The effects of subchronic exposure to NeemAzal T/S on zebrafish (Danio rerio)

The objective of the study was to investigate the subchronic effects of the insecticidal preparation NeemAzal T/S at concentrations of 3, 10 and 30?mg/L on the mortality, growth and histopathology of juvenile zebrafish (Danio rerio), as well as its related effects on selected indices of oxidative stress during a 28-day toxicity test. The juvenile growth toxicity test was performed according to OECD Guidelines 215 (Fish, Juvenile Growth Test). The results of this study indicate that these tested concentrations of NeemAzal T/S (containing 1% of the active ingredient azadirachtin A) affect fish growth and have a negative influence on the indices of oxidative stress in the juvenile stage of zebrafish, as well as cause mild histopathological changes in liver tissue.     

Antioxidant enzymatic activities and lipid peroxidation in liver and ovary of zebrafish (Danio rerio) exposed to deltamethrin

Deltamethrin (DM) is being used as a substitute for organochlorines and organophosphates in pest control because of its low environmental persistence and toxicity. But it has become an environmental contaminant as it has been used widely. In this study, we investigated the effect of DM (technical grade) on the antioxidant system of adult zebrafish. For this, six-month-old fish were exposed to 2, 4 and 6?μg/L of DM for 96?h. The tissues selected were liver and ovary. Our data showed that exposure to DM increases CAT (catalase), SOD (superoxide dismutase), GPx (glutathione peroxidase, antioxidant enzymes), LPO (lipid peroxidation, non-enzymatic antioxidant) and GST (glutathione S-transferase, detoxifying enzyme) in liver and ovary. Increased GST could detoxify the toxicant; still there could be enough DM to cause oxidative stress. It appears from our study that zebrafish used compensatory mechanisms in eliminating reactive oxygen species. These data will be useful as oxidative stress is being used as a biomarker for aquatic pollution.     

Polyaspartic acid alleviates heavy metal toxicity in zebrafish (Danio rerio)

The present study was conducted to investigate the effect of polyaspartic acid (PASP) on Pb, Cu, and Cd toxicity in zebrafish (Danio rerio). After 24 hours’ toxicity test on zebrafish, LC50, survival rates of zebrafish and catalase (CAT) activity were determined. The results revealed that the 24-hour survival rates of zebrafish exposed to Pb (P?P?P?P?in vitro and in vivo mechanisms.     

五氯酚和八氯代二苯并二噁英复合暴露对斑马鱼胚胎发育的毒性效应

为了评价环境中五氯酚(PCP)和八氯代二苯并二噁英(OCDD)对水环境以及鱼类的影响,以斑马鱼为模式生物,研究了PCP和OCDD对其胚胎发育的单一及复合毒性效应.结果表明,PCP单独暴露(浓度25μg·L-1～5mg·L-1)对斑马鱼胚胎发育具有较强的毒性效应,可导致胚胎孵化率显著下降,死亡率、畸形率显著上升,而OCDD单独暴露(200、500μg·L-1)对斑马鱼胚胎发育没有明显的毒性效应;OCDD与环境浓度的PCP复合暴露(OCDD+PCP1:250μg·L-1+25μg·L-1;OCDD+PCP2:250μg·L-1+50μg·L-1)对斑马鱼胚胎的存活与发育等没有显著影响,对斑马鱼胚胎内CYP1A基因表达以及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)的酶活力也没有显著影响,在实验浓度下二者共存没有明显的复合毒性效应。     

Behavioural toxicity in zebrafish (Danio rerio) exposed to waterborne zinc and chromium(VI)

This study is to investigate the behavioural responses of zebrafish to the toxic effects of waterborne zinc and chromium(VI). Swimming performance of fish was tracked by a real-time toxicity bio-monitoring system and transformed into data to calculate individual and fish school behavioural endpoints. Behavioural responses under metals were in accordance with the Stepwise Behavioural Response model. Significant and concentration-dependent effects on swimming speed and turning times were found at concentrations equal to or higher than 3.5 mg L^-1 for zinc and 0.7 mg L^-1 for chromium. Swimming height significantly declined in concentrations equal to or higher than 3.5 mg L^-1 under zinc exposure, while for the chromium treatment, it significantly increased in lower concentration exposures and decreased in higher concentration exposures. Average distance and dispersion of fish school decreased at the beginning but there were no obvious changes during the whole exposure period. Behavioural responses under metal exposures were time-dependent: avoidance (increased speed and turning times and decreased average distance and dispersion) might be the first response and followed by height changes. Changes in behavioural endpoints ‘swimming speed and height’ can be used as early stress responses for acute aquatic metal contamination as part of ecological risk assessment.     

Impact of environmental concentrations of beta-cypermethrin on the antioxidant system in the brain and liver of zebrafish (Danio rerio)

Beta-cypermethrin (beta-CYP) is a widely used pyrethroid pesticide, the extensive application of which may potentially cause damage to non-target organisms. To investigate the effect of beta-CYP on the antioxidant system of aquatic animals, adult zebrafish were exposed to environmentally relevant dosages (0.01, 0.1 and 1.0 μg/L) of beta-CYP. The activities of four antioxidant enzymes in zebrafish liver and brain tissue were tested after 7, 15 and 30 days of exposure. Our results showed that exposure of beta-CYP could induce different levels of increase in hepatic superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR) and glutathione peroxidase (GPx) activities at 7 and 15 days post exposure (dpe), but caused apparent inhibition of hepatic SOD, GR and GPx activities at 30 dpe. Unlike in liver tissue, SOD and CAT activities in zebrafish brain did not show any apparent response to beta-CYP during the whole treatment period. In addition, increased brain GPx activities were observed at 7 and 30 dpe.     

Single and joint effects of HHCB and cadmium on zebrafish (Danio rerio) in feculent water containing bedloads

As an important type of emerging pollutants, ecological toxicity and risk of artificial musks are increasingly concerned. Thus, single and joint toxic effects of 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethylcyclopenta-gamma-2-benzopyran (HHCB) as one of the most widely applied artificial musks and cadmium (Cd) as an toxic metal on zebrafish (Danio rerio) were investigated by the exposure of zebrafish to various concentrations of HHCB or/and Cd in feculent water containing bedloads. The results indicated that the joint effect of HHCB and Cd changed during different exposure times within 120 h. The index of the antioxidant enzyme system including superoxide dismutase (SOD) and peroxidase (POD), and malondialdehyde (MDA) were sensitive and induced in the zebrafish stressed by Cd, and content of soluble protein (SP) was sensitive to HHCB and could be used as a biomarker for HHCB. Joint effects on antioxidant enzymes depended more on the effect of single Cd in the first one or two days. However, in the rest exposure days, the effect of HHCB began to dominate in the joint effect during the exposure process.     

三甲基氯化锡对斑马鱼(Danio rerio)生理生化特性的影响

为初步探讨三甲基氯化锡(trimethyltin chloride,TMT)对鱼类的毒性效应以及评价环境中TMT的安全性,采用静态鱼类急性毒性试验法测定了TMT对斑马鱼(Danio rerio)生理生化指标的影响;参考TMT的96 h-LC50值,设定3个浓度(0.39、0.78和1.17 mg·L-1)处理斑马鱼,测...     

The microcystins-induced DNA damage in the liver and the heart of zebrafish,Danio rerio

Microcystins (MCYST) are the freshwater cyanobacterial toxins, known to induce hepatocellular carcinoma, necrosis, intrahepatic bleeding, as well as human and livestock mortality. Within hepatocytes, MCYST selectively bind to protein phosphatases 1 and 2A, resulting in severe liver damage. The toxicology of MCYST in mice and rats has been well studied, but little is known regarding genotoxicity in aquatic animals. In this study, the zebrafish, Danio rerio was exposed to crude extract of Microcystis aeruginosa bloom. Liver and heart were examined for MCYST-induced toxicity. Light microscopy at 36?h revealed severe, widespread apoptotic necrosis of the majority of hepatocytes, and cytoskeletal deformation in myocardiocytes. Hepatocytes were dissociated with cell shrinkage and margination of nuclear chromatin. Laddering of genomic DNA from the liver and heart of the exposed fish in an increment of 180–200?bp was consistent with apoptosis. Fluorimetric analysis of DNA unwinding was carried out to determine the DNA strand breakage. After 36?h exposure, the % double-stranded DNA was significantly reduced in hepatocytes and myocardiocytes. In conclusion, the results obtained in this study indicate that, the extract of M. aeruginosa bloom is genotoxic to fish. The DNA damage observed in this study may be attributed to the activation of DNA endonucleases. This model of DNA damage may contribute for identifying novel molecular mechanisms of interest for therapeutic application.     

Metabolic responses in zebrafish (Danio rerio) exposed to zinc and cadmium by nuclear magnetic resonance -based metabolomics

Heavy metals are common marine and soil pollutants that are mainly the result of industrial activity, and are a threat to the environment and human health. In this study, ¹H nuclear magnetic resonance (NMR)-based metabolomics was applied to adult Danio rerio to monitor the metabolic change as a response to ZnCl₂ and CdCl₂ exposure at different concentrations for 72?h. NMR spectroscopy was used to identify and quantify the metabolites extracted from D. rerio. The metabolite profiles of the control and heavy metal exposed group were classified by partial least squares – discriminant analysis (PLS-DA) analysis, and potential contaminant-specific biomarkers were suggested. For the ZnCl₂-exposed zebrafish, the levels of ATP, aspartate and NAD⁺ were increased, whereas the levels of formate, inosine, hypoxanthine and succinate decreased. In addition, the CdCl₂-exposed zebrafish showed an increase in the levels of ATP and formate and a decrease in the levels of glutamate, inosine and glutathione. Overall, Zn and Cd may lead to neurotoxicity, disturbances in the energy metabolism and oxidative stress. Our finding demonstrated that the application of NMR-based metabolomics might be useful for detecting the toxicity caused by sub-lethal concentrations of heavy metal contaminants in the environment.     

脱氧鬼臼毒素对斑马鱼（Danio rerio）代谢酶和抗氧化酶活性的影响（Effects of Deoxypodophyllotoxin on Activities of Metabolic Enzymes and Antioxidant Enzymes of Danio rerio）

为初步探讨脱氧鬼臼毒素(Deoxypodophyllotoxin,DOP)对鱼类的毒性效应,采用静态鱼类急性毒性试验法测定了DOP对斑马鱼(Daniorerio)代谢酶和抗氧化酶活性的影响;参照DOP的96hLC50值,设定4个浓度(0.5、1.0、2.0、4.0mg·L-1)处理斑马鱼,48h后测定斑马鱼肌肉组织腺苷三磷酸酶(ATPase)、过氧化氢酶(CAT)和超氧化物歧化酶(SOD)活性.结果表明,DOP对斑马鱼96hLC50值为3.49mg·L-1,安全浓度为0.36mg·L-1.DOP在0.5~4.0mg·L-1对斑马鱼肌肉总ATPase、Na+-K+-ATPase和Ca2+-ATPase的作用趋势均表现为:1.0和2.0mg·L-1显著激活(p<0.05,p<0.01);4.0mg·L-1显著抑制(p<0.05),抑制率分别为24.4%、12.3%和33.8%.DOP在0.5~4.0mg·L-1对斑马鱼肌肉抗氧化酶的作用趋势与对ATPase的作用趋势一致,表现为:1.0、2.0mg·L-1DOP对SOD和CAT均表现为显著激活(p<0.05,p<0.01);4.0mg·L-1DOP对SOD和CAT均表现为显著抑制(p<0.05),抑制率分别为34.1%和31.9%.以上结果表明,DOP对斑马鱼为中等毒性;斑马鱼肌肉组织ATPase、CAT和SOD可能是脱氧鬼臼毒素的作用标靶之一.     

铜及其与四环素的联合暴露对斑马鱼胚胎的毒性效应

近年来工业和养殖业中铜和四环素的滥用,导致了一定程度的水环境污染问题。为探究铜与四环素对水生生物的毒害作用,选择斑马鱼作为受试生物,研究了铜及其与四环素的联合暴露对斑马鱼胚胎的毒性效应,并进一步探索了其中可能的致毒机制。结果表明:铜在低浓度下(10%致死浓度LC10=2.5μg·L~(-1),10%效应浓度EC_(10)=0.1μg·L~(-1))明显延迟了斑马鱼胚胎的孵化、卵黄囊吸收、头部、鱼鳔和体长等生长指标的发育,同时在心脏区域引起了明显的细胞凋亡效应。幼鱼体内总铜含量检测结果显示低浓度下铜的生物利用度相对更高。基因表达结果显示环境浓度的铜可能通过影响神经和心脏相关基因的表达引起斑马鱼胚胎的神经发育和心脏发育异常。铜和四环素的联合暴露实验结果表明二者的复合污染类型为拮抗作用,且两者相互作用可以形成络合物。综合以上结论,说明环境浓度的铜可能通过细胞凋亡、分子水平的变化等方式对水生生物的早期生长发育产生危害,如延迟生长发育、神经及心脏发育异常,另外铜可通过和四环素等环境中其他污染物的结合改变铜的生物有效性和毒性。     

全氟辛烷磺酸(PFOS)急性暴露对斑马鱼鳃显微结构的影响

采用光镜和电镜的方法,探讨了高浓度全氟辛烷磺酸(PFOS)急性暴露对斑马鱼(Danio rerio)鳃显微结构的影响.暴露组PFOS浓度分别为0.5、1、2mg·L-1,同时设对照组,连续暴露7d,取鳃制备石蜡切片,进行光镜和扫描电镜观察.结果表明,7d后PFOS暴露组斑马鱼鳃部均有不同程度的损伤,石蜡切片上可观察到暴露组鱼鳃上皮细胞坏死脱落,鳃小片断裂或融合,并伴有鳃丝充血,且病变程度随PFOS浓度的升高而加重;扫描电镜下可观察到暴露组鱼鳃鳃丝表面分泌物增多,鳃丝细胞脱落或水肿,鳃小片前端细胞破损.高浓度PFOS急性暴露可在短期内对斑马鱼鳃组织造成严重损伤,且损伤程度存在剂量依赖效应.     

五氯酚和八氯代二苯并二嗯英复合暴露对斑马鱼胚胎发育的毒性效应

为了评价环境中五氯酚(PCP)和八氯代二苯并二嚼英(OCDD)对水环境以及鱼类的影响,以斑马鱼为模式生物,研究了PCP和OCDD对其胚胎发育的单一及复合毒性效应.结果表明,PCP单独暴露(浓度25μg·L-1～5mg·L-1)对斑马鱼胚胎发育具有较强的毒性效应,可导致胚胎孵化率显著下降,死亡率、畸形率显著上升,而OCDD单独暴露(200、500μg·L-1)对斑马鱼胚胎发育没有明显的毒性效应;OCDD与环境浓度的PCP复合暴露(OCDD PCP1:250μg·L-1 25μg·L-1;OCDD PCP2:250μg·L-1 50μg·L-1)对斑马鱼胚胎的存活与发育等没有显著影响,对斑马鱼胚胎内CYP1A基因表达以及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)的酶活力也没有显著影响,在实验浓度下二者共存没有明显的复合毒性效应.     

Evaluation of developmental toxicity and apoptotic induction of the aqueous extract of Millettia pachycarpa using zebrafish as model organism

Extensive and indiscriminate use of synthetic compounds and natural compounds obtained from plant sources have resulted in serious threats to the aquatic ecosystem and human health. Aqueous extract of the root of the plant, Milletia pachycarpa Benth, is currently used for killing fish in the state of Manipur, India. Moreover, this plant is also used as traditional medicine in this region. Although it is widely used in traditional medicine, there is limited information available regarding the adverse effects and mechanism underlying its toxicity. This study examined the effects of exposure to aqueous extract of M. pachycarpa (AEMP) on early embryonic development of zebrafish embryos and mechanisms underlying toxicity. Zebrafish embryos treated with different concentrations of the AEMP produced embryonic lethality and developmental defects. The 96-hr-LC₅₀ of AEMP was found to be 4.276 µg/mL. Further, multiple developmental abnormalities such as pericardial edema, yolk sac edema, spinal curvature, swim bladder deflation, decreased heart rate, and delayed hatching were also observed in a dose-dependent manner. Zebrafish embryo showing moderate-to-severe developmental defects following AEMP exposure cannot swim properly. Further, this study examined oxidative stress and apoptosis in embryos exposed to AEMP. Enhanced production of ROS and apoptosis was found in brain, trunk, and tail of zebrafish embryos treated with AEMP. Data suggest that oxidative stress and apoptosis are associated with AEMP-induced embryonic lethality and developmental toxicity in zebrafish embryos.     

Toxicity of depleted uranium on isolated liver mitochondria: a revised mechanistic vision for justification of clinical complication of depleted uranium (DU) on liver

Depleted uranium (DU) is widely used in military anti-armor weapons. Recent evidence suggested that oxidative stress and mitochondrial dysfunction may contribute to DU-induced toxicity. However, the underlying mechanisms of DU toxicity in mitochondria are not well understood. In this study, liver mitochondria were obtained from Wistar rats treated with DU in the form of uranyl acetate (UA) (0.5, 1 or 2 mg/kg i.p.) using differential centrifugation. For in vitro experiments, control rat liver mitochondria were incubated with different concentrations of UA (50, 100 or 200 μM) for 1 hr. Mitochondrial reactive oxygen species (ROS) production, collapse of mitochondrial membrane potential, and mitochondrial swelling were examined by flow cytometry. Mitochondrial sources of ROS formation were determined using specific substrates and inhibitors. Extent of lipid peroxidation (LPO) and glutathione (GSH) oxidation, and also complex II and IV activities were detected via spectroscopy. Further, the concentration of ATP and ATP/ADP ratio was measured using luciferase enzyme and release of cytochrome c from mitochondria which was detected by ELISA kit. UA induced succinate-supported mitochondrial ROS production, elevated LPO levels, GSH oxidation, and mitochondrial complex II inhibition. UA also induced mitochondrial permeability transition and increase in cytochrome c release which subsequently disturbed oxidative phosphorylation and reduced the mitochondrial ATP concentration. Data suggest that mitochondrial oxidative stress and uncoupling of oxidative phosphorylation may play key roles in DU-induced hepatic toxicity.     

壬基酚和双酚A对雄性斑马鱼(Danio rerio)卵黄蛋白原mRNA的诱导效应

为了对野外环境中内分泌干扰物更快速有效地进行检测,运用逆转录-聚合酶链反应(RT-PCR)技术检测了17β-雌二醇(E2)、壬基酚(NP)和双酚A(BPA)对雄性成体斑马鱼肝脏合成卵黄蛋白原(VTG)mRNA的诱导作用,并初步探讨了VTGmRNA体内代谢动力学模式.结果表明,随着暴露浓度的升高(E2:20、100、250、500ng·L-1;NP:20、100、250、500μg·L-1;BPA:50、100、250、500、1000μg·L-1),VTGmRNA表达量均显著增加,VTGmRNA表达量与暴露浓度具有明显的剂量-效应关系.在10d的暴露期内,随着暴露时间的延长,VTGmRNA表达量均显著增加,VTGmRNA表达量与暴露时间呈明显的时间-效应关系;停止暴露后,VTGmRNA表达量迅速下降,E2诱导的表达消失得最快(4d),而NP、BPA诱导的表达持续时间相对较长(6d).     

利用大型蚤和斑马鱼评价腈纶废水好氧-厌氧处理过程的急性毒性和遗传毒性变化

腈纶废水毒性较强且难以去除,对腈纶废水毒性及污水处理过程毒性削减能力进行评价,可为开发该类废水毒性减排技术提供科学依据。利用大型蚤及斑马鱼评价某腈纶废水的急性毒性和遗传毒性,及厌氧-好氧工艺对该废水毒性的削减能力。采用大型蚤活动抑制率和斑马鱼致死率表征废水急性毒性,采用斑马鱼肝细胞彗星尾矩表征废水遗传毒性。腈纶废水处理前对大型蚤和斑马鱼的急性毒性单位(TU)分别为1.2和2.9,经厌氧-好氧工艺处理后分别降至0.4和0.5。遗传毒性结果表明腈纶废水对斑马鱼肝细胞造成DNA损伤作用,经处理后遗传毒性仍显著高于阴性对照组。理化指标与毒性指标相关性分析表明,该废水氨氮与毒性显著相关,推测氨氮可能是该废水中的重要致毒因子之一。研究结果表明该腈纶废水采用现有厌氧-好氧工艺无法有效削减毒性,对受纳水体水生态环境造成潜在危害。     

林丹短期暴露下的斑马鱼(Brachydanio rerio)组织学变化

以斑马鱼(Brachydaniorerio)为受试生物,初步探讨了林丹对斑马鱼的急性毒性以及不同浓度(0.01μg·L-1、1.0μg·L-1、100.0μg·L-1)的林丹暴露36d后对斑马鱼鳃和肝组织结构变化的影响.结果表明,林丹对斑马鱼96h-LC50为97.98μg·L-1,95%可信限为94.38￣101.72μg·L-1.当林丹浓度为0.01μg·L-1时,斑马鱼的鳃和肝组织结构未发生明显变化,而暴露在1.0μg·L-1和100.0μg·L-1林丹溶液中的斑马鱼的鳃和肝组织结构变化显著.斑马鱼鳃组织变化为:上皮细胞残损、脱落,鳃小片上皮细胞水肿,柱细胞变形.斑马鱼肝组织变化为:部分肝细胞肿大,细胞核萎缩变形或偏离细胞中心,胞质疏松,空泡明显增加,细胞质中可见脂沉积.与1.0μg·L-1林丹暴露相比,暴露在100μg·L-1林丹溶液中的斑马鱼鳃和肝组织结构受到的损害更为严重.     

对硝基酚对大型蚤和斑马鱼胚胎的毒性

采用静态生物毒性试验方法,研究了对硝基酚对大型蚤的急性毒性和斑马鱼胚胎发育影响.结果表明,大型蚤幼蚤接触不同浓度的对硝基酚后,活动会受到不同程度的抑制,甚至死亡,48hEC50值为2.55mg·L-1.对硝基酚对斑马鱼胚胎有蓄积致毒作用,最敏感性指标为83h未孵化、83h卵凝结;实验还发现对硝基酚具有低浓度促进斑马鱼胚胎发育和孵化,高浓度抑制发育并延缓孵化的作用.对硝基酚对大型蚤的毒性与对斑马鱼胚胎的发育影响相比,大型蚤对对硝基酚反应更为敏感.