RESUMO
Embryonic zebrafish represent a useful test system to screen substances for their ability to perturb development. The exposure scenarios, endpoints captured, and data analysis vary among the laboratories who conduct screening. A lack of harmonization impedes the comparison of the substance potency and toxicity outcomes across laboratories and may hinder the broader adoption of this model for regulatory use. The Systematic Evaluation of the Application of Zebrafish in Toxicology (SEAZIT) initiative was developed to investigate the sources of variability in toxicity testing. This initiative involved an interlaboratory study to determine whether experimental parameters altered the developmental toxicity of a set of 42 substances (3 tested in duplicate) in three diverse laboratories. An initial dose-range-finding study using in-house protocols was followed by a definitive study using four experimental conditions: chorion-on and chorion-off using both static and static renewal exposures. We observed reasonable agreement across the three laboratories as 33 of 42 test substances (78.6%) had the same activity call. However, the differences in potency seen using variable in-house protocols emphasizes the importance of harmonization of the exposure variables under evaluation in the second phase of this study. The outcome of the Def will facilitate future practical discussions on harmonization within the zebrafish research community.
RESUMO
The embryonic zebrafish is a useful vertebrate model for assessing the effects of substances on growth and development. However, cross-laboratory developmental toxicity outcomes can vary and reported developmental defects in zebrafish may not be directly comparable between laboratories. To address these limitations for gaining broader adoption of the zebrafish model for toxicological screening, we established the Systematic Evaluation of the Application of Zebrafish in Toxicology (SEAZIT) program to investigate how experimental protocol differences can influence chemical-mediated effects on developmental toxicity (i.e., mortality and the incidence of altered phenotypes). As part of SEAZIT, three laboratories were provided a common and blinded dataset (42 substances) to evaluate substance-mediated effects on developmental toxicity in the embryonic zebrafish model. To facilitate cross-laboratory comparisons, all the raw experimental data were collected, stored in a relational database, and analyzed with a uniform data analysis pipeline. Due to variances in laboratory-specific terminology for altered phenotypes, we utilized ontology terms available from the Ontology Lookup Service (OLS) for Zebrafish Phenotype to enable additional cross-laboratory comparisons. In this manuscript, we utilized data from the first phase of screening (dose range finding, DRF) to highlight the methodology associated with the development of the database and data analysis pipeline, as well as zebrafish phenotype ontology mapping.
RESUMO
There is a need for fast, efficient, and cost-effective hazard identification and characterization of chemical hazards. This need is generating increased interest in the use of zebrafish embryos as both a screening tool and an alternative to mammalian test methods. A Collaborative Workshop on Aquatic Models and 21st Century Toxicology identified the lack of appropriate and consistent testing protocols as a challenge to the broader application of the zebrafish embryo model. The National Toxicology Program established the Systematic Evaluation of the Application of Zebrafish in Toxicology (SEAZIT) initiative to address the lack of consistent testing guidelines and identify sources of variability for zebrafish-based assays. This report summarizes initial SEAZIT information-gathering efforts. Investigators in academic, government, and industry laboratories that routinely use zebrafish embryos for chemical toxicity testing were asked about their husbandry practices and standard protocols. Information was collected about protocol components including zebrafish strains, feed, system water, disease surveillance, embryo exposure conditions, and endpoints. Literature was reviewed to assess issues raised by the investigators. Interviews revealed substantial variability across design parameters, data collected, and analysis procedures. The presence of the chorion and renewal of exposure media (static versus static-renewal) were identified as design parameters that could potentially influence study outcomes and should be investigated further with studies to determine chemical uptake from treatment solution into embryos. The information gathered in this effort provides a basis for future SEAZIT activities to promote more consistent practices among researchers using zebrafish embryos for toxicity evaluation.
