Transcriptomics Points-of-Departure (tPODs) to Support Hazard Assessment of Benzo[ a]pyrene in Early-Life-Stage Rainbow Trout

Abstract

New approach methods (NAMs) are urgently needed to address the significant ethical and economic concerns associated with live animal testing as well as the low throughput associated with current toxicity testing frameworks. NAMs such as rapid mechanistic early-life-stage fish assays are promising alternatives to current hazard assessment approaches, as they can be used to derive toxicity thresholds and guide decision-makers on identifying or prioritizing chemicals of concern. This study aimed to derive benchmark concentrations from RNaseq data (transcriptomic points-of-departure; tPOD) from a short-term exposure study with early life stages of rainbow trout (RBT; Oncorhynchus mykiss) using benzo[a]pyrene (B[a]P) as the model compound. tPODs were then calibrated with higher organizational-level responses observed during an extended 28 day exposure period. RBT were exposed from 1 to 28 days post-hatch (dph) to 0.079, 0.35, 1.5, 7.4, and 29 μg/L (28 d time weighted average measured) B[a]P, as well as 0.05% dimethyl sulfoxide and water only controls. Benchmark concentration analysis of transcriptomic responses at 4 dph, based on the most sensitive transcriptomic features, yielded tPODs between 0.028 and 0.47 μg/L B[a]P. At 28 dph, Cyp1a1 exhibited significantly increased catalytic activity, with biochemical POD, bPOD_EROD,28dph of 0.599 μg/L B[a]P, while morphometric analysis showed significant growth inhibition in terms of length, with apical POD, aPOD_length,28dph of 1.77 μg/L B[a]P, with a notable decreasing trend in body weight. A toxicity pathway model constructed from genes and apical end points exhibiting concentration-dependent responses provided further evidence supporting the utility of tPODs from short-term RBT early-life-stage assay to support chemical risk assessment to guide decision-makers in chemical testing prioritization.

Keywords: RNAseq; apical responses; benchmark concentration (BMC); chemical risk assessment; fish; new approach methods (NAMs); toxicity pathway.