Cryptic speciation and blurred species boundaries of the earthworm: A challenge for soil-based toxicological risk assessments.

The toxicological risk assessment of chemicals is largely based on the Organization for Economic Co-operation and Development (OECD) guidelines. These internationally approved methodologies help shape policy and political strategy of environment and human health issues. Risk assessments which pertain to soil biota 'recruit' sentinel organisms, including the earthworm Eisenia fetida. Despite E. fetida being morphologically similar to Dendrobaena veneta, they are characterized by a several-fold difference in sensitivity to xenobiotics. Worms, sold as either as pure E. fetida stocks or E. fetida/D. veneta mixed cultures, were obtained from five commercial suppliers. The species identity of 25 earthworms was determined by sequencing the cytochrome c oxidase subunit 2 (COII). We revealed that only one of 25 worms was E. fetida, the remaining worms were all identified as D. veneta. This underlines the notion that E. fetida and D. veneta are easily mis-identified. The occurrence of cryptic speciation combined with the well-documented species-specific variation in toxicological responses highlights the pressing need to accurately classify earthworms to species level prior to any toxicological research. Only this will ensure the validity and reliability of risk assessments.

Polyester-derived microfibre impacts on the soil-dwelling earthworm Lumbricus terrestris.

Microplastic (MP) pollution is everywhere. In terrestrial environments, microfibres (MFs) generated from textile laundering are believed to form a significant component of MPs entering soils, mainly through sewage sludge and compost applications. The aim of this study was to assess the effect of MFs on a keystone soil organism. We exposed the earthworm Lumbricus terrestris to soil with polyester MFs incorporated at rates of 0, 0.1 and 1.0 %w/w MF for a period of 35 days (in the dark at 15 °C; n = 4 for each treatment). Dried plant litter was applied at the soil surface as a food source for the earthworms. We assessed earthworm vitality through mortality, weight change, depurate production and MF avoidance testing. In addition, we measured stress biomarker responses via the expression of metallothionein-2 (mt-2), heat shock protein (hsp70) and superoxide dismutase (sod-1). Our results showed that exposure and ingestion of MFs (as evidenced by subsequent retrieval of MFs within earthworm depurates) were not lethal to earthworms, nor did earthworms actively avoid MFs. However, earthworms in the MF1.0% treatment showed a 1.5-fold lower cast production, a 24.3-fold increase in expression of mt-2 (p < 0.001) and a 9.9-fold decline in hsp70 expression (p < 0.001). Further analysis of soil and MF samples indicated that metal content was not a contributor to the biomarker results. Given that burrowing and feeding behaviour, as well as molecular genetic biomarkers, were modulated in earthworms exposed to MFs, our study highlights potential implications for soil ecosystem processes due to MF contamination.