Advancing the Effect-Directed Identification in Combined Pollution: Using Pathways to Link Effects and Toxicants

The difficulty in associating diverse pollutants with mixture effects has led to significant challenges in identifying toxicants in combined pollution. In this study, pathways were used to link effects and toxicants. By pathways evaluated by the concentration-dependent transcriptome, individual effects were extended to molecular mechanisms encompassing 135 pathways corresponding to 6 biological processes. Accordingly, mechanism-based identification of toxicants was achieved by constructing a pathway toxicant database containing 2413 chemical–pathway interactions and identifying pathway active fragments of 72 pathways. The developed method was applied to two different wastewaters, industrial wastewater OB and municipal wastewater HL. Although lethality and teratogenesis were both observed at the individual level, different molecular mechanisms were revealed by pathways, with cardiotoxicity- and genotoxicity-related pathways significantly enriched in OB, and neurotoxicity- and environmental information processing-related pathways significantly enriched in HL. Further suspect and nontargeted screening generated 59 and 86 causative toxicants in OB and HL, respectively, among which 29 toxicants were confirmed, that interacted with over 90% of enriched pathways and contributed over 50% of individual effects. After upgrading treatments based on causative toxicants, consistent removal of toxicants, pathway effects, and individual effects were observed. Mediation by pathways enables mechanism-based identification, supporting the assessment and management of combined pollution.