It has not been known how immune responses in soil invertebrates occur against microplastics (MPs). This study aims to investigate the effects of MPs on endocytosis, including phagocytosis and pinocytosis, of immune cells of soil invertebrates in the soil ecosystem in the process of bacterial infection. We employed polystyrene micro- plastics (similar to 1 mu m PS MPs) to treat earthworm Eisenia andrei during the infection of Escherichia coli for in vitro (1, 5, 10, and 50 mg/L) and in vivo (1, 10, and 1000 mg/kg dry soil) assays. The results of in vitro migration assay revealed that MPs caused inhibitory effects on the phagocytosis, pinocytosis and oxidative stress in coelomocytes. Soil bioassay also confirmed that endocytosis of coelomocytes and mitochondrial damages in the intestinal epithelium were significantly altered in the polluted soil with MPs. Thus, MPs induced adverse effects to inhibit bacterial endocytosis, which may disturb the immune system of soil invertebrates. This study is the first report on the inhibition of phagocytosis in the soil invertebrates by MPs. These findings contribute to understanding the response of soil invertebrates, which play important roles in the soil food web with cellular level towards microplastic pollution in soil.

Nanoplastics (NPs) are currently everywhere and environmental pollution by NPs is a pressing global problem. Nevertheless, until now, few studies have concentrated on the mechanisms and pathways of cytotoxic effects and immune dysfunction of NPs on soil organisms employing a multidimensional strategy. Hence, earthworm immune cells and immunity protein lysozyme (LZM) were selected as specific receptors to uncover the underlying mechanisms of cytotoxicity, genotoxicity, and immunotoxicity resulting from exposure to polystyrene nanoplastics (PS-NPs), and the binding mechanisms of PS-NPs-LZM interaction. Results on cells indicated that when earthworm immune cells were exposed to high-dose PS-NPs, it caused a notable rise in the release of reactive oxygen species (ROS), resulting in oxidative stress. PS-NPs exposure significantly decreased the cell viability of earthworm immune cells, inducing cytotoxicity through ROS-mediated oxidative stress pathway, and oxidative injury effects, including reduced antioxidant defenses, lipid peroxidation, DNA damage, and protein oxidation. Moreover, PS-NPs stress inhibited the intracellular LZM activity in immune cells, resulting in impaired immune function and immunotoxicity by activating the oxidative stress pathway mediated by ROS. The results from molecular studies revealed that PS-NPs binding destroyed the LZM structure and conformation, including