Green autofluorescence eleocytes from earthworm as a tool for detecting environmental iron pollution

Abstract The riboflavin content of eleocytes is involved in immune responses in earthworms and serves as a sensitive bioindicator of soil condition, such as metal ions pollution. Previous studies have shown that the transition metals Ni and Cu robustly decrease riboflavin content of eleocytes when earthworms were exposed to transition metals-polluted soil. However, there is no previous study of earthworm responses to environmental stressors associated with the transition metal Fe. Therefore, this study aimed to uncover the effect of Fe exposure on the riboflavin content of eleocytes from Eisenia fetida (E. fetida) exposed to 1 mM FeCl3 for 4 days using agar gels as exposure media, and riboflavin exposure as a positive control. In this study, we found that two cell types exist in the coelomic fluid of E. fetida. One cell type is the amoebocytes (about 70%), and the other is riboflavin-storing eleocytes (about 20%). Interestingly, eleocytes can spontaneously emit green autofluorescence, and the mean fluorescence intensity (MFI) and percentages of this cell population were decreased when exposed to FeCl3 in vitro. These findings were also confirmed by in vivo experiments. However, the riboflavin content of celomic fluid supernatants and total coelomocyte numbers were unaffected. These results indicate that Fe3+ can potentially quench the fluorescence of riboflavin in eleocytes. Therefore, by detecting the number and MFI of eleocytes, we provide a novel link between Fe3+ and the green autofluorescent cells of earthworms, which can be used as a novel tool for detecting the iron pollution in soil and the environment.