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Development of Bioanalytical Techniques to Assess the Potential Human Health Impacts of Recycled Water

Project: 10-07
Year Released: 2014
Type: Report

Program: Principal
Funding Partner: Bureau of Reclamation
Total Investment: $839,446 ($249,789 Cash; $589,657 In-kind)

Principal Investigator: Beate Escher, The University of Queensland (UQ)−National Research Center for Environmental Toxicology (Entox)


Bioanalytical tools are cell-based bioassays that give a measure of the toxicity and presence of known and unknown chemicals in complex environmental samples. Improved detection of bioactive chemicals in water will improve risk assessment methods and inform future water management options, particularly in the context of water reclaimed from impaired sources such as wastewater or stormwater.

Goals and Objectives

The project advances the science and application of bioanalytical tools for water quality assessment. The specific project objectives were (A) to review literature and give an overview on bioanalytical tools that can assess likely impacts on human health, (B) to validate novel endpoints and implement them together with existing bioassays for benchmarking water quality of recycled water, and (C) to develop interpretation guidelines.

Research Approach

Task 1: Literature Review. The past years have seen increased application of bioanalytical tools for water quality assessment of wastewater, recycled water and surface waters. As a first step we will compile activities and recent applications in a global context. This will allow pre-selection of bioassays that have already been validated or partially validated for application in water quality assessment based on mode of action. These are defined as category 1 bioassays.

An additional and unique feature of the literature review is that it will involve evaluation of the application of bioanalytical tools of relevance to human health. This will lead to identification of a suite of category 2 bioassays, i.e., those that target a relevant mode of toxic action and that have been validated with single chemicals, for example during the Tox21 program (Gibb, 2008) but that (a) have not yet been adapted to water quality assessment and/or (b) lack a TEQ concept but appear promising after the literature review.

Task 2: Validation Plan. For validation, bioanalytical tools will be selected that were deemed as potentially suitable by the literature review and that have been previously applied and established by the core partner laboratories and/or associated laboratories. In addition new endpoints/bioassays that have been identified as category 2 bioassays during the literature review will be implemented by the core laboratories for comparison with existing established bioassays, if required. The suitability of category 2 bioassays to be included into the battery will be assessed with experiments using defined positive and negative controls and with the same samples to which category 1 bioassays were applied, but no wider inter-laboratory comparison will be performed with the category 2 bioassays within the current project. If several bioassays based on the same endpoint (covering a relevant mode of toxic action) are identified and it is not clear what bioassay is best suited to assess this endpoint, different bioassays covering the same mode of action but different stages of the toxicity pathway or involving different visualisations of the effect (e.g., gene reporter assays with different reporter genes) will be included. The core of the validation plan is an inter-laboratory comparison to assure repeatability, robustness and reliability of responses.

Task 3: Interpretation Guidelines. Difficulties in the interpretation of results are an obstacle to the acceptance of bioanalytical tools for monitoring treatment efficiency and assessment of water quality. Responses are usually reported as % effect at a given sample dilution/enrichment or as an effect concentration eliciting a defined endpoint (e.g., 50% inhibition of enzyme activity or exceedance of a defined effect threshold). This type of information can be very confusing for people who are not very familiar with bioassays and dose response assessment. Therefore, we seek to implement a uniform and easily followed way to interpret the toxicity measurements and to express the results. The observed effect in every selected bioassay will be expressed as toxic equivalent concentrations (TEQ) relative to an appropriate reference compound. No qualitative assays will be selected that do not allow dose-response assessment or translation of results into TEQ, as this would preclude derivation of log removal using bioassays. The existence of a TEQ concept will be a selection criterion for choosing a bioassay or will be one of the first steps to be addressed during the implementation of category 2 bioassays. Bioassays respond to mixture of chemicals with a common mode of toxic action. Therefore, a one-to-one translation from chemical guideline values to bioassay based guideline values for all compounds may not be possible. Instead we seek to make initial steps towards establishment of bioassay based trigger values. Exceedance of these trigger values will indicate that a more detailed analysis, including chemical analysis or more definitive toxicity assessment, is warranted.

Findings and Conclusions

All goals of this project were achieved and the results of the study are a major leap toward the implementation and acceptance of bioanalytical tools to assess the quality of recycled water. The purpose of this study was not to investigate the occurrence of bioactivity at a large number of facilities but rather to demonstrate the proficiency and reproducibility of a battery of test assays that could be employed in reuse scenarios to screen complex mixtures. Thus, the most important aspect of the study was demonstrating that the assays were effective in screening both highly challenged waters (i.e., secondary effluents from WWTPs) as well as highly treated waters (i.e., post-reverse osmosis waters).

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