Single-cell characterization of skin response to a bite by West Nile virus-infected mosquito reveals fibroblast-mediated barrier to transmission

ABSTRACT Cutaneous events at the mosquito bite site that determine orthoflavivirus transmission efficiency remain largely uncharacterized. Here, we report single-cell RNA-sequencing of skin from immunocompetent mice bitten by West Nile virus-infected mosquitoes, capturing early response at a critical transmission bottleneck. Fibroblasts were the dominant cells exposed to infectious saliva. While neutrophils and Folr2+ macrophages diminished, monocyte-derived and antigen-presenting macrophages, and lymphocytes became more abundant. Cell-cell communication analysis revealed the central roles of fibroblasts and myeloid cells in induced immune-related signaling. Transcriptional profiling defined cell type-specific responses to infectious bite integrating immune modulation, skin repair and metabolic remodeling. Using in vivo gene silencing, we demonstrated that fibroblast-expressed LRRC15 (leucine rich repeat-containing 15) functions as a cutaneous restriction factor, limiting viral replication in skin, viral dissemination to draining lymph nodes, and disease severity. Collectively, our analyses provide cellular and molecular understanding of bite-initiated arboviral transmission, establishing skin-resident fibroblasts as frontline defender cells. HIGHLIGHTS Single-cell profiling captures early cutaneous response to West Nile virus-infected mosquito bites with cell type resolution. Skin-resident fibroblasts are the dominant frontline cells exposed to infectious mosquito saliva. Infectious bite reconfigures cutaneous immune cells, and activates multi-directional signaling between immune and structural skin cell types. Bite-induced fibroblast-expressed LRRC15 restricts viral transmission and attenuates disease severity. Competing Interest Statement The authors have declared no competing interest. Footnotes The revised version include an improvement in the annotation of myeloid cells.