Extracellular traps release by coelomocytes in Eisenia andrei earthworm are involved in encapsulation process

Earthworms have several strategies for pathogen elimination including: cytotoxicity, phagocytosis, production of reactive oxygen species and formation of extracellular traps (ETs). In turn, an effective response against larger pathogens, such as parasites, is based on the encapsulation followed by melanization and formation of brown bodies. In our work, we elucidated interactions between ETs and encapsulation. We found that such interactions are time-dependent and involved humoral components of the coelomic fluids. Moreover, it was observed that in vitro encapsulation of agarose beads was much more effective when beads were coated with lipopolysaccharide (LPS) and when process took place in the presence of the coelomic fluid humoral components. Their presence was also crucial for ETs formation. Moreover, we found that protease inhibitors reduced agarose bead-induced encapsulation and ETs formation and that presence of humoral components of coelomic fluid and ETs enhanced encapsulation of the beads and the melanization processes. In contrast, induced by LPS-bead ET formation was not dependent on reactive oxygen species (ROS). Moreover, we discovered that in the first steps of encapsulation process, extracellular DNA release may help in the formation of the brown bodies. Therefore, it could be summarized that earthworms utilize a combination of cytotoxicity, phagocytosis, ROS production, and ET formation for pathogen elimination. The encapsulation process, enhanced by humoral components and ETs, plays a significant role in their immune response, particularly against large pathogens.