Fe (9 mg/kg), and Ca (2500 mg/kg). Rats were fed for 5 weeks and then orally dosed with 1g of diet containing rice extrinsically labeled with 109Cd. From 0.25 to 64 days after dosing, 109Cd and total Cd concentrations were determined in intestinal segments. Shortly after dosing, 109Cd, as a percentage of the dose, was about 4 times higher in the duodenum of marginally fed rats than in that of control rats (10% vs 40%, respectively). Sixty-four days after dosing, 109Cd was 10 times higher in marginally fed rats than in controls; however, of the amount at day 1, <0.1% remained at day 64. After 5 weeks, the concentration of elemental Cd in the duodenum of the marginally fed rats was 8 times higher than that of control rats (24 microg/g dry wt. vs 2.9 microg/g dry wt., respectively). Cd concentrations in liver and kidney were 5 times higher in the marginally fed rats than those in controls (liver, 0.81 microg/g dry wt. vs 0.14 microg/g dry wt.; kidney, 4.7 microg/g dry wt. vs 0.92 microg/g dry wt., respectively). These data suggest that marginal intakes of Zn, Fe, and Ca cause the accumulation of Cd in the duodenum, which results in a greater rate of Cd absorption and a greater accumulation in the internal organs. Results are discussed in relation to mineral nutrient status and risk assessment of Cd in natural food sources.     

Effects of dietary Cd and Cu on feeding and growth rates of the landsnail Helix engaddensis

Effects of dietary cadmium and copper on feeding and growth rates of adult Helix engaddensis snails were studied for a 1-month period. The concentrations of cadmium were 0, 50, 100, 200, 400, and 800 microg Cd/g dry food, while those of copper were 0, 4, 20, 100, 500, and 2500 microg Cu/g dry food. Both metals inhibited feeding and growth rates significantly after 1 week and 3 weeks of exposure to Cd and Cu, respectively. Inhibition caused by Cd was found to be irreversible, which indicates toxicity, while that of Cu was reversible, indicating starvation because snails identified and refused to consume contaminated food. After 4 weeks of exposure, the NOEC of Cd was 50 microg/g and the LOEC was 100 microg/g, while those for Cu were 20 and 100 microg/g, respectively. EC50,75,90,100 (growth and feeding) values were calculated, and indicated that H. engaddensis snails are suitable for short-term toxicity bioassays.     

The importance of the gut and its contents in prey as a source of cadmium to predators

Many mammalian and avian predators consume the whole carcass of their small mammal prey. Thus, the contaminant load of the gut and its contents may be of toxicological significance, but pollutant loads in the gut of prey species are often not measured and their role in food-chain transfer of contaminants is largely unquantified. We investigated the importance of the gut and its contents in prey as a source of cadmium (Cd) to predators by measuring the Cd concentrations in the guts (including contents), in various body tissues, and in the intact bodies of wood mice (Apodemus sylvaticus). The mice had been fed for up to 365 d on either control, a medium-Cd (MCd) diet, or a high-Cd (HCd) diet that had nominal Cd concentrations of 0, 8, and 40 g/g wet weight, respectively. In wood mice fed contaminated diets, Cd was progressively bioaccumulated in all the body tissues that were analyzed. The gut (and contents) and the liver and kidneys (the critical organs) contained most of the accumulated Cd and were the only significant contributors to the total body burden (TBB). The contribution of the gut (and contents) to the TBB was greater in mice fed MCd diet than in those given HCd feed but declined with age (equivalent to exposure duration) in both groups as Cd in the critical organs increasingly became the most important source of TBB Cd. However, the gut and its contents contributed between 50 and 100% of the Cd TBB in mice fed for up to six months on MCd diet; this was the diet that most closely mimicked the estimated average Cd intake of wood mice on contaminated sites. The results of this study highlight the importance of including measures of Cd load in the gut when modeling food-chain transfer from small mammals to their predators.     

Expression of the mouse metallothionein-I and -II genes provides a reproductive advantage during maternal dietary zinc deficiency.

The function of metallothionein in zinc homeostasis was examined by using mice homozygous for knockout (KO) of the metallothionein-I or -II (MT-I and MT-II) genes. Pregnant MT-I/II KO mice or control mice were fed a zinc-deficient (1 microg/g or 5 microg/g) diet or a zinc-adequate (50 microg/g) diet during specific periods of pregnancy, and the effects on morphogenesis of the embryos were determined at day 14 of pregnancy (day 1 = vaginal plug). In the homozygous MT-I/II KO, as well as in the nontransgenic control mice, severe dietary zinc deficiency (1 microg/g) beginning on day 1 of pregnancy was embryotoxic and teratogenic, and the majority of the embryos in both strains were dead by mid-gestation. However, 53% of the surviving embryos in the MT-I/II KO mice were morphologically abnormal compared to only 32% of the embryos in the control mice. In subsequent experiments, moderate dietary zinc deficiency (5 microg/g beginning on day 1 of pregnancy or 1 microg/g dietary zinc beginning on day 8 of pregnancy) exerted teratogenic, but not embryotoxic effects. Embryos in the MT-I/II KO mice were 260 to 290% as likely to develop abnormally than were embryos in the control mice fed these same diets. These results demonstrate that the expression of the MT-I and -II genes in pregnant females improves reproductive success during maternal dietary zinc deficiency.     

The assessment of episodic metal pollution. II. The effects of cadmium and copper enriched diets on tissue contaminant analysis in rainbow trout (Oncorhynchus mykiss)

Rainbow trout, Oncorhynchus mykiss, were fed on a commercial feed enriched with either Cd or Cu for 32 days and allowed to recover on normal food for 12 days. The body burden of fish fed Cd-enriched diet increased from about 0.05 to 0.39 g/g wet weight, reflected by elevation in the Cd content of gill, liver, blood plasma, kidney, skin, mucus, and whole gut. Muscle Cd content did not alter. Most organs remained contaminated with Cd after 12 days depuration. The body burden of animals fed Cu-enriched diet increased from about 1.2 to 1.5 g/g wet weight, with the gill, whole gut, blood plasma, skin, and mucus contaminated after 32 days, no changes in the liver and muscle occurred. All organs, except the gill and kidney, returned to control values after 12 days depuration. In exposed animals, 76 and 53% of the Cd or Cu body burden, respectively, were contained in the gut, unlike waterborne exposure. Differences in the body distribution of toxicants after intermittent waterborne and dietary exposure may identify the principal routes of pollutant uptake. Ratios of toxicant concentrations in gill/liver may establish the exposure status of fish, but cannot be used to differentiate dietary and waterborne contamination. Analysis of mucus may identify waterborne Cu exposure; dietary and waterborne contributions to mucus Cd content are more difficult to establish.     

Subcellular cadmium distribution, accumulation, and toxicity in a predatory gastropod, thais Clavigera, fed different prey

Bioaccumulation and toxicity of Cd were investigated in a marine predatory whelk, Thais clavigera, after being fed with the rock oyster, Saccostrea cucullata, or the herbivorous snail, Monodonta labio, for up to four weeks. The oysters and snails had different subcellular Cd distributions and concentrations in their bodies given their different metal-handling strategies and were exposed to dissolved Cd for two weeks before being fed to the whelks. After four weeks of dietary exposure, the Cd body concentrations in T. clavigera increased from 3.1 microg/g to between 22.9 and 41.8 microg/g and to between 22.7 and 24.1 microg/g when they were fed with oyster and snail prey, respectively. An increasing proportion of Cd was found to be distributed in the metallothionein (MT)-like proteins and organelle fractions, whereas the relative distribution in the metal-rich granules fraction decreased when the whelks were fed Cd-exposed prey. At the highest Cd dosage, more Cd was distributed in the pool of metal-rich granules when the whelks were fed the oysters than when they were fed the snails. Among all the biomarkers measured (MT induction, condition index, lipid peroxidation, and total energy reserve including carbohydrate, lipid, and protein), only MT showed a significant difference from the control treatments, and MT was the most sensitive biomarker for dietary Cd exposure. No toxicity was found in the whelks fed different Cd-exposed prey as revealed by various biomarkers at the different biological levels. Our results imply that metal fractionation in prey can alter the subsequent subcellular metal distribution in predators and that dietary Cd toxicity to the whelks was low, even when the accumulated Cd body concentrations were high.     

How terrestrial snails can be used in risk assessment of soils

Among soil invertebrates, terrestrial snails are herbivorous and detritivorous organisms exposed to polluted soils by both digestive and cutaneous routes. Using laboratory-reared snails (Helix aspersa aspersa), we describe how the effects of contaminants on survival and growth of snails can be evaluated in laboratory bioassays. A national ring test was performed to assess the effect of Cd added to the soil or to the food. The ecotoxicity of sewage sludge also was evaluated. The present results demonstrate that toxicity depends on both the pollutants and the exposure route. Cadmium was sixfold more toxic for snails exposed via food contamination (median effective concentration [EC50], 68-139 microg/g) than via soil contamination (EC50, 534-877 microg/g), whereas the opposite occurred with the sewage sludge (EC50, 55% of sludge in the food and 10% of waste in the soil). A logistic relationship linked growth inhibition and internal Cd concentrations, which can reach 2,000 microg/g in the viscera of snails exposed to 626 microg/g in the food. No clear trend was found between Cu, Zn, Pb, Cr, and Ni concentrations in the sludge and in snail tissues. These data enabled the development of an international standard, which should enhance the use of terrestrial gastropods for both fundamental research and routine risk assessment in the terrestrial environment.     

Mineral status of female rats affects the absorption and organ distribution of dietary cadmium derived from edible sunflower kernels (Helianthus annuus L.)

The intake of food cadmium (Cd) in microg/day over time can increase the body burden of this element. Some human populations that consume subsistence rice-based diets low in calcium (Ca), iron (Fe), and zinc (Zn) are more susceptible to Cd poisoning than populations that consume more nutritious diets. This study determined the effects of marginal deficiencies of these essential elements on the absorption and organ retention of Cd from a natural food that contains Cd, edible sunflower kernels (Helianthus annuus L.; SFK). Weanling female rats were fed diets containing 20% SFK in a 2x2x2 factorial design with marginal and adequate amounts of Ca, Zn, and Fe. Marginal Zn (11 mg/kg) and Fe (13 mg/kg), and Cd (0.18 mg/kg) were derived solely from 20% SFK. These amounts of Fe and Zn represented 39 and 90% of the NRC requirement for the rat, respectively. The marginal dietary Ca concentration (2.5 g/kg) was one-half the NRC requirement. After 5 weeks on the experiment, rats were fed 1 g of their respective diets containing SFK extrinsically labeled with 37 kBq 109Cd, and absorption was determined by whole-body counting techniques. Rats were then killed and organs collected for 109Cd assays. No effect of treatment on weight gain was observed; however, when dietary Zn was low, feeding marginal Ca elevated Cd absorption by 50% (P<0.05) over those fed adequate Ca and Zn. Feeding marginal Fe elevated Cd absorption >2.5-fold (P<0.001) over those fed adequate Fe. In contrast, the naturally occurring Zn in SFK that provided 90% of the rat's requirement was enough to deter excessive absorption of Cd and enough to alone prevent significant elevation of organ Cd. Organ content of 109Cd and Cd followed the same general pattern as whole-body absorption. These data show that marginal nutritional deficiencies of Ca and Fe can readily enhance the body burden of Cd that comes from the diet. Also, some natural competitors of Cd, such as Zn, contained in foods can independently minimize Cd absorption.     

Dose-dependent effects of dietary Pb and Zn on feeding and growth rates of the landsnail Helix engaddensis

Dose-dependent effects of dietary lead and zinc on feeding and growth rates of adult Helix engaddensis snails were studied over a 1-month period followed by a 2-week recovery period. Snails were fed on an artificial diet containing the following lead or zinc concentrations= 0, 20, 100, 500, 2500, and 12,500 microg/g dry food. At the end of the 6 weeks, mortality rate among snails fed on Pb-contaminated diet was 18.3% (11/60) and the respective value for Zn-contaminated diet was 50% (30/60). Both metals significantly reduced growth and feeding rates. Snails were found to be sensitive to zinc but tolerant to lead. During the recovery period, snails fed on Zn-contaminated diet failed completely to feed or grow normally, whereas snails fed on Pb-contaminated diet showed signs of slow improvement in terms of feeding and growth rates. The NOEC and LOEC for lead were 100 and 500 microg/g (-1), respectively, while the respective concentrations for zinc were 20 and 100 microg/g (-1).     

Genistein may prevent cadmium-induced bone loss in ovariectomized rats

This study compared the ability of genistein, a soy isoflavone, with that of 17 beta-estradiol to prevent bone loss in cadmium (Cd)-exposed ovariectomized (OVX) rats during growth. Female Wistar rats (4 weeks old) were either sham-operated (SH; n = 9/group) or OVX and placed on experimental diets (n = 9/group): OVX; OVX rats fed 50 ppm of CdCl(2) (OVX-Cd); OVX fed 50 ppm of CdCl(2) and 10 microg/kg of body weight genistein (OVX-Cd-G); and OVX fed 50 ppm of CdCl(2) and 10 microg/kg of body weight estrogen (OVX-Cd-E). All rats were given free access to AIN-76 modified diet and drinking water, with or without Cd, for 8 weeks. The OVX groups gained more body weight than the SH group. Femoral weight was increased by feeding genistein and estradiol, whereas femoral length among groups was not significantly different. Femoral Cd content was significantly higher in the OVX-Cd group than in the other groups. Both serum osteocalcin and calcium (Ca) concentrations, as well as urinary Ca, were significantly higher in the OVX-Cd group than in the other groups. Urinary excretion of Cd was significantly increased in Cd-OVX-G rats, and fecal Cd excretion was increased by feeding both genistein and estradiol. Femoral histomorpological changes in proliferative cartilage and hypertrophic cells in the OVX-Cd group showed that both cell types were decreased by feeding Cd, and irregular arrangements were observed in proliferative cells. However, both cells types exhibited normal distribution in OVX-Cd-G and OVX-Cd-E groups. These findings suggest that Cd/OVXinduced osteopenia or osteoporosis probably results from an increase in bone turnover. Genistein may be involved in stimulating Cd excretion and inhibiting Ca excretion from bone.     

The garden snail (Helix aspersa) as a bioindicator of organophosphorus exposure: effects of dimethoate on survival, growth, and acetylcholinesterase activity

The garden snail (Helix aspersa) is currently used as bioindicator of metallic pollution. Our objective was to extend its use to organic chemicals by studying the effects and tissue concentrations of the organophosphorus pesticide dimethoate following dietary uptake. After exposure for four weeks to increasing doses of pesticide in the diet, the median lethal concentration (LC50) was determined to be 3,700 microg/g food. Clinical signs indicated a no-observed-effect concentration of 100 microg/g and a lowest-observed-effect concentration of 250 microg/g. The growth parameters were decreased with increasing exposure to the pesticide. The median effective concentration (EC50), which was evaluated based on both shell diameter and dry weight inhibitions, was 665 and 424 microg/g, respectively, and the EC10 was 180 and 145 microg/g, respectively. Accumulation in the viscera was related to the amount of dimethoate in the food. The bioconcentration factors were low (>6 x 10(-3)). Acetylcholinesterase (AChE) activity was strongly decreased (80% from 250 microg/g). In conclusion, we demonstrated that the species H. aspersa could be a useful sentinel organism for organophosphorus contamination surveys. Among the effects measured, the inhibition of AChE activities and clinical signs were the most sensitive, followed by the growth parameters. These results confirm the suitability of the garden snail for development of sublethal toxicity tests using primary consumers and aboveground organisms.     

Pyrene biotransformation products as biomarkers of polycyclic aromatic hydrocarbon exposure in terrestrial Isopoda: concentration-response relationship,and field study in a contaminated forest

In this study, biotransformation products of pyrene were measured in the hepatopancreas of terrestrial isopods as biomarkers of polycyclic aromatic hydrocarbon (PAH) exposure. These products--pyrene-1-glucoside, pyrene-1-sulfate, an unknown pyrene conjugate, and 1-hydroxypyrene--were quantitated using high-performance liquid chromatography (HPLC) with fluorescence detection. In a controlled exposure experiment, a linear relationship was established between pyrene exposure and pyrene metabolite concentrations in the hepatopancreas of Porcellio scaber Latr. To this end, isopods of the species P. scaber were exposed to a range of pyrene concentrations spiked to their food. A linear response was found for all pyrene conjugates in the range 0.67 to 67 microg/g of pyrene (dry wt). Hepatopancreatic pyrene metabolite concentrations were also measured in isopods (P. scaber and Oniscus asellus L.) from PAH-contaminated field sites. The sites and the inhabiting isopods were located in a gradient of atmospheric PAH deposition caused by a nearby blast furnace plant. The highest levels of conjugated 1-hydroxypyrene in the hepatopancreas were 3.8 pmol/g fresh weight (pyrene-1-glucoside) and 2.8 pmol/g fresh weight (pyrene-1-sulfate) (expressed on whole-body basis). The levels of the pyrene metabolites correlate with reported pyrene concentrations in spite of these sites. As pyrene is one of the most predominant PAHs, analysis of its metabolites provides a good tool for environmental risk assessment of ecosystems with regard to PAH exposure, bioavailability, and biotransformation.     

Metal and metallothionein concentrations in Scoter (Melanitta spp.) from the Pacific northwest of Canada, 1989-1994

Eighty-six surf ( Melanitta perspicillata) and nine white winged (M. fusca) scoters were collected from 1989-1994 at 11 locations in British Columbia and the Yukon. Their kidney and liver tissues were analyzed for cadmium (Cd), copper (Cu), zinc (Zn), and metallothionein (MT) concentrations. Individual kidney Cd values ranged from 2.4 microg/g dry weight (DW) in birds from northern Vancouver Island to 390.2 microg/g (DW) in birds from the Queen Charlotte Islands, which is in the range of values potentially associated with kidney damage. Birds from the Queen Charlotte Islands also had high kidney Zn concentrations, which were significantly higher than values in birds from Naniamo and Westham Island on the south coast of BC. Cd values were significantly (p < 0.001) correlated with both Zn and MT concentrations in kidney tissue (r = 0.66 and 0.62, respectively). Male surf scoters had significantly higher kidney Cd and Zn levels than females, with mean Cd values of 47.3 and 19.7 microg/g DW (p < 0.002) for males and females, respectively. Mean Zn values were 120.8 and 108.0 microg/g DW (p < 0.05) for males and females, respectively. However, no sex differences were observed for either Cu or MT concentrations in kidney tissue. Individual MT values ranged from 2.7 to 416.8 microg/g wet weight (WW). Individual kidney Cu values ranged from 15.1 to 48.4 microg/g DW for both sexes. Kidney and liver Cd concentrations were significantly correlated (r = 0.90, p < 0.05) with kidney levels almost always higher than liver values. Although Cd accumulation by the Skidegate scoters was high, comparable tissue Cd values have been documented in other seabirds collected from the same general area, indicating elevated Cd concentrations in marine food chains around the Queen Charlotte Islands (Haida Gwaii) region of the Pacific coast.     

Dose-Dependent Effects of Cadmium on the Growth of Snails in Toxicity Bioassays

The effects on survival and growth of exposure to cadmium (Cd) in the food were analyzed in juvenile snails (age one month, mean weight 1 g) of the two subspecies Helix aspersa aspersa (H.a.a.) and H. aspersa maxima (H.a.m.). The experiments lasted for four weeks and the animals were fed with special snail food containing 0-, 50-, 100-, 200-, 400-, and 800 μg Cd/g of dry food. No significant adverse effect (NOEC) was noted at 50 μg/g for three weeks. A negative effect of Cd on growth was noted from 100 to 800 μg/g and plotting the growth coefficient variation against the Cd concentration led to an estimate of the EC₇₅ at day 14 of 370 μg/g in H.a.a. and 470 μg/g in H.a.m. and at day 28 of 290 μg/g in H.a.a. and 330 μg/g in H.a.m. Juvenile snails are thus a suitable material for use as bioindicators in the assay of contamination of food. Received: 4 May 1996/Revised: 24 December 1996     

Inhibition of acetylcholinesterase activity in the terrestrial isopod Porcellio scaber as a biomarker of organophosphorus compounds in food

This paper describes the toxicity of organophosphorus pesticide diazinon in juvenile and adult terrestrial isopods Porcellio scaber (Isopoda, Crustacea). The woodlice were exposed to different concentrations of diazinon added to food (5, 10, 50, and 100 or 150 micrograms/g dry food). Weight change and food assimilation efficiency were determined two and four weeks after the exposure. The activity of acetylcholinesterase (AChE) in surviving animals was measured at the end of the experiment. The results show that woodlice exposed to diazinon do not significantly differ from controls in growth and feeding rate. The reduction of AChE activity was observed at the lowest diazinon exposure (5 and 10 micrograms/g dry food). These results suggest that AChE activity might prove a useful biomarker, indicating low levels of organophosphates in food.