Ecotoxicological risk assessment using a multi-scale approach in the estuarine copepod Eurytemora affinis applied to an ecdysone agonist

Previous transcriptomic analyses have demonstrated that exposure to tebufenozide (TEB) induces significant alterations in gene expression in the crustacean Eurytemora affinis (calanoid copepod; E. affinis), supporting its suitability as a model organism for investigating the ecotoxicological effects of ecdysone agonists in estuarine environments. Based on these findings, we hypothesized that TEB exposure may disrupt biological processes across multiple levels of organization in E. affinis. We conducted an integrated assessment of upstream (epigenetics) and downstream (locomotion, larval development) endpoints through multiple assays. Additionally, we examine the indirect effects of TEB by evaluating the changes in the copepod microbiota. We showed that TEB had no impact on global DNA methylation (HPLC-MS/MS) in both males and females. In contrast, significant effects were observed on the swimming behaviour of males, with copepods exposed to TEB being more affected by the light/dark transition and showing hyperactivity after 4 days. The larval development was also shown to be accelerated by TEB after 4 days. Finally, significant changes in microbiota were highlighted, including a decrease in both alpha and beta diversity indices, along with an increase in the abundance of the Gammaproteobacteria class. The effects observed in E. affinis show that TEB could impact fitness and increase vulnerability of non-target organisms. The strategy of prioritising the responses to be investigated on the basis of gene expression has proven effective. The study reinforces the relevance of using E. affinis for environmental risk assessment in estuaries, as it can be used to assess multiple endpoints at different biological scales.