Combined toxicity of organophosphate flame retardants and polyethylene microplastics on Eisenia fetida: Biochemical and molecular insights

Microplastics and organophosphate flame retardants (OPFRs), a subset of organophosphate esters (OPEs), are frequently co-detected in terrestrial ecosystems, raising concerns about their combined ecological risks. In this study, the epigeic earthworm Eisenia fetida was exposed to polyethylene microplastics (PEs) (0.5 g kg⁻¹), three chlorinated OPFRs (TCEP (tris(2-chloroethyl) phosphate), TCPP (tris(1-chloro-2-propyl) phosphate), and TDCPP (tris(1,3-dichloro-2-propyl) phosphate), each at 0.002 g kg⁻¹), and their binary mixtures for 7 and 28 days. A suite of physiological, biochemical, and molecular biomarkers, including enzyme activities (SOD, CAT, GST, AChE, ATPases), oxidative damage markers (MDA, 8-OHdG), and gene expressions (sod, hsp70, tctp), were evaluated to assess sublethal toxic responses. A time-dependent shift in the correlation between sod transcription and SOD enzyme activity was observed, indicating possible early-stage post-transcriptional regulation and later-stage transcriptional control. The results revealed compound-specific and time-dependent toxicities. TDCPP exhibited the highest individual toxicity, suppressing antioxidant enzymes and disrupting ion transport. Co-exposure with PEs attenuated TDCPP-induced effects, likely via reduced bioavailability. In contrast, PEs enhanced the toxicity of TCPP, especially in oxidative and genotoxic responses. TCEP induced moderate but delayed biochemical changes. This study underscores the dual role of PEs as both carriers and modulators of co-occurring pollutants, and highlights the need for mixture-based risk assessments in soil ecosystems.