Review of ecologically relevant in vitro bioassays to supplement current in vivo tests for whole effluent toxicity testing - Part 1: Apical endpoints.

Whole effluent toxicity (WET) testing is commonly used to ensure that wastewater discharges do not pose an unacceptable risk to receiving environments. Traditional WET testing involves exposing animals to (waste)water samples to assess four major ecologically relevant apical endpoints: mortality, growth, development, and reproduction. Recently, with the widespread implementation of the 3Rs to replace, reduce and refine the use of animals in research and testing, there has been a global shift away from in vivo testing towards in vitro alternatives. However, prior to the inclusion of in vitro bioassays in regulatory frameworks, it is critical to establish their ecological relevance and technical suitability. This is part 1 of a two-part review that aims to identify in vitro bioassays that can be used in WET testing and relate them to ecologically relevant endpoints through toxicity pathways, providing the reader with a high-level overview of current capabilities. Part 1 of this review focuses on four apical endpoints currently included in WET testing: mortality, growth, development, and reproduction. For each endpoint, the link between responses at the molecular or cellular level, that can be measured in vitro, and the adverse outcome at the organism level were established through simplified toxicity pathways. Additionally, literature from 2015 to 2020 on the use of in vitro bioassays for water quality assessments was reviewed to identify a list of suitable bioassays for each endpoint. This review will enable the prioritization of relevant endpoints and bioassays for incorporation into WET testing.