Effects of water chemistry variables on gill binding and acute toxicity of cadmium in rainbow trout (Oncorhynchus mykiss): A biotic ligand model (BLM) approach

This study investigated the short-term (3 h) cadmium binding characteristics of the gills, as well as the influence of various water chemistry variables [calcium, magnesium, sodium, pH, alkalinity and dissolved organic carbon (DOC)] on short-term gill accumulation and acute toxicity of cadmium in juvenile freshwater rainbow trout. The cadmium binding pattern revealed two types of cadmium binding sites in the gill: (i) saturable high affinity sites operating at a low range of waterborne cadmium concentration, and (ii) non-saturable low affinity sites operating at a higher range of cadmium concentration. Among the water chemistry variables tested, only calcium and DOC significantly reduced both gill accumulation and toxicity of cadmium. Interestingly, alkalinity (15-90 mg L(-1) as CaCO(3)) did not influence the gill cadmium accumulation but a significant increase in toxicity was recorded at a higher alkalinity level (90 mg L(-1)). Affinity constants (log K) for binding of competing cations (Cd(2+) and Ca(2+)) to the biotic ligand and for binding of Cd(2+) to DOC were derived separately from the 3 h gill binding tests and the 96 h toxicity tests. In general, the values agreed well, indicating that both tests targeted the same population of high affinity binding sites, which are likely Ca(2+) uptake sites on the gills. These parameters were then incorporated into a geochemical speciation model (MINEQL+) to develop a biotic ligand model for predicting acute toxicity of cadmium in trout. The model predictions exhibited a good fit with the measured toxicity data except for high alkalinity and pH.     

Modulation of glutathione-related antioxidant defense system of fish chronically treated by the fungicide propiconazole

Recently, residual fungicides are generally recognized as relevant sources of aquatic environmental pollutants. However, the toxicological effects of these contaminants have not been adequately researched. In this study, the chronic effect of PCZ, a triazole-containing fungicide commonly present in aquatic environment, on GSH-related antioxidant system and oxidative stress indices of rainbow trout (Oncorhynchus mykiss) were investigated. Fish were exposed at sub-lethal concentrations of PCZ (0.2, 50 and 500 μg/L) for 7, 20 and 30 days. GSH levels and GSH-related enzyme activities, including GPx, GR and GST, were quantified in three tissues—liver, gill and muscle. The levels of LPO and CP were also measured as makers of oxidative damage. In addition, the correlations of the measured parameters in various tissues were evaluated by using PCA. The results of this study indicate that chronic exposure of PCZ has resulted in different responses in various tissues and the gill was the most sensitive tissue; however, before these parameters are used as potential biomarkers for monitoring residual fungicides in aquatic environment, more detailed experiments in laboratory need to be performed in the future.     

Immunohistochemical localization of rainbow trout ladderlectin and intelectin in healthy and infected rainbow trout (Oncorhynchus mykiss)

In the present study, the pattern of immuno-reactive ladderlectin and intelectin in healthy rainbow trout is compared to rainbow trout infected with a variety of infectious agents. In healthy rainbow trout, both proteins were localized to individual epithelial cells of the gill and intestine and both proteins were clearly demonstrated within cytoplasmic granules of polymorphonuclear leucocytes and macrophages/monocytes found in blood vessels, hepatic sinusoids, renal interstitium, mucosal epithelium and submucosa of normal intestine. In tissue from infected rainbow trout, there was an overall relative increase in both lectins compared to healthy fish and both proteins were detected in extra-cellular spaces surrounding bacteria, fungi and protozoa. Increased distribution and density of both RTLL and RTInt was demonstrated along mucosal surfaces and within inflammatory leucocytes in infected tissues and immune related organs. These findings represent one of the few examples of in vivo association of defence lectins and infectious agents.     

Radiotracer Studies on Waterborne Copper Uptake,Distribution, and Toxicity in Rainbow Trout and Yellow Perch: A Comparative Analysis

Rainbow trout (Oncorhynchus mykiss) are often used to estimate important biotic ligand model (BLM) parameters, such as metal-binding affinity (log K) and capacity (B_max). However, rainbow trout do not typically occupy metal-contaminated environments, whereas yellow perch (Perca flavescens) are ubiquitous throughout most of North America. This study demonstrates that dynamic processes that regulate Cu uptake at the gill differ between rainbow trout and yellow perch. Rainbow trout were more sensitive to acute aqueous Cu than yellow perch, and toxicity was exacerbated in soft water relative to similar exposures in hard water. Whole body Na loss rate could account for acute Cu toxicity in both species, as opposed to new Cu uptake rate that was not as predictive. Time course experiments using radiolabelled Cu (⁶⁴Cu) revealed that branchial Cu uptake was rather variable within the first 12 h of exposure, and appeared to be a function of Cu concentration, water hardness, and fish species. After 12 h, new branchial Cu concentrations stabilized in both species, suggesting that metal exposures used to estimate BLM parameters should be increased in duration from 3 h to 12+ h. In rainbow trout, 71% of the new Cu bound to the gill was exchangeable (i.e., able to either enter the fish or be released back to the water), as opposed to only 48% in yellow perch. This suggests that at equal exposure concentrations, proportionally more branchial Cu can be taken up by rainbow trout than yellow perch, which can then go on to confer toxicity. These qualitative differences in branchial Cu handling between the two species emphasize the need to develop BLM parameters for each species of interest, rather than the current practice of extrapolating BLM results derived from rainbow trout (or other laboratory-reared species) to other species. Data reported here indicate that a one-size-fits-all approach to predictive modeling, mostly based on rainbow trout studies, may not suffice for making predictions about metal toxicity to yellow perch—that is, a species that inhabits metal-contaminated lakes around northern Canadian industrial operations.     

Occurrence and synthesis of a non-thionein, zinc-binding protein in the rainbow trout (Salmo gairdneri)

A non-thionein, Zn-binding protein (ZBP) was induced in Donaldson strain rainbow trout (Salmo gairdneri) by 7 mg/kg, i.p. injections of divalent Zn ion. The Sephacryl S-200 used for supernatant fractionation had to be saturated with Zn to recover quantitatively the Zn-ZBP complex. The ZBP was present in liver and kidney, but was absent from gill and spleen. The apparent molecular weights of the liver and kidney ZBP as estimated by gel filtration were 17,300 +/- 1300 (SD; N = 11) and 18,100 (N = 1), respectively. Starvation induced hepatic ZBP synthesis whereas cycloheximide inhibited hepatic ZBP synthesis. The quantity of hepatic ZBP synthesized varied with the temperature of the water in which the trout resided. The maximum quantity of ZBP in the liver following a single 7 mg/kg Zn injection (17 micrograms Zn/g liver wet weight) occurred at 24 hr.     

Hexavalent Chromium Reduction in Soils Contaminated with Chromated Copper Arsenate Preservative

The toxicity and mobility of chromium in the environment greatly depends upon its speciation. The reduction of hexavalent chromium to trivalent chromium in a soil environment was examined by spiking three soil types (sandy, clayey, and organic soils) with a common wood preservative solution known as chromated copper arsenate (CCA). Chromium in the CCA preservative solution exists in the hexavalent form. The total and hexavalent chromium concentrations (mg/kg) were measured over a period of 11 months. Leachable chromium concentrations (mg/L) were assessed using the synthetic precipitation leaching procedure (SPLP). The degree and rate of hexavalent chromium reduction were similar for the sand and clayey soil, but much greater for the organic soil. Most of the chromium reduction occurred within the first month of the experiment. At the end of the experiment, approximately 50% of the hexavalent chromium was converted to the trivalent form in the sand and clayey soils. Hexavalent chromium concentrations were below detection in the organic soil at the end of the experiment. Nearly all of the chromium observed in the SPLP leachates was in the form of hexavalent chromium. Chromium leaching was thus greatest in the sand and clay soils where the hexavalent chromium persisted. The results indicate that hexavalent chromium in soils can persist for considerable time periods, in particular in soils with low organic matter content.

     

An alternative model to study the association of rainbow trout (Oncorhynchus mykiss L.) pathogens with the gill tissue

This study describes the development of a trout gill perfusion model, consisting of an excised branchial arch from rainbow trout (Oncorhynchus mykiss L.), perfused via the afferent branchial artery and suspended in a circular organ chamber filled with Ringer solution. Different perfusion fluids were tested: Ringer, Cortland, Ringer + procaine, Ringer + adrenalin, Cortland + procaine, Cortland + adrenalin and Cortland + dextran 1%. The latter perfusion fluid proved to be satisfactory, maintaining the gill tissue in a healthy condition outside the body of the fish for at least 180 min. Using this model, the interaction of damaging agents with the trout gill tissue may be studied under carefully controlled conditions. The trout gill perfusion model leads effectively to a reduction in the number of experimental animals to be used and also involves an elimination of pain and/or suffering, which is as good as complete.     

Interaction between dietary calcium supplementation and chronic waterborne zinc exposure in juvenile rainbow trout, Oncorhynchus mykiss

This study investigated the effects of dietary Ca2+ on branchial Ca2+ and Zn2+ uptake, new and total zinc accumulation in target tissues (gill, liver and kidney), calcium and zinc homeostasis, and acute tolerance to waterborne zinc in fish chronically exposed to waterborne zinc. Juvenile rainbow trout (Oncorhynchus mykiss) were maintained on a calcium-enriched diet [41.2 mg vs. 21.2 mg (control) calcium/g dry wt. of food] and chronic waterborne zinc exposure (2.3 micromol/L), both separately and in combination, for 28 days. Calcium-supplemented diet in the absence of waterborne zinc significantly reduced branchial Ca2+ and Zn2+ influx rates, and new and total zinc accumulations in target tissues relative to control. However it did not protect against the acute zinc challenge. In contrast, waterborne zinc exposure significantly increased branchial Ca2+ and Zn2+ influx rates, new and total zinc concentrations in target tissues, and acute zinc tolerance relative to control. Interestingly, no such changes in any of these parameters were recorded in fish treated simultaneously with elevated dietary Ca2+ and waterborne zinc, except acute zinc tolerance which was highest among all the treatments. Thus, we conclude that the interactions between elevated dietary Ca2+ and waterborne zinc can protect freshwater fish against waterborne zinc toxicity.     

The effect of mycotoxin deoxynivalenol (DON) on the oxidative stress markers in rainbow trout (Oncorhynchus mykiss,Walbaum 1792)

Deoxynivalenol (DON) is one of the most frequently detected mycotoxins in agricultural commodities used as animal feedstuff in Central Europe. This study focuses on determining effects of diets containing DON on oxidative stress markers and detoxifying enzymes in rainbow trout (Oncorhynchus mykiss). The fish were fed with commercial pellets containing DON at a concentration of 2 mg kg^?1. Selected enzymes were measured in liver, gill and caudal kidney of the fish after 23 and 32 days of the experiment. Significant differences between the control and experimental groups were observed concerning activities of glutathione peroxidase (GPx) in kidney, glutathione reductase (GR) in gill and kidney, catalase (CAT) in kidney and liver and glutathione S‐transferase (GST) in gill and liver. No significant differences were found for superoxide dismutase (SOD) gene expression, lipid peroxidation (TBARS) and the ferric reducing ability of plasma (FRAP). The data show that DON in the diet at the concentration below EC recommendation (2006/576/EC) induces oxidative stress in the rainbow trout.     

A comparison of cadmium-induced metallothionein gene expression and Me2+ distribution in the tissues of cadmiumsensitive (rainbow trout; Salmo gairdneri) and tolerant (stone loach; Noemacheilus barbatulus) species of freshwater fish

1. The accumulation of cadmium in the liver, kidney and gills of rainbow trout and stone loach was measured during exposure of the fish to the metal at 3 smg/l in their aquarium water. The pattern of accumulation of the toxic metal in the individual organs was different between the two species.2. The tissue concentrations of metallothionein-specific mRNA and metallothionein protein were also determined in these organs from the same fish. In rainbow trout, the induction of metallothionein gene expression resulted in a gradual increase in metallothionein concentration in gill over the course of the experiment whereas increases in metallothionein in the liver and kidney were detected only at the later time points of analysis (beyond 19 weeks). By contrast, in the same tissues from stone loach, relatively minor changes were quantified in specific mRNA and metallothionein concentrations.3. Throughout the experimental period, tissue concentrations of zinc and copper were determined in the liver, kidney and gills of the rainbow trout and stone loach. Subtle decreases were observed in the zinc concentration of gills in rainbow trout and substantial increases were observed in the hepatic copper concentrations in both species at the later time points of analysis.4. The ability of cadmium to induce metallothionein gene expression and its subsequent ability to compete for the sequestration sites on the newly-synthesized protein is discussed with regard to the relative levels of cadmium, zinc and copper in the organs studied and differing regimes of cadmium administration.     

六价铬对薏米人工湿地微生物群落数量的影响

通过桶栽构筑微型模拟垂直流薏米人工湿地(CAW),以1/2 Hoaglands营养液为营养源,在营养液中添加不同浓度的Cr6+(0,5,20,40,60mg/L,以K2Cr2O7配置),各浓度处理均以不种植薏米湿地(NPW)为对照,以研究铬(Cr6+)对薏米人工湿地基质真菌、细菌及放线菌群落结构数量的影响。结果表明:(1)真菌、细菌、放线菌的数量在薏米人工湿地(CAW)中明显多于无植物对照处理(NPW);(2)中低浓度(5、20mg/L)Cr6+对CAW真菌,对NPW细菌、放线菌数量有促进作用,薏米人工湿地微生物对低中浓度的Cr6+胁迫有一定的耐受能力。     

Evaluating the toxicity of environmental concentrations of waterborne chromium (VI) to a model teleost, Oncorhynchus mykiss: a comparative study of in vivo and in vitro

Toxic effects of environmental concentrations (50, 100, and 200μg/L) of waterborne chromium (VI) were evaluated in rainbow trout by comparison of in vitro and in vivo assays. Multiple biomarkers were measured including oxidative stress indices and antioxidant response parameters in liver and brain, as well as Na(+)-K(+)-ATPase in gill. Superoxide dismutase (SOD) and glutathione reductase (GR) activities were significantly induced (1.54-fold and 1.37-fold, respectively) in fish brain in vivo, but no significant differences were observed in any other biomarker or in vivo test group. Oxidative stress was apparent in vitro as significantly higher levels of oxidative indices, with the highest induction of TBARS and CP found in brain at 200μg/L Cr(VI) (2.41-fold and 1.95-fold, respectively), and SOD and GR activities and reduced glutathione in brain were significantly inhibited (65%, 44%, and 36%, respectively). In vitro Na(+)-K(+)-ATPase activity in gill was also significantly inhibited at concentrations of 100 and 200μg/L (69% and 45%, respectively). Short-term exposure to environmental concentrations of Cr(VI) does not therefore evoke marked effects in fish in vivo. Based on the present results, a set of in vitro tests with tissue homogenate can be evoked more remarkable effects by the lower concentrations of Cr(VI) than in vivo, which could provide some useful information and might be a potential alternative approach for monitoring heavy metal pollution in aquatic environments. However, it needs more detailed studies in other area, such as hormonal response or genotoxicity, before these findings could be applied in the field investigation.     

Intra- and extra-cellular sources of T3 binding to putative thyroid hormone receptors in liver, kidney, and gill nuclei of immature rainbow trout, Oncorhynchus mykiss.

The sources of extracellular and intracellular 3,5,3'-triiodo-L-thyronine (T3) binding to putative thyroid hormone receptors in liver, kidney, and gill nuclei were determined in vivo for immature rainbow trout at 12 degrees C. Both [131I]T3 and [125I]T4 were injected intraperitoneally, the plasma and tissues were examined at isotopic equilibrium at 20 h, and the proportions of intracellular [125I]T3 and extracellular [131I]T3 saturably bound in the nucleus were determined. Comparable total amounts of T3 were saturably bound in the nuclei of liver (7.2), kidney (8.0), and gill (9.7 moles x 10(-13) .mg DNA-1), but the percentage of nuclear T3 generated within the target cell was greater for gill (76%) than for liver (50%) and kidney (28%). Both gill and liver possess a low Km T4 5'monodeiodinase which could be responsible for the high proportion of the nuclear T3 generated within those tissues.     

Removal of hexavalent chromium using distillery sludge

Batch mode experiments were conducted to study the removal of hexavalent chromium from aqueous and industrial effluent using distillery sludge. Effects of pH, contact time, initial concentration and adsorbent dosage on the adsorption of Cr(VI) were studied. The data obeyed Langmuir and Freundlich adsorption isotherms. The Langmuir adsorption capacity was found to be 5.7 mg/g. Freundlich constants K(f) and n were 2.05 [mg/g(L/mg)(n)] and 3.9, respectively. Desorption studies indicated the removal of 82% of the hexavalent chromium. The efficiency of adsorbent towards the removal of chromium was also tested using chromium-plating wastewater.     

Peripheral serotonin dynamics in the rainbow trout (Oncorhynchus mykiss)

Serotonin (5-hydroxytryptamine, 5-HT) occurs in a wide range of tissues throughout the body of the rainbow trout. Results reported here indicate that the main peripheral sources of serotonin are the intestinal tract and the gill epithelium (levels above 1500 ng/g). The high intestinal serotonin concentration is mostly due to serotoninergic nerve fibres, which are present at high density in the intestinal wall. Only about 2% of serotonin is associated with mucosal enterochromaffin cells. In the remaining tissues studied serotonin concentration was below 160 ng/g: the highest concentrations were seen in the anterior and posterior kidneys, followed by the liver, heart, and spleen. 5-Hydroxyindolacetic acid (5-HIAA) levels, except in plasma, were generally lower than serotonin levels, and were below our detection limits in heart, spleen and posterior kidney. Acute d-fenfluramine treatment (5 or 15 mg/kg i.p.) significantly increased 5-HIAA/5-HT ratio in the anterior intestine, pyloric caeca and plasma. Serotonin released from intestinal serotoninergic fibres in response to d-fenfluramine treatment is metabolized locally, and only a small part reaches the blood, from where it can be taken up and metabolized by other peripheral tissues, such as the liver and gill epithelium. The non-metabolized serotonin pool in the blood appears to be located extracellularly, not intracellularly as in mammals. In view of these findings, we present an overview of peripheral serotonin dynamics in rainbow trout.     

Extracellular carbonic anhydrase in the dogfish, Squalus acanthias: a role in CO2 excretion.

In Pacific spiny dogfish (Squalus acanthias), plasma CO(2) reactions have access to plasma carbonic anhydrase (CA) and gill membrane-associated CA. The objectives of this study were to characterise the gill membrane-bound CA and investigate whether extracellular CA contributes significantly to CO(2) excretion in dogfish. A subcellular fraction containing membrane-associated CA activity was isolated from dogfish gills and incubated with phosphatidylinositol-specific phospholipase C. This treatment caused significant release of CA activity from its membrane association, a result consistent with identification of the dogfish gill membrane-bound CA as a type IV isozyme. Inhibition constants (K(i)) against acetazolamide and benzolamide were 4.2 and 3.5 nmol L(-1), respectively. Use of a low dose (1.3 mg kg(-1) or 13 micromol L(-1)) of benzolamide to selectively inhibit extracellular CA in vivo caused a significant 30%-60% reduction in the arterial-venous total CO(2) concentration difference, a significant increase in Pco(2) and an acidosis, without affecting blood flow or ventilation. No effect of benzolamide on any measure of CO(2) excretion was detected in rainbow trout (Oncorhynchus mykiss). These results indicate that extracellular CA contributes substantially to CO(2) excretion in the dogfish, an elasmobranch, and confirm that CA is not available to plasma CO(2) reactions in rainbow trout, a teleost.     

Apoptotic effects and glucose-6-phosphate dehydrogenase responses in liver and gill tissues of rainbow trout treated with chlorpyrifos

We investigated apoptotic effects and changes in glucose-6-phosphate dehydrogenase (G6PD) enzyme activity in liver and gill tissues of fish exposed to chlorpyrifos. Three different chlorpyrifos doses (2.25, 4.5 and 6.75 μg/L) were administrated to rainbow trout at different time intervals (24, 48, 72 and 96 h). Acute exposure to chlorpyrifos showed time dependent decrease in G6PD enzyme activity at all concentrations (p < 0.05). Immunohistochemical results showed that chlorpyrifos caused mucous cell loss in gill tissue and apoptosis via caspase-3 activation in fish. The present study suggested that chlorpyrifos inhibits G6PD enzyme and causes mucous cell loss in gill and apoptosis in gill and liver tissues.     

虹鳟spindlin基因克隆及不同倍性的表达分析

spindlin基因是减数分裂纺锤体相关因子,为了研究spindlin基因在二倍体和三倍体雌性虹鳟减数分裂过程中出现的差异,通过cDNA末端快速扩增(RACE)技术获得spindlin基因cDNA 4529 bp(GenBank登录号:MN378564),其中3′非编码区(UTR)和5′非编码区(UTR)分别长3662 bp和141 bp,开放阅读框(ORF)长726 bp,编码241个氨基酸,该蛋白质序列的相对分子量为28.3 kD,理论等电点值为5.94,无跨膜结构。同源性分析表明,虹鳟(Oncorhynchus mykiss)与银大马哈鱼(Oncorhynchus kisutch)同源最高,高达99.59%。系统发育进化树显示,虹鳟与大鳞大马哈鱼(Oncorhynchus tshawytscha)和红点鲑(Salvelinus alpinus),聚为一支。实时荧光定量(RT-PCR)结果显示, spindlin基因在二倍体雌性虹鳟卵巢、肾、肝、脾、肌、鳃、心、眼、肠和鳍组织中均有表达,其中,在卵巢中的表达量极显著高于其他组织(P<0.01)。对于二倍体雌性虹鳟,在受精后24...