Sublethal effects of Cu and Cr on Eisenia andrei: Life-cycle endpoints and biomarker integration for soil assessment

Soil contamination by heavy metals poses profound ecological challenges owing to their persistent nature and inherent toxicity. This study aims to assess sublethal effects of copper (Cu) and chromium (Cr) on earthworm Eisenia andrei by integrating life-cycle endpoints and biomarkers to develop an early-warning framework for soil contamination and ecological risk assessment. Natural soils were spiked with increasing concentrations of Cu (0 -1600 mg/kg) and Cr (0-200 mg/kg) and organismal responses were evaluated by assessing survival, growth (after 10 or 28 days) and reproduction (after 56 days), and a suite of physiological and biochemical biomarkers, including membrane stability (LMS), accumulation of lipofuscin (LF) and unsaturated neutral lipids (NL), synthesis of metallothioneins, and activities of Ca²⁺-ATPase (CaATP), glutathione S-transferase, catalase, and acetyl-CoA oxidase (AOX) after 10 days. The reproduction endpoint emerged as the most sensitive indicator of toxicity, with estimated effective concentrations (EC50) of 355 mg/kg for Cu and 150 mg/kg for Cr. Conversely, effects on survival and growth were primarily observed at the highest concentrations. Among biomarkers, LMS and LF were the most sensitive parameters, with EC50 values of 157 and 142 mg/kg, respectively. For Cr, AOX was the most sensitive biomarker (EC50 of 131 mg/kg), followed by LMS. For Cr, AOX was the most sensitive biomarker (EC50 of 131 mg/kg), followed by LMS. The findings highlight that the selected biomarkers can detect sublethal stress from metal contamination prior to observable effects on life-cycle parameters. This study underscores the usefulness of an integrated approach for early detection of soil contamination and proactive ecological management and preservation of soil health.