Investigating endocrine-disrupting properties of chemicals in fish and amphibians: Opportunities to apply the 3Rs.

Many regulations are beginning to explicitly require investigation of a chemical's endocrine-disrupting properties as a part of the safety assessment process for substances already on or about to be placed on the market. Different jurisdictions are applying distinct approaches. However, all share a common theme requiring testing for endocrine activity and adverse effects, typically involving in vitro and in vivo assays on selected endocrine pathways.

Polyester-derived microfibre impacts on the soil-dwelling earthworm Lumbricus terrestris.

Microplastic (MP) pollution is everywhere. In terrestrial environments, microfibres (MFs) generated from textile laundering are believed to form a significant component of MPs entering soils, mainly through sewage sludge and compost applications. The aim of this study was to assess the effect of MFs on a keystone soil organism.

Opportunistic disease in yellow perch in response to decadal changes in the chemistry of oil sands-affected waters.

Oil sands-affected water from mining must eventually be incorporated into the reclaimed landscape or treated and released. However, this material contains petrogenic organic compounds, such as naphthenic acids and traces of polycyclic aromatic hydrocarbons. This has raised concerns for impacts of oil sands process-affected waters on the heath of wildlife and humans downstream of receiving environments.

Plastic Bag Derived-Microplastics as a Vector for Metal Exposure in Terrestrial Invertebrates.

Microplastics are widespread contaminants in terrestrial environments but comparatively little is known about interactions between microplastics and common terrestrial contaminants such as zinc (Zn). In adsorption experiments fragmented HDPE bags c. one mm in size showed similar sorption characteristics to soil.

Characteristic and functional analysis of toll-like receptors (TLRs) in the lophotrocozoan, Crassostrea gigas, reveals ancient origin of TLR-mediated innate immunity.

The evolution of TLR-mediated innate immunity is a fundamental question in immunology. Here, we report the characterization and functional analysis of four TLR members in the lophotrochozoans Crassostreagigas (CgTLRs). All CgTLRs bear a conserved domain organization and have a close relationship with TLRs in ancient non-vertebrate chordates.

Test concentration setting for fish in vivo endocrine screening assays.

Fish in vivo screening methods to detect endocrine active substances, specifically interacting with the hypothalamic-pituitary-gonadal axis, have been developed by both the Organization for Economic Co-operation and Development (OECD) and United States Environmental Protection Agency (US-EPA). In application of these methods, i.e.

Immunotoxic effects of oil sands-derived naphthenic acids to rainbow trout.

Naphthenic acids are the major organic constituents in waters impacted by oil sands. To investigate their immunotoxicity, rainbow trout (Oncorhynchus mykiss) were injected with naphthenic acids extracted from aged oil sands tailings water. In two experiments, rainbow trout were injected intraperitoneally with 0, 10, or 100 mg/kg of naphthenic acids, and sampled after 5 or 21 d.

Mode of sexual differentiation and its influence on the relative sensitivity of the fathead minnow and zebrafish in the fish sexual development test.

Exogenous treatment of fish with natural sex hormones and their mimics has been shown to influence gonadal differentiation resulting in biased phenotypic sex-ratios. This has lead to the development of the Fish Sexual Development Test (FSDT) as a method for the detection of endocrine active chemicals. Proposed test organisms include the medaka, zebrafish (ZF) and stickleback, although the guideline also allows for inclusion of species such as the fathead minnow (FHM), provided the test duration allows for sufficient sexual differentiation.

Short-term exposure to the environmentally relevant estrogenic mycotoxin zearalenone impairs reproduction in fish.

Zearalenone (ZON) is one of the worldwide most common mycotoxins and exhibits estrogenic activity in the range of natural steroid estrogens such as 17β-estradiol (E2). The occurrence of ZON has been reported in drainage water, soil, wastewater effluents and rivers, but its ecotoxicological effects on fish have hardly been investigated. In this study the estrogenic potency of the ZON was compared to E2 in a recombinant yeast estrogen screen (rYES) and the effects of waterborne ZON exposure on reproduction, physiology and morphology of zebrafish (Danio rerio) were investigated in a 42-day reproduction experiment.

Estrogenic wastewater treatment works effluents reduce egg production in fish.

Estrogenic chemicals found within wastewater treatment work (WwTW) effluents have been shown individually to inhibit reproduction in fish, but the impact of the WwTW effluents themselves and the complex mixtures of environmental estrogens and other endocrine disrupting chemicals they contain has not been established. In this investigation, the effect of exposure to three WwTW effluents, with differing levels of estrogenic activity, was assessed on egg production in pair-breeding fathead minnow. Exposure to two of the three effluents tested resulted in a reduced egg production (by 28% for effluent I at a dilution of 50% and by 44% for effluent III at full strength), which was proportional to the estrogenic content of the effluents.

A practicable laboratory flow-through exposure system for assessing the health effects of effluents in fish.

The knowledge that exposure to estrogenic wastewater treatment work (WwTW) effluents induces a range of reproductive abnormalities in fish has highlighted the need to understand the wider health effects of effluents. Access to laboratory-based testing systems for WwTW effluents could greatly facilitate this endeavour. In this investigation, a laboratory-based test system was developed and applied for WwTW effluents using fathead minnows (Pimephales promelas).

Gene expression profiling for understanding chemical causation of biological effects for complex mixtures: a case study on estrogens.

Gene expression profiling offers considerable potential for identifying chemical causation of effects induced in exposures to complex mixtures, and for understanding the mechanistic basis for their phenotypic effects. We characterized gene expression responses in livers and gonads of fathead minnow (Pimephales promelas) exposed (for 14-21 days) to estrogenic wastewater treatment works final effluents with varying potencies and assessed the extent to which these expression profiles mapped with those induced by individual steroid estrogens present in the effluents (17beta-estradiol and 17alpha-ethinylestradiol) and, thus, were diagnostic of estrogen exposure. For these studies, we adopted a targeted approach (via real-time PCR) with a suite of 12 genes in liver and 21 genes in gonad known to play key roles in reproduction, growth and development (processes controlled by estrogens) and responses were compared with effects on phenotypic end points indicative of feminization.

Health impacts of estrogens in the environment, considering complex mixture effects.

Background: Environmental estrogens in wastewater treatment work (WwTW) effluents are well established as the principal cause of reproductive disruption in wild fish populations, but their possible role in the wider health effects of effluents has not been established.

Objectives: We assessed the contribution of estrogens to adverse health effects induced in a model fish species by exposure to WwTW effluents and compared effects of an estrogen alone and as part of a complex mixture (i.e.

Associations between altered vitellogenin concentrations and adverse health effects in fathead minnow (Pimephales promelas).

Mechanism specific biomarkers are used in ecotoxicology to identify classes of chemicals and to inform on their presence in the environment, but their use in signalling for adverse effects has been limited by a poor understanding of their associated links with health. In this study an experimental analysis was undertaken to investigate how induction or suppression of an estrogen-dependent biomarker, vitellogenin (VTG), related to health effects in fathead minnow (Pimephales promelas, FHM). Exposure to an oestrogen agonist, estradiol (29 and 60 ng/L), resulted in rapid induction of VTG (elevated plasma concentrations within 2 days of exposure) in male FHM that was subsequently slow to clear from the plasma (concentrations remained elevated 70 days after cessation of exposure).

Gene expression profiles revealing the mechanisms of anti-androgen- and estrogen-induced feminization in fish.

Environmental anti-androgens are increasingly being recognized as potential contributing factors in the chemically induced feminization of wild fish because, by blocking androgen action, they can produce phenotypic effects similar to environmental estrogens. The molecular mechanisms by which anti-androgens and estrogens exert feminizing effects, however, have not been systematically compared. Using a targeted approach, we profiled the expression responses of a suite of 22 genes involved in reproduction, growth and development (processes controlled by androgens and estrogens) in the liver and gonad in adult male and female fathead minnow (Pimephales promelas) exposed to the model anti-androgen flutamide and the model synthetic estrogen 17alpha-ethinylestradiol (EE(2)).

An optimised experimental test procedure for measuring chemical effects on reproduction in the fathead minnow, Pimephales promelas.

The production of viable offspring is fundamental to the survival of any population. Tests that quantify effects on reproduction can, therefore, inform on the potential for long-term health effects of exposure to endocrine active chemicals. Surprisingly little is known, however, about the reproductive capacity of laboratory fish species used for chemical testing.

Multiple molecular effect pathways of an environmental oestrogen in fish.

Complex interrelationships in the signalling of oestrogenic effects mean that environmental oestrogens present in the aquatic environment have the potential to disrupt physiological function in fish in a more complex manner than portrayed in the present literature. Taking a broader approach to investigate the possible effect pathways and the likely consequences of environmental oestrogen exposure in fish, the effects of 17beta-oestradiol (E(2)) were studied on the expression of a suite of genes which interact to mediate growth, development and thyroid and interrenal function (growth hormone GH (gh), GH receptor (ghr ), insulin-like growth factor (IGF-I) (igf1), IGF-I receptor (igf1r ), thyroid hormone receptors-alpha (thra) and -beta (thrb) and glucocorticoid receptor (gr )) together with the expression analyses of sex-steroid receptors and ten other genes centrally involved in sexual development and reproduction in fathead minnow (fhm; Pimephales promelas). Exposure of adult fhm to 35 ng E(2)/l for 14 days induced classic oestrogen biomarker responses (hepatic oestrogen receptor 1 and plasma vitellogenin), and impacted on the reproductive axis, feminising "male" steroidogenic enzyme expression profiles and suppressing genes involved in testis differentiation.

An assessment of the model of concentration addition for predicting the estrogenic activity of chemical mixtures in wastewater treatment works effluents.

The effects of simple mixtures of chemicals, with similar mechanisms of action, can be predicted using the concentration addition model (CA). The ability of this model to predict the estrogenic effects of more complex mixtures such as effluent discharges, however, has yet to be established. Effluents from 43 U.

Relative potencies and combination effects of steroidal estrogens in fish.

The natural steroids estradiol-17beta (E2) and estrone (E1) and the synthetic steroid ethynylestradiol-17alpha (EE2) have frequently been measured in waters receiving domestic effluents. All of these steroids bind to the estrogen receptor(s) and have been shown to elicit a range of estrogenic responses in fish at environmentally relevant concentrations. At present, however, no relative potency estimates have been derived for either the individual steroidal estrogens or their mixtures in vivo.