Single-cell RNA sequencing reveals the landscape of immune cells and the characterization of viral tropism in Japanese encephalitis virus-infected mouse brain

JEV is a neurotropic pathogen that causes lethal encephalitis in humans. The high susceptibility and massive proliferation of JEV in neurons lead to neuronal damage and dramatical inflammation in central nerves system (CNS). However, the mechanisms by which JEV preferentially infects neurons and induces massive neuroinflammation are largely unknown. Here, we found that JEV mainly distributed in cerebral cortex, striatum and thalamus in brain of infected mice, indicating the tissue or cell type bias. Cells in these regions of JEV-infected mice showing different symptoms (mild, moderate and severe) were isolated and subjected to scRNA-seq. 88000 single cells were obtained based on top differentially expressed genes, and 34 clusters and 10 major cell types were identified. As demonstrated by scRNA-seq and flow-cytometry assay, activated microglia cells and infiltrating immune cells including monocyte & macrophage, T cells and NK cells which are responsible for inflammatory response were increased in JEV-infected brain in a symptom severity-dependent manner. Furthermore, the subclusters of these cells and the cell-cell interaction network were analyzed. More communications between individual cells and significant increase in ligand receptor pairs associated with tight junctions, chemokines and antigen-presenting molecules were shown in JEV-infected mouse brain, suggesting upregulation of cellular permeability, inflammation and antiviral response. To identify which subtype(s) of neurons is/are targeted by JEV, 27874 neuronal cells were reclustered to 15 subsets. By analyzing the correlation between the expression of JEV E gene and neuronal genes, we found that Baiap2high neurons are highly permissive to JEV.