Reflecting on 20 years of OECD 225: Advancing standardised ecotoxicological testing with Lumbriculus variegatus

Sediments are key environmental compartments that accumulate and release chemical contaminants, leading to chronic exposure of benthic communities. The freshwater oligochaete Lumbriculus variegatus is the only sediment-dwelling species recommended for sediment risk assessment under OECD guidelines, but it is used far less frequently than Chironomus sp. To analyse why it is underused, we systematically reviewed 303 ecotoxicological studies with L. variegatus published since the development of OECD Guideline 225 in 2007, identifying 91 studies that met predefined inclusion criteria and methodological quality standards. Only 12 % of chronic sediment studies (≥28 days) strictly followed OECD 225 without deviations, while 51 % did not refer to the guideline at all. Conventional endpoints (mortality, growth, reproduction) were assessed in 91 % of studies. However, life-stage differentiation was rarely applied, and 27 % of chronic studies failed to meet the reproduction criterion in controls. Overall, 44 % of studies extended OECD 225 by including additional endpoints. Behavioural endpoints (feeding, locomotion, sediment avoidance, bioturbation) were used in 30 % of studies and were frequently more sensitive than conventional measures. Thirty-four comparative studies between L. variegatus and at least one other standard sediment species (Chironomus sp., Hyalella azteca, or Tubifex tubifex) showed that none of the species was generally much more sensitive than the oligochaete. Our review identifies substantial gaps in standardisation, endpoint selection, and experimental design in L. variegatus studies. We recommend revising OECD 225 to incorporate additional sublethal endpoints, update the reproduction criterion and synchronisation time, and differentiate life stages in growth and reproduction assessments. These improvements would enhance the ecological relevance and regulatory applicability of L. variegatus in sediment ecotoxicology.