The project ZET-O-MAP aimed to give zebrafish embryo toxicity testing a mechanistic foundation, turning scattered public data into a knowledge base for regulatory use. It was funded by the European Chemical Industry Council's Long-Range Research Initiative (Cefic-LRI).

Congenital malformations affect around 3% of newborns, but identifying which chemicals disrupt embryonic development remains difficult. The zebrafish embryo teratogenicity assay (ZETA) is a promising alternative to mammalian testing. It is fast, transparent, and vertebrate-based. However, regulators often require mechanistic evidence before accepting such assays. A visible malformation says little about why it occurred. ZET-O-MAP set out to close this gap by linking molecular events to morphological outcomes.

Carried out by Fraunhofer SCAI, Fraunhofer ITEM, and the Flemish Institute for Technological Research (VITO), the project mined morphological findings from published zebrafish tests and compiled transcriptomics datasets from public repositories. These heterogeneous datasets were reprocessed through a single, uniform pipeline, using the R-ODAF framework, which was developed in the sister project LRI-C4 as a standard for regulatory-grade reproducibility. This approach aimed to make studies generated on different platforms genuinely comparable.

SCAI researchers integrated harmonized data into a machine-readable knowledge graph, supported by newly developed ontologies for morphological effects and enriched with time-resolved “spatio-temporal maps” of gene expression. Three test substances were additionally profiled experimentally in concentration series with RNA sequencing at four developmental stages. The resulting knowledge map links adverse outcomes to mechanisms mirroring known mammalian toxicology.

The results are publicly accessible via the ZET-O-MAP Graph Visualizer, an online tool for exploring substance-molecule-effect relationships. By linking molecular drivers to malformations, the project strengthens the case for regulatory acceptance of the ZETA assay, thus helping safer substances reach the market faster, with fewer animals used.

Project duration: March 2021 until February 2023