Richard T. Di Giulio

Prooxidant and antioxidant mechanisms in aquatic organisms

Abstract A consequence of all aerobic life is the production of potentially harmful, partially reduced species of molecular oxygen (oxyradicals) that occurs as a result of normal oxygen metabolism. In view of this potential for harm, aerobic organisms have evolved a complex armamentorium of defenses to protect them against the possible ravages of oxyradical production. A recent growing body of evidence indicates a marked penchant for augmented oxyradical production in aquatic organisms resulting largely from anthropogenic pollutant outfalls. Two consequences of environmental contamination might be envisaged; certain compounds which cause increased oxyradical flux could exacerbate the deleterious effects of oxyradicals or the increased fluxes could affect induction of antioxidant enzymes thereby overcoming the stresses imposed by the contaminating compound(s). This review describes the various adaptational mechanisms and responses, including the toxicological consequences, associated with oxyradical production. The broad phylogenetic spectrum of examples discussed herein illustrate that while there are numerous differences among the phyla with respect to both pro- and anti-oxidant mechanisms to establish a vigorous basis of inquiry, enough striking similarities also exist to suggest the use of aquatic organisms as alternative models to mammals in various areas of oxyradical research, e.g. aging and cancer. In addition, the potential use of anti-oxidant responses, e.g. induction of specific anti-oxidant enzymes as biomarkers of environmental contamination is reviewed.

The Toxicology of Fishes

Ecological Risk Assessments (ERA) are performed to evaluate the likelihood of adverse ecological effects occurring as a result of exposure to physical or chemical stressors. These stressors are defined as any biological, physical, or chemical factor that causes adverse responses in the environment. An ERA evaluates any potential harm that human activities have on living organisms within ecosystems. Within the framework of an Ecological Risk Assessment, scientific information is developed, organized and presented so that it is relevant to environmental decisions.General Principles Introduction, R.T. Di Giulio and D.E. Hinton Bioavailability of Chemical Contaminants in Aquatic Systems, R.J. Erickson, J.W. Nichols, P.M. Cook, and G.T. Ankley Toxicokinetics in Fishes, K.M. Kleinow, J.W. Nichols, W.L. Hayton, J.M. McKim, and M.G. Barron Biotransformation in Fishes, D. Schlenk, M. Celander, E.P. Gallagher, S. George, M. James, S.W. Kullman, P. van den Hurk, and K. Willett Receptor-Mediated Mechanisms of Toxicity, M.E. Hahn and E.V. Hestermann Reactive Oxygen Species and Oxidative Stress, R.T. Di Giulio and J.N. Meyer Key Target Systems and Organismal Effects Liver Toxicity, D.E. Hinton, H. Segner, D.W.T. Au, S.W. Kullman, and R.C. Hardman The Osmoregulatory System, S.E. Wendelaar Bonga and R.A.C. Lock Toxic Responses of the Fish Nervous System, S.P. Bradbury, R.W. Carlson, T.R. Henry, S. Padilla, and J. Cowden The Endocrine System, P. Thomas The Immune System of Fish: A Target Organ of Toxicity, E. Carlson and J.T. Zelikoff Chemical Carcinogenesis in Fishes, J.M. Rotchell, M.R. Miller, D.E. Hinton, R.T. Di Giulio, and G.K. Ostrander Toxicity Resistance, P.A. Van Veld and D.E. Nacci Methodologies and Applications Exposure Assessment and Modeling in the Aquatic Environment, D. Mackay and L. Milford Fish Toxicity Studies, G.M. Rand Biomarkers, D. Schlenk, R. Handy, S. Steinert, M.H. Depledge, and W. Benson Aquatic Ecosystems for Ecotoxicological Research: Considerations in Design Analysis for Fish, T.W. La Point, J.H. Kennedy, J.K. Stanley, and P. Balci Ecological Risk Assessment, D.R. Mount and T.R. Henry Case Studies Mining Impacts on Fish in the Clark Fork River, Montana: A Field Ecotoxicology Case Study, S.N. Luoma, J.N. Moore, A. Farag, T.H. Hillman, D.J. Cain, and M. Hornberger Toxicology of Synthetic Pyrethroid Insecticides in Fish: A Case Study, J.R. Coats Reproductive Impairment of Great Lakes Lake Trout by Dioxin-Like Chemicals, D.E. Tillitt, P.M. Cook, J.P. Giesy, W. Heideman, and R.E. Peterson The Effects of Polycyclic Aromatic Hydrocarbons in Fish from Puget Sound, Washington, L.L. Johnson, M.R. Arkoosh, C.F. Bravo, T.K. Collier, M.M. Krahn, J.P. Meador, M.S. Myers, W.L. Reichert, and J.E. Stein Effects of the Exxon Valdez Oil Spill on Pacific Herring in Prince William Sound, Alaska, G.D. Marty Case Study: Pulp and Paper Mill Impacts, M.G. Dube, K.R. Munkittrick, and L.M. Hewitt Estrogenic Effects of Treated Sewage Effluent on Fish: Steroids and Surfactants in English Rivers, C.R. Tyler, E.J. Routledge, and R. van Aerle Index

Effects of Black Rock Harbor sediments on indices of biotransformation, oxidative stress, and DNA integrity in channel catfish

Selected biochemical responses were measured in channel catfish (Ictalurus punctatus) exposed in the laboratory to sediments obtained from either of two sites in Long Island Sound, CT. These sites were Black Rock Harbor, which is highly contaminated with various aromatic hydrocarbons, and a relatively uncontaminated reference site. Livers and bile were removed from fish on days 2, 7, 14, and 28 following the initiation of sediment exposures and examined for responses associated with (1) xenobiotic biotransformation, (2) oxidative stress, and (3) DNA integrity. Relative to reference sediment exposures in fish, exposures to Black Rock Harbor sediments elicited on at least two of the four sampling points significant (P < 0.05) increases in: (1) phase I biotransformation enzyme activities (EROD and ECOD) and concentrations of bile metabolites of polycyclic aromatic hydrocarbons (PAHs); (2) antioxidant enzyme activities (SOD and catalase), concentrations of reduced and oxidized glutathione, and malondialdehyde (an index of lipid peroxidation); and (3) the fraction of DNA as single-stranded DNA following alkaline unwinding (an index of strand breaks). The results of this study support hypotheses concerning mechanistic relationships in benthic fish among the metabolism of sediment-associated aromatic hydrocarbons, the generation of free radical intermediates, and mutagenesis. Furthermore, these results support the utility of biochemical responses as tools for assessing contaminant exposures and sublethal effects in aquatic animals.

Sulfidation of Silver Nanoparticles: Natural Antidote to Their Toxicity

Nanomaterials are highly dynamic in biological and environmental media. A critical need for advancing environmental health and safety research for nanomaterials is to identify physical and chemical transformations that affect the nanomaterial properties and their toxicity. Silver nanoparticles, one of the most toxic and well-studied nanomaterials, readily react with sulfide to form Ag(0)/Ag2S core-shell particles. Here, we show that sulfidation decreased silver nanoparticle toxicity to four diverse types of aquatic and terrestrial eukaryotic organisms (Danio rerio (zebrafish), Fundulus heteroclitus (killifish), Caenorhabditis elegans (nematode worm), and the aquatic plant Lemna minuta (least duckweed)). Toxicity reduction, which was dramatic in killifish and duckweed even for low extents of sulfidation (about 2 mol % S), is primarily associated with a decrease in Ag(+) concentration after sulfidation due to the lower solubility of Ag2S relative to elemental Ag (Ag(0)). These results suggest that even partial sulfidation of AgNP will decrease the toxicity of AgNPs relative to their pristine counterparts. We also show that, for a given organism, the presence of chloride in the exposure media strongly affects the toxicity results by affecting Ag speciation. These results highlight the need to consider environmental transformations of NPs in assessing their toxicity to accurately portray their potential environmental risks.

Synergistic embryotoxicity of polycyclic aromatic hydrocarbon aryl hydrocarbon receptor agonists with cytochrome P4501A inhibitors in Fundulus heteroclitus.

Widespread contamination of aquatic systems with polycyclic aromatic hydrocarbons (PAHs) has led to concern about effects of PAHs on aquatic life. Some PAHs have been shown to cause deformities in early life stages of fish that resemble those elicited by planar halogenated aromatic hydrocarbons (pHAHs) that are agonists for the aryl hydrocarbon receptor (AHR). Previous studies have suggested that activity of cytochrome P4501A, a member of the AHR gene battery, is important to the toxicity of pHAHs, and inhibition of CYP1A can reduce the early-life-stage toxicity of pHAHs. In light of the effects of CYP1A inhibition on pHAH-derived toxicity, we explored the impact of both model and environmentally relevant CYP1A inhibitors on PAH-derived embryotoxicity. We exposed Fundulus heteroclitus embryos to two PAH-type AHR agonists, β-naphthoflavone and benzo(a)pyrene, and one pHAH-type AHR agonist, 3,3′,4,4′,5-pentachlorobiphenyl (PCB-126), alone and in combination with several CYP1A inhibitors. In agreement with previous studies, coexposure of embryos to PCB-126 with the AHR antagonist and CYP1A inhibitor α-naphthoflavone decreased frequency and severity of deformities compared with embryos exposed to PCB-126 alone. In contrast, embryos coexposed to the PAHs with each of the CYP1A inhibitors tested were deformed with increased severity and frequency compared with embryos dosed with PAH alone. The mechanism by which inhibition of CYP1A increased embryotoxicity of the PAHs tested is not understood, but these results may be helpful in elucidating mechanisms by which PAHs are embryotoxic. Additionally, these results call into question additive models of PAH embryotoxicity for environmental PAH mixtures that contain both AHR agonists and CYP1A inhibitors.

Cumulative impacts of mountaintop mining on an Appalachian watershed

Mountaintop mining is the dominant form of coal mining and the largest driver of land cover change in the central Appalachians. The waste rock from these surface mines is disposed of in the adjacent river valleys, leading to a burial of headwater streams and dramatic increases in salinity and trace metal concentrations immediately downstream. In this synoptic study we document the cumulative impact of more than 100 mining discharge outlets and approximately 28 km2 of active and reclaimed surface coal mines on the Upper Mud River of West Virginia. We measured the concentrations of major and trace elements within the tributaries and the mainstem and found that upstream of the mines water quality was equivalent to state reference sites. However, as eight separate mining-impacted tributaries contributed their flow, conductivity and the concentrations of selenium, sulfate, magnesium, and other inorganic solutes increased at a rate directly proportional to the upstream areal extent of mining. We found strong linear correlations between the concentrations of these contaminants in the river and the proportion of the contributing watershed in surface mines. All tributaries draining mountaintop-mining-impacted catchments were characterized by high conductivity and increased sulfate concentration, while concentrations of some solutes such as Se, Sr, and N were lower in the two tributaries draining reclaimed mines. Our results demonstrate the cumulative impact of multiple mines within a single catchment and provide evidence that mines reclaimed nearly two decades ago continue to contribute significantly to water quality degradation within this watershed.

HERITABLE ADAPTATION AND FITNESS COSTS IN KILLIFISH (FUNDULUS HETEROCLITUS) INHABITING A POLLUTED ESTUARY

Adaptation to contaminants in the environment has been studied extensively in microbes, insects, and plants, and increasing evidence suggests that certain vertebrate populations as well are evolving in response to pollution. Here, we show that F1 and F2 laboratory-raised offspring of killifish (Fundulus heteroclitus, also known as mummichog) from a highly contaminated site on the Elizabeth River (Virginia, USA) are more resistant to the toxicity of Elizabeth River sediments than are offspring of reference site killifish. This resistance is more marked in the F1 than in the F2 generation, but it remains significant in the F2 generation, indicating that the resistant phenotype in the feral Elizabeth River killifish is based on both genetic and nongenetic mechanisms. In addition, both the F1 and F2 generation offspring of the Elizabeth River killifish are more susceptible to other stressors, both anthropogenic (photoenhanced toxicity) and natural (hypoxia), suggesting that the changes that have conferred resistance to the toxicity of the Elizabeth River sediments carry a cost of reduced fitness in other contexts. Corresponding Editor: J. E. McDowell.

Biotic and abiotic interactions in aquatic microcosms determine fate and toxicity of Ag nanoparticles: part 2-toxicity and Ag speciation.

To study the effects of complex environmental media on silver nanoparticle (AgNP) toxicity, AgNPs were added to microcosms with freshwater sediments and two species of aquatic plants (Potamogeton diversifolius and Egeria densa), followed by toxicity testing with microcosm surface water. Microcosms were designed with four environmental matrices in order to determine the contribution of each environmental compartment to changes in toxicity: water only (W), water + sediment (WS), water + plants (WP), and water + plants + sediment (WPS). Silver treatments included AgNPs with two different coatings, gum arabic (GA-AgNPs) or polyvinylpyrollidone (PVP-AgNPs), as well as AgNO(3). Water samples taken from the microcosms at 24 h postdosing were used in acute toxicity tests with two standard model organisms, early life stage zebrafish (Danio rerio) and Daphnia magna. Speciation of Ag in these samples was analyzed using Ag L3-edge X-ray absorption near edge spectroscopy (XANES). Silver speciation patterns for the nanoparticle treatments varied significantly by coating type. While PVP-AgNPs were quite stable and resisted transformation across all matrices (>92.4% Ag(0)), GA-AgNP speciation patterns suggest significantly higher transformation rates, especially in treatments with plants (<69.2% and <58.8% Ag(0) in WP and WPS, respectively) and moderately increased transformation with sediments (<85.6% Ag(0)). Additionally, the presence of plants in the microcosms (with and without sediments) reduced both the concentration of Ag in the water column and toxicity for all Ag treatments. Reductions in toxicity may have been related to decreased water column concentrations as well as changes in the surface chemistry of the particles induced by organic substances released from the plants.

Heavy metals in tissues of waterfowl from the Chesapeake Bay, USA

Concentrations of cadmium, lead, copper and zinc were measured in 774 livers, 266 kidneys and 271 ulnar bones from 15 species of ducks obtained from the Chesapeake Bay region. A major purpose of this study was to elucidate relationships between food habits and tissue accumulations of heavy metals in Chesapeake Bay waterfowl. Liver and kidney concentrations of cadmium were highest among two carnivorous seaduck species, Clangula hyemalis and Melanitta deglandi. In contrast, lead concentrations in tissues were generally highest in largely herbivorous species, such as Anas platyrhynchos, Anas rubripes and Anas strepera. Spent shot may be an important source for tissue burdens of lead in these ducks. No marked trends were observed between food habits and tissue concentrations of the nutrient elements, copper and zinc.

Oxidant, mixed-function oxidase and peroxisomal responses in channel catfish exposed to a bleached kraft mill effluent

Channel catfish (Ictalarus punctatus) were exposed to 0, 10, 20, and 40% bleached kraft pulp and paper mill effluent (BKME) for up to two weeks. Fish were sacrificed on days 1, 3,7, and 14 of exposure. Blood, liver, and gill tissues were assayed for biochemical responses associated with oxidative stress (superoxide dismutase, catalase, and glutathione peroxidase activities and methemoglobin, reduced glutathione, and malondialdehyde concentrations), peroxisome proliferation (palmitoyl-CoA oxidase and lauroyl-CoA oxidase activities) and the mixed-function oxidase system (cytochrome P-450 difference spectra and ethoxyresorufin O-deethylase activities). There were significant dose-related increases in hepatic catalase and ethoxyresorufin O-deethylase activities and significant decreases in hepatic reduced glutathione concentrations. There were also significant increases in lauroyl-CoA oxidase and palmitoyl-CoA oxidase activities, both components of the fatty acid beta-oxidation system and located in peroxisomes. No statistically significant changes were seen in the other components studied. The increases in catalase and the fatty-acid oxidase activities suggest that BKME exposure may have resulted in peroxisome proliferation in catfish hepatic tissue.