A preliminary investigation into the use of biomarkers for the monitoring of an ocean outfall

The use of several biomarkers of exposure for monitoring a treated mixed effluent from the Latrobe Valley Ocean Outfall was examined. Sand flatheads (Platycephalus bassensis) were exposed in the laboratory to two dilutions of effluent (1.3 and 2.5%, v/v) for three days. Cytochrome P450 content, 7-ethoxyresorufin-O-deethylase (EROD) activity, and biliary metabolites of polycyclic aromatic hydrocarbons (PAH), 2-chlorosyringaldehyde and 2,6-dichlorosyringaldehyde, were measured. There were no statistically significant differences in cytochrome P450 content or EROD activity between control and exposed fish. Analysis of the bile by gas chromatography/mass spectrometry of fish exposed to 2.5% effluent found no increases above background in the levels of PAHs, or specific chlorinated phenolics that are found in bleached hardwood pulp effluents. © 1996 by John Wiley & Sons, Inc.     

Exposure to pollutants present in Iguaçu River Southern Brazil affect the health of Oreochromis niloticus (Linnaeus, 1758): Assessment histological,genotoxic and biochemical

Urban sewage is a source of major contamination in aquatic systems and contributes to environmental and human health disturbances. This study investigates the effects of sewage-polluted waters from Iguaçu River on the health of juvenile Oreochromis niloticus. Two hundred four specimens were exposed to riverine water in four groups: no diluted, 25 and 50 % diluted water and a control group without tested water for 72 days. Biological samples were obtained for histopathological, neurotoxicity, antioxidant defenses, genotoxicity, metallothionines expression and polycyclic aromatic hydrocarbons (PAHs) metabolites. The results showed histopathological alterations in liver and gills, genotoxic alteration in erythrocytes, reduction of acetylcholinesterase activity in brain and muscle, activation of antioxidant defenses in the liver, recruitment of metals by metallothionein and the detection of PAHs metabolites in bile. These results demonstrate that juveniles of O. niloticus are susceptible to Iguaçu River exposure water and they can be used as indicator of water quality.     

Subchronic exposure to diflubenzuron causes health disorders in neotropical freshwater fish,Prochilodus lineatus

The action of diflubenzuron (DFB) was evaluated in a freshwater fish, Prochilodus lineatus, after exposure to 0.06, 0.12, 0.25, or 0.50 mg L^?1 DFB for 14 days. Erythrocyte nuclear abnormalities (ENA), the gill activity of Na⁺/K⁺‐ATPase, H⁺‐ATPase and carbonic anhydrase (CA), and lipid peroxidation (LPO) and histopathological changes in the gills and liver were determined. The number of micronuclei increased in fish exposed to 0.25 and 0.50 mg L^?1 DFB. Plasma Cl^? and the CA activity decreased, while the activity of Na⁺/K⁺‐ATPase and of H⁺‐ATPase increased in fish exposed to 0.25 and 0.50 mg L^?1 DFB. LPO did not change in the gills but increased in the liver of fish exposed to 0.25 and 0.50 mg L^?1 DFB. In the gills, histopathological changes indicated disperse lesions and slight to moderate damage in fish exposed to 0.50 mg L^?1 DFB, whereas in the liver, these changes were significantly greater in fish exposed to 0.25 and 0.50 mg L^?1 DFB, indicating moderate to severe damage. Continuous exposure to DFB is potentially toxic to P. lineatus, causing heath disorders when the fish is exposed to the two highest DFB concentrations, which are applied to control parasites in aquaculture and to control mosquito populations in the environment. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 533–542, 2016.     

Toxic effects of an oil spill on fish early life stages may not be exclusively associated to PAHs: studies with Prestige oil and medaka (Oryzias latipes)

Polycyclic aromatic hydrocarbons (PAHs) are assumed to be the primary determinant of oil petroleum toxicity. Since the PAH content in Prestige oil was relatively high, we investigated the effects of different oil fractions (crude or weathered oil -0.05 to 50 g/L, and shaken or sonicated water accommodated fractions, WAFs, 25-100%, v/v) on the embryo-larval development of medaka (Oryzias latipes). Concentrations of summation operator16PAHs analyzed in the incubation medium were highest in the shaken WAF followed by the crude oil, the sonicated WAF and the weathered oil. Both oils (> or =0.25 g/L) induced developmental abnormalities whereas no significant effects were seen in the WAF exposures. In vivo morphometric analysis of the surface of the gallbladder during advanced embryo organogenesis (192 h post-fertilization, hpf) revealed significant dilation in both WAF exposures (>3 x 10(4) microm(2) at > or =25%, v/v, compared to <1.7 x 10(4) microm(2) at 0%, v/v) followed by the crude oil (>2.2 x 10(4) microm(2) at > or =0.05 g/L). Fluorescent aromatic compounds were observed in the gallbladder and the yolk sac of 168-hpf embryos exposed to all oil fractions. Results suggest the presence of components in both oils capable of penetrating the chorion and inducing a toxicity not observed in the WAFs. Hence, the hazard and risk assessment of Prestige oil should not be based solely on the presence of PAHs since proximity or direct contact may induce toxicity not associated exclusively to these compounds. This research offers a new hypothesis for explaining the reported biological observations, which could be correlated to direct oil exposure rather than the traditional mechanism of waterborne PAH exposure. Further research is needed to identify those oil components responsible for toxicity.     

Gasoline effects on biotransformation and antioxidant defenses of the freshwater fish Prochilodus lineatus

Biochemical biomarkers in the Neotropical freshwater fish Prochilodus lineatus were evaluated following acute exposures to the water-soluble fraction of gasoline (WSFG). Fish were exposed to the WSFG diluted to 5% in water (WSFG group) or only to water (Control group) for 6, 24 and 96 h and the gills and liver were removed for the biochemical analyses. Fish exposed to WSFG for 24 and 96 h showed significant increase in the activity of 7-ethoxyresorufin-O-deethylase (EROD) and glutathione-S-transferase (GST) both in liver and gills, pointing toward phase I and phase II biotransformation of the compounds present in the WSFG. The results also indicated the activation of antioxidant defenses in both the liver and gills after fish exposure to WSFG. The liver showed activation of catalase (CAT) and glutathione peroxidase (GPx) after 96 h exposure. An increase in hepatic content of reduced glutathione (GSH) together with decreased glutathione reductase (GR) activity was observed after 24 and 96 h of exposure to WSFG. In the gills, only catalase (CAT) activity augmented after 6 and 24 h of exposure and GSH content increased after 24 h of WSFG exposure. However, in both the organs, activation of the antioxidant defenses was not enough to prevent oxidative damage since they showed lipid peroxidation (LPO) at one of the experimental times: the liver after 6 h and the gills only after 96 h of exposure to WSFG. This may indicate better adaptation of the liver to longer exposures, starting from 24 h. As the gills are the first organ to be exposed to xenobiotics, the antioxidant defenses were triggered immediately upon exposure to WSFG and were able to prevent the occurrence of LPO during the initial times.     

Uptake and Biotransformation of 6,7-Dimethylquinoline and 6,8-Dimethylquinoline by Rainbow Trout (Salmo Gairdneri)

1. 6,7-Dimethylquinoline (6,7-DMQ) is readily taken up by rainbow trout and bioconcentrated in tissue after exposure to ca 1?mg/1 for 7.5?h. Mean bioconcentration factors (from water) were 21, 18, 6 and 14 for bile, liver, muscle and carcass respectively. Mean tissue concentrations after 69-96?h depuration were ND, ND, 0.54 and 0.48 μg/g for bile, liver, muscle and carcass respectively.

2. Major metabolites, following exposure to 6,7-DMQ, were conjugates (glucuronide or sulphate) of 7-hydroxymethyl-6-methylquinoline and 6-hydroxymethyl-7-methylquinoline. Mean concentration of metabolites in the bile were 500μg/g after 7.5?h exposure to ca 1?mg/1 and 1367 μg/g after 9.5?h exposure to ca 1?mg/1 and 69?h depuration.

3. 6,8-Dimethylquinoline (6,8-DMQ) is also readily bioconcentrated in fish tissue after exposure to ca 1?mg/1. Mean bioconcentration factors (from water) were 23, 20, 13 and 25 for bile, liver, muscle and carcass respectively. Mean tissue concentrations after 7?h exposure to ca 1?mg/1 and 63?h depuration were 4.0,0.67,0.49, and 3.2 μg/g respectively for bile, liver, muscle and carcass.

4. Major metabolites, following exposure to 6,8-DMQ were conjugates (glucuronide or sulphate) of 6,8-dimethyl-5-hydroxyquinoline, 6,8-dimethyl-7-hydroxyquinoline. 6,8-dimethyl-3-hydroxyquinoline and 6-hydroxymethyl-8-methylquinoline. Mean concentration of metabolites in the bile were 1278 μg/g after exposure to ca 1?mg/1 for 8?h and 1031 μg after exposure to ca 1?mg/1 for 7h and 63?h depuration.     

Influence of 17α-ethynylestradiol on CYP1A, GST and biliary FACs responses in male African sharptooth catfish (Clarias gariepinus) exposed to waterborne Benzo[a]Pyrene

Dose dependent effects of Benzo[a]pyrene (BaP) on cytochrome P4501A (CYP1A), glutathione-S-transferase (GST) and fluorescent aromatic compounds (FACs) metabolites biomarker responses were studied in African sharptooth catfish (Clarias gariepinus) following 24 h of waterborne exposures. Based on biomass of C. gariepinus in different tanks, BaP concentrations of 1.60, 3.44, and 18.21 μg/L that corresponded to 0.5, 1.0, and 5.0 mg/kg body weight were used. Significant induction of EROD activities in gill filaments was observed at all doses and the accumulation of FACs metabolites in bile was significantly different between groups. Accumulation of FACs metabolites in bile strongly correlated (r ² = 0.99) with BaP doses. Hepatic EROD activities were undetectable and no effect on GST activities was observed. The highest dose of BaP from the dose dependent study was further studied to assess the interactive and temporal responses of C. gariepinus on CYP1A, GST, and FACs metabolites biomarkers following exposure to either BaP alone, 17α-ethynylestradiol (EE₂) alone or a combination of both compounds at concentrations of 54.17 μg/L for BaP, 51.38 μg/L for EE₂ and 54.44 μg/L for each of both compounds. Based on biomass in each tank, these concentrations corresponded to 5 mg/kg body weight. While a group of six fish was sacrificed on day 0 from the control tank only, other groups of six fish were sacrificed after 1, 3, and 6 days of exposure from the control and exposed groups. Maximum induction of gill filament and hepatic EROD activities was observed after 1 day of exposure. Both EROD activities in gill filaments and liver were significantly induced by exposure to BaP alone or co-administration with EE₂. Gill filament EROD induction was significantly inhibited (50%) by co-administration of BaP and EE₂ compared to administration of BaP alone. Levels of FACs in bile for BaP and BaP + EE₂ exposed groups were significantly different from the control at all doses and time points. A significant induction of GST activities was observed in fish exposed to BaP and BaP + EE₂ after 3 days. Exposure to EE₂ alone caused significant induction of this enzyme after day 6. This study reports for the first time the significant antagonistic influence of EE₂ on BaP in gills of fish following waterborne exposures. The results also indicate that chemical mixtures may affect biomarker responses differently from compounds administered alone and that the sensitivity of CYP1A to interactive chemicals is different in gills and liver.     

胆汁酸-肠道菌群轴在2型糖尿病中的作用

目的 分析2型糖尿病(type 2 diabetes mellitus,T2DM)患者和正常人群胆汁酸与肠道菌群差异,探讨胆汁酸-肠道菌群轴在T2DM中的作用。方法 利用代谢组学、16S rRNA测序手段分别对T2DM患者中的血清胆汁酸含量及粪便肠道微生物进行检测及分析,结合斯皮尔曼相关性分析,明确胆汁酸-肠道菌群在T2DM中的代谢对话关系。结果 血清胆汁酸含量和肠道微生物的丰度在T2DM患者与正常人群中存在一定的差异。与正常人群相比,甘氨熊脱氧胆酸、牛磺鹅脱氧胆酸、甘氨鹅脱氧胆酸的含量在T2DM患者中显著降低;T2DM患者中肺炎克雷伯菌属、普拉梭菌属的相对丰度较正常人群明显升高,而狄氏副拟杆菌属、普雷沃菌属、艾克曼菌属、双歧杆菌属的相对丰度明显降低;斯皮尔曼相关性分析表明甘氨熊脱氧胆酸与狄氏副拟杆菌属、艾克曼菌属呈正相关,与克雷伯氏菌属呈负相关。结论 胆汁酸-肠道菌群轴是维持机体稳态的必要因素,在T2DM中发挥重要作用。     

Accumulation and toxicity of CuO and ZnO nanoparticles through waterborne and dietary exposure of goldfish (Carassius auratus)

Dietary and waterborne exposure to copper oxide (CuO) and zinc oxide (ZnO) nanoparticles (NPs) was conducted using a simplified model of an aquatic food chain consisting of zooplankton (Artemia salina) and goldfish (Carassius auratus) to determine bioaccumulation, toxic effects, and particle transport through trophic levels. Artemia contaminated with NPs were used as food in dietary exposure. Fish were exposed to suspensions of the NPs in waterborne exposure. ICP‐MS analysis showed that accumulation primarily occurred in the intestine, followed by the gills and liver. Dietary uptake was lower, but was found to be a potential pathway for transport of NPs to higher organisms. Waterborne exposure resulted in about a 10‐fold higher accumulation in the intestine. The heart, brain, and muscle tissue had no significant Cu or Zn. However, concentrations in muscle increased with NP concentration, which was ascribed to bioaccumulation of Cu and Zn released from NPs. Free Cu concentration in the medium was always higher than that of Zn, indicating CuO NPs dissolved more readily. ZnO NPs were relatively benign, even in waterborne exposure (p ≥ 0.05). In contrast, CuO NPs were toxic. Malondialdehyde levels in the liver and gills increased substantially (p < 0.05). Despite lower Cu accumulation, the liver exhibited significant oxidative stress, which could be from chronic exposure to Cu ions. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 119–128, 2015.     

In Vivo Effects of Environmental Concentrations of Produced Water on the Reproductive Function of Polar Cod (Boreogadus saida)

Offshore oil and gas drilling processes generate operational discharges such as produced water (PW), a complex mixture of seawater with polycyclic aromatic hydrocarbons (PAH) and alkylphenols (AP). Some of these compounds may interact with the endocrine system of marine organisms and alter reproductive functions. In this study, polar cod were exposed for up to 28 d to a mixture of PAH, alkylated PAH, and AP simulating the composition of North Sea PW, at low and high concentrations (1:2000 and 1:1000 dilution of the original concentrate, respectively). Potential adverse effects of PW on polar cod physiology were investigated through biomarkers of biotransformation (hepatic ethoxyresorufin O-deethylase [EROD] activity and bile PAH metabolites), endocrine disruption (plasma vitellogenin [VTG] levels and sex steroid concentrations), and gonad histology. Plasma sexual steroid levels in fish were not markedly affected by PW exposure, while higher plasma VTG concentrations were measured in females exposed to the high PW treatment for 7 and 28 d. In males exposed to the higher PW concentration, inhibition of spermatogenesis was observed after 28 d in addition to increase of melano-macrophage occurrence in testis. Females exposed to the high PW treatment for 21 d showed a significant increase of atresia incidence. Finally, a significant decrease in oocyte number was observed in high PW exposed female ovaries after 28 d of exposure.     

Evaluation of the Gill Filament-Based EROD Assay in African Sharptooth Catfish (Clarias gariepinus) as a Monitoring Tool for Waterborne PAH-Type Contaminants

The ability of African sharptooth catfish (Clarias gariepinus) in inducing cytochrome P-450 class 1A (CYP1A) and glutathione S-transferase (GST) biomarkers was determined in liver and gill filaments after 4 days of waterborne exposure to the polycyclic aromatic hydrocarbon, benzo[a]pyrene (B[a]P). Male (n=6) and female (n=6) fish were exposed to B[a]P added to the water (30 μg/l) corresponding to 5 mg/kg total body weight. Five female and six male fish, exposed to acetone alone added to the water served as controls. The 7-ethoxyresurufin-O-deethylase (EROD) activity was measured in hepatic microsomes and gill filaments. In addition, GST activity was determined in the hepatic cytosolic fraction and fluorescent aromatic compounds (FACs) in bile and biliverdin. Benzo[a]pyrene strongly induced EROD activities in gill filaments of both sex. Levels of FACs per ml of bile were 17-fold higher in exposed fish compared to the controls. Correlations between induction of EROD activities in gill filaments and liver and between induction of EROD activities in gill filaments and levels of biliary FACs metabolites were strong. GST activities in the hepatic cytosolic fraction were similar amongst the treated and control groups. This is the first report on studies determining EROD activities in gill filaments and hepatic tissue, FACs in bile and GST in hepatic tissues of C. gariepinus after waterborne exposure to B[a]P. The findings suggest that the gill filament-based EROD assay in C. gariepinus can be used to monitor the pollution of AhR agonists in aquatic ecosystems in eastern and southern African countries.     

Genotoxic effects of produced waters in mosquito fish (Gambusia affinis)

The aim of this study was to assess the potential genotoxic effects of produced water (PW) from an Italian on-shore oil plant. Produced water is a complex mixture containing residual hydrocarbons, trace elements, naturally occurring radioactive material and potentially toxic treatment chemicals such as biocides, dispersants, detergents and scale inhibitors used in oil production. The test organism, mosquito fish (Gambusia affinis), was divided into male and female groups and exposed for 8 days in the laboratory to 50% concentrations of different produced waters: PW before treatment and after settling treatment. The fish were also exposed to lower concentrations (10%) of the same PW for 30 days. DNA damage was evaluated in erythrocytes by single cell gel electrophoresis (Comet assay) and micronucleus test, while an oxidative stress biomarker, was assessed. Polycyclic aromatic hydrocarbons (PAH) metabolites in bile were also evaluated. A higher sensitivity in biomarker responses was found in females in comparison to males. An increase in DNA strand breaks was observed in both genders after 30 days exposure and a statistically significant increase of micronucleated cells was found in females after 8 days exposure. A positive correlation between presence of micronucleated cells and PAH metabolites in bile was also observed.     

The effects of elutriates from PAH and heavy metal polluted sediments on Crassostrea gigas (Thunberg) embryogenesis,larval growth and bio-accumulation by the larvae of pollutants from sedimentary origin

The release, bio-availability and toxicity of contaminants, when sediments are resuspended have been examined, studying concurrently their effects on the embryogenesis and on the larval growth of the Crassostrea gigas larvae and their bio-accumulation in those organisms. Three characteristic sediments have been selected (one contaminated by PAHs, a second by heavy metals and the last by the both pollutants). The organisms were directly exposed to elutriates obtained from each sediment or fed on algae (Isochrysis galbana) contaminated with the same elutriates. The elutriates used in this study show contamination levels similar to those observed in some polluted coastal and estuary environments. The larval growth test has appeared to be more sensitive that the embryotoxicity test. The biological effects and the contaminant bio-accumulation were more pronounced when larvae were directly exposed to different elutriates. In the case of PAHs, the contamination of algae was sufficient to lead to effect on the larval growth of the Crassostrea gigas. In all cases, a fraction of contaminants adsorbed on suspended particles was bio-available and accumulated by the larvae. This study has shown that resuspending polluted sediments constitutes a threat to pelagic organisms and than the C. gigas larval growth may be proposed as a test to protect the most sensitive areas.     

Reproductive toxicity of the water accommodated fraction (WAF) of crude oil in the polychaetes Arenicola marina (L.) and Nereis virens (Sars)

Accidental pollution incidents are common in the marine environment and are often caused by oil-related activities. Here the potential of such an incident to disrupt reproduction in two polychaete species is investigated, using an environmentally relevant preparation of weathered Forties crude oil, i.e. the water accommodated fraction (WAF). Oocytes were collected and exposed to three concentrations of WAF for 1h prior to the addition of sperm, so that fertilization took place under exposure conditions. Fertilization success was significantly reduced in both species by an exposure to WAF concentrations equivalent to 0.38 mgL(-1) PAHs, to just 26.8% in Arenicola marina compared to 76% in Nereis virens. The effects of WAF exposure on fertilization were greatly enhanced at lower sperm concentrations in N. virens, with a complete lack of fertilization reactions observed at sperm concentrations of 10(3)sperm per mL. We therefore suggest a mechanism of toxicity related to sperm swimming behaviour, resulting in reduced sperm:egg collision rates. WAF was found to reduce post-fertilization development rates and have teratogenic effects on early embryonic stages in both species, which exhibited abnormal cleavage patterns and high levels of fluctuating asymmetry. These results illustrate how the presence of crude oil in its soluble form in seawater at the time of a spawning event for either A. marina or N. virens could impact on fertilization success with implications for the fertilization ecology of these free spawning marine invertebrates.     

Structure Elucidation and Thermospray High-Performance Liquid Chromatography/Mass Spectroscopy (HPLC/MS) of the Microbial and Mammalian Metabolites of the Antimalarial Arteether

Microbial metabolism studies of the antimalarial drug arteether (1) have shown that arteether is metabolized to six new metabolites in addition to those previously reported (3). Large-scale fermentations with Cunninghamella elegans (ATCC 9245) and Streptomyces lavendulae (L-105) have resulted in the characterization of these metabolites primarily by two-dimensional nuclear magnetic resonance (2D-NMR) methods as 9-hydroxyarteether (2), a ring rearrangement metabolite (3), 3-hydroxy-11-epi-deoxydihydroartemisinin (4), 9-hydroxyarteether (5), 2-hydroxyarteether (6), and 14-hydroxyarteether (7). Thermospray mass spectroscopy/high-performance liquid chromatographic analyses have shown that four of these metabolites (2, 5, 6, 7) are also present in rat liver microsome preparations.     

In vitro/in vivo investigations to examine the gender differences in the pharmacokinetics of novel oral Janus kinase (JAK) inhibitor ASP015K and sulfate metabolite M2 in rats

Abstract

1. Although marked gender differences have been reported for the exposure level of the sulfate metabolite M2 of ASP015K in rats, no such differences have been reported for the unchanged drug. To clarify the cause of these pharmacokinetic gender differences, we investigated the in vitro hepatic sulfation, glucuronidation, and cytochrome P450 (CYP) metabolism of ASP015K in rat liver cytosols or rat liver microsomes. Further, in vivo excretion and metabolic profiles were investigated using rat urine, bile, and feces post-ASP015K administration.

2. In vitro metabolism study using liver cytosols clearly suggested that the gender differences in the M2 exposure were mainly attributed to the female-predominant ASP015K metabolism mediated by sulfotransferase (SULT). Metabolic profiles in urine and bile from male rats suggested that the major elimination pathway of ASP015K is glucuronidation in rats. No remarkable gender differences in the in vitro glucuronidation were observed.

3. The contribution of the sulfation pathway to the clearance of ASP015K was markedly lower than that of the glucuronidation pathway in both male and female rats. These results might explain why gender differences were not marked for ASP015K exposure but were for M2.     

The histopathological,cytopathological and ultrastructural effects of carbaryl on gills of Oreochromis niloticus (Linnaeus, 1758)

Ultrastructural and histopathological reponses in the organs of living organisms are important and useful tools to determine the health condition and the effects of pollutants, such as pesticides, on the organisms. The aim of this study is to determine possible histopathological, cytopathological and ultrastructural alterations in gills of Oreochromis niloticus individuals exposed to 850 μg/L carbaryl standart at 7th, 14th and 21st days with light and electron microscopes. The fish were exposed to carbaryl for 21 days and the histopatological, ultrastructural and cytopathological alterations occuring in the gill tissues of organisms were determined by light, Scanning and Transmission Electron Microscopes (SEM and TEM). At the end of the study, it was observed that carbaryl caused both histopathological and cytopathological changes in the gills of O. niloticus. It has been determined that the most of the pathological changes in the exposed organisms are the metabolic defence reactions.     

An assessment of the importance of exposure routes to the uptake and internal localisation of fluorescent nanoparticles in zebrafish (Danio rerio), using light sheet microscopy

A major challenge in nanoecotoxicology is finding suitable methods to determine the uptake and localisation of nanoparticles on a whole-organism level. Some uptake methods have been associated with artefacts induced by sample preparation, including staining for electron microscopy. This study used light sheet microscopy (LSM) to define the uptake and localisation of fluorescently labelled nanoparticles in living organisms with minimal sample preparation. Zebrafish (Danio rerio) were exposed to fluorescent gold nanoparticles (Au NPs) and fluorescent polystyrene NPs via aqueous or dietary exposure. The in vivo uptake and localisation of NPs were investigated using LSM at different time points (1, 3 and 7?days). A time-dependent increase in fluorescence was observed in the gut after dietary exposure to both Au NPs and polystyrene NPs. No fluorescence was observed within gut epithelia regardless of the NP exposure route indicating no or limited uptake via intestinal villi. Fish exposed to polystyrene NPs through the aqueous phase emitted fluorescence signals from the gills and intestine. Fluorescence was also detected in the head region of the fish after aqueous exposure to polystyrene NPs. This was not observed for Au NPs. Aqueous exposure to Au NPs resulted in increased relative swimming distance, while no effect was observed for other exposures. This study supports that the route of exposure is essential for the uptake and subsequent localisation of nanoparticles in zebrafish. Furthermore, it demonstrates that the localisation of NPs in whole living organisms can be visualised in real-time, using LSM.