Can roach, Rutilus rutilus, adapt to the harmful effects of oestrogen exposure from waste water treatment work effluents?

Summary
Major worldwide attention has focused on the observations of disruptions of reproduction in both wildlife and human populations that can result from exposure to chemicals that interfere with the body's hormone signalling systems (so-called endocrine disrupting chemicals; EDCs) impacting on reproductive health. We have shown that reproduction in fish (roach, a common member of the carp family of fish) living in many UK Rivers has been damaged by exposure to EDCs contained in the wastewater treatment works (WwTW) effluents and the chemicals responsible for these effects include natural oestrogen hormones and pharmaceutical oestrogens in the contraceptive pill.
Feminised roach have a reduced capability to breed under competitive breeding conditions and we have evidence (as yet, unpublished) for reduced breeding population sizes in wild roach living in rivers with a high effluent content. Populations with low numbers of breeding individuals lose genetic variation over multiple generations with a greater risk of extinction. Nevertheless, we find that in some stretches of these rivers with high oestrogenic exposure roach populations appear to be reproductively self-sustaining. Establishing whether fish (here roach) have adapted to oestrogenic contaminants, how they do this (the mechanisms) and the possible fitness costs of these adaptations are essential in understanding resilience (and thus sustainability) of fish populations living in these polluted environments.
We will examine whether exposure of roach populations to oestrogenic WwTW effluents over multiple generations has resulted in genetic selection and the impacts of selection on the susceptibility in male fish to develop oestrogen-induced effects associated with negative fitness consequences. We will examine for selection by identifying functionally significant changes in their genetic make up. Specifically we will look for differences in genetic poylmorphisms (so called single nucleotide polymorphisms -SNPs) in (1) specific (candidate) genes that we know are important in oestrogen signalling of reproductive (and other life) processes and (2) by using methods that allow us to scan the whole genome for these genetic alterations (using a technique called RAD-tag genotyping). Adopting these methods we expect to find 'footprints' of selection in fish living in rivers with a high oestrogenic effluent content, which should allow identification of novel adaptive processes and suggest important mechanisms of toxicity or survival in effluent rich environments. Uniquely, we are able to do this work because we have in depth knowledge (15 years of study) of the wild populations of roach in the selected UK Rivers that we propose to study.
We will establish whether roach populations exposed to high levels of oestrogenic effluent over many generations have adapted to become less responsive to environmental oestrogens and thus are now less susceptible to their associated adverse effects by comparing responses to a controlled exposure to the contraceptive oestrogen, ethinyloestradiol. We will use roach collected from the same clean and WwTW effluent contaminated sites as for the population genetic analyses described above and fish will be exposed for a period of 1 year and effects quantified on responses that have negative fitness consequences in male fish, including ovotestis.
This work is intended to greatly improve understanding of the ability of fish to adapt to exposure to environmental pollutants (here oestrogen) and how they do so, in turn helping to inform on fish population resilience in UK rivers receiving WwTW effluent discharges. The work will have importance in the regulation of discharges for the better protection of our aquatic resources and biodiversity and is of very wide interest to the government regulatory bodies, environment protection groups, industry and the wider public.

Endocrine disrupting chemicals (EDCs) potentially have very wide-reaching deleterious environmental impacts and our findings will be of considerable interest (inter)nationally to academia, pharmaceutical and chemical industries, government regulatory bodies, environment protection groups, and the wider public. The major part of our impact activities will be centred on training, engagement with industry partners and (inter)national government regulatory bodies, and informing the wider public.

The approach we propose in this proposal will apply advanced molecular techniques to provide an advanced understanding on the ability of fish to adapt to oestrogenic EDCs, how they do this and the fitness consequences of these adaptations. We expect the work to benefit, both nationally and internationally industry and government regulatory bodies, by supporting evidence-based decision-making on EDCs, a large group of academic researchers working in similar or related fields and by advancing the use of cutting edge molecular and genome methods and understanding adaptive (evolutionary) processes; the water and chemical (including pharmaceutical) industries through better understanding the implications of exposure to EDCs on wildlife health allowing industry to consider remediation strategy options, including additional treatment technologies; fisheries managers, as widespread adaptation would influence management of fish using hatchery stock; NGOs and the public, who will benefit from better evidence of EDC effects on wildlife and that may also affect their own health.

At this time we do not see that this project has any IP with immediate industrial application. If such an opportunity arises however (e.g. in the discovery of molecular biomarkers) we will gain direct support on commercialisation opportunities and be provided with advice on the protection of IP through our University's Research & Knowledge Transfer sections.

The project team has a strong record in public engagement and outreach activities and will make active use of our respective university press offices to maximise publicity opportunities to ensure the results of our work are widely disseminated. The PIs and RCo-I will also participate in a number of public events and scientific discussions on environmental pollution and these will include; the annual National Science and Engineering Week at Exeter, Cafe Scientifique at Exeter, presentations at national and regional levels to the Association of River Trusts, the Environment Agency Fisheries Teams, Natural England and the European Commission. Both PIs participate actively in a series of school-focused events and in higher education programmes at 'Public Open Days' and take on work experience students in their laboratories.

Both RCo-I and RA recruited to this project will be expected to contribute substantially to the communication activities. The RCo-I and RA will attend a NERC training course on science communication. We will ensure that all personnel recruited to this projected are provided with the opportunity to develop their awareness of, and skills in, science communication and knowledge transfer by; a) designating them major roles in the organization and running of the proposed SETAC and Exeter workshops (including as a presenter); b) encouraging the RCo-I and RA to author articles for the popular press and to engage in public discussion related events outlined above; c) working with university publicity team to design project website and briefing leaflets.

Key measures of success will be in the delivery of high quality science with high impact, as evidenced by publication of the work in the best peer reviewed scientific journals, workshop participation and delivery, disseminations at internationally leading events (scientific, government, regulatory), and uptake of this work into these sectors and influence on policy development for EDCs.