Toxicity and Behavioral Responses of Environmentally Relevant Microplastics in Earthworms ( <i>Eisenia fetida</i> ): Ingestion, Survival, Reproduction, and Avoidance Across Common Polymer Types

Microplastics (MPs) are recognized as ubiquitous contaminants in terrestrial ecosystems, yet their ecological risks to soil invertebrates remain poorly categorized, particularly at environmentally relevant exposure levels. This study investigated avoidance behaviour, ingestion, and toxicity of four common MP polymers (polyethylene; PE, polypropylene; PP, polyester/poly(ethylene terephthalate); PET, and polyacrylamide; PAM) in the earthworm Eisenia fetida using particle-based exposure concentrations. Standardized avoidance, reproduction, and ingestion assays were conducted across a wide concentration range (1,000-1,000,000 MP/kg dry soil) under controlled laboratory conditions. Avoidance responses were weak and highly variable across all polymers, with no consistent concentration-dependent patterns observed. Survival exceeded 90% in all treatments, and reproductive output showed no clear concentration-response relationship, with juvenile production at the highest exposure levels not differing from controls. MP ingestion was minimal at lower exposure concentrations, with sporadic detections occurring across polymers, but MPs across the polymers were detectable above similar threshold concentrations (∼80,800-92,800 MP/kg). Above these thresholds, ingestion increased predictably with log-transformed exposure concentration, showing comparable post-threshold ingestion patterns across polymers. Estimated bioaccumulation factors (BAFs), calculated at a standardized exposure concentration of 1,000,000 MP/kg, were low (0.0042-0.0124), indicating limited internal accumulation of MPs over the 28-day exposure period. Collectively, these results suggest that Eisenia fetida does not actively avoid MP-contaminated soils and does not retain MPs to a significant extent under the conditions tested. By integrating behavioural, ingestion, and toxicity endpoints and reporting exposure in particle-based units, this study provides ecologically relevant insights into MP-earthworm interactions and contributes to improving terrestrial MP risk assessment frameworks.