Metabolic disruption in salmonids following co-exposure to road runoff contaminants 6PPD-Quinone and 9,10-anthraquinone.

Components of road runoff have been correlated to adverse health outcomes in fish and aquatic invertebrates residing in urban water bodies. Recently, a transformation product of the antioxidant chemical added to car tires, N-(1,3-Dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD-quinone, 6PPD-q), was found to induce urban stormwater mortality syndrome (URMS) in salmonids. Exposure to 6PPD-q has the potential to increase the sensitivity of salmonids to other co-occurring contaminants commonly detected in surface waters, such as the polycyclic aromatic hydrocarbon 9,10-anthraquinone (AQ). To investigate the mechanisms of 6PPD-q toxicity, fish were exposed to established sublethal concentrations of 6PPD-q and AQ, both separately and in combination, over a five-day period. Non-targeted MS-based metabolomics analysis was conducted on liver samples. Using in-house and open-source spectral libraries, 260 metabolites were confidently annotated in liver tissues. This study provides mechanistic insights into 6PPD-q and AQ toxicity in salmonids, highlighting oxidative stress, mitochondrial dysfunction, and impaired detoxification as key drivers of sublethal effects. Findings also suggest that AQ may mitigate 6PPD-q toxicity by enhancing antioxidant defenses and xenobiotic metabolism. Additionally, quantitative RNA sequencing of liver tissues revealed species-specific transcriptomic responses to chemical exposures, with coho salmon exhibiting more pronounced differential gene expression relative to chinook and rainbow trout. Transcriptomic analysis identified key modulation of genes involved in apoptosis, immune response, vascular permeability, and energy metabolism, deepening mechanistic understanding of 6PPD-q toxicity. SYNOPSIS: This study reveals species-specific 6PPD-q toxicity mechanisms and highlights co-exposure effects, informing risk assessments for road runoff-derived pollutant mixtures.