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

Cutaneous events determining transmission of mosquito-borne orthoflaviviruses remain largely uncharacterized. Here, we report single-cell RNA-sequencing of skin from immunocompetent mice exposed to West Nile virus-infected mosquitoes, capturing early response at a critical transmission bottleneck. Skin-resident fibroblasts, keratinocytes, and myeloid cells were exposed to infectious saliva. At the bite site, neutrophils and mast cells diminished, while lymphoid cells augmented. Cell-cell communication analysis showed that structural skin cells activate immune signaling targeting myeloid populations, which signal to lymphoid cells. Transcriptional profiling revealed cell subtype-specific responses integrating immune activation, skin repair and metabolic remodeling. Using in vivo gene silencing, we demonstrated that fibroblast-expressed LRRC15 (leucine rich repeat-containing 15) functions as a restriction factor, limiting viral replication in skin, viral dissemination to draining lymph nodes, and disease severity. Collectively, our transcriptomic-resolution and functional analyses provide cellular and molecular understanding of bite-initiated arboviral transmission, establishing skin-resident fibroblasts as frontline defender cells. HIGHLIGHTS* Single-cell RNA-sequencing captures early cutaneous response to West Nile virus-infected mosquito bites. * Skin-resident fibroblasts, keratinocytes and myeloid cells are exposed to infectious mosquito saliva. * Infectious bite induces rapid cellular remodeling and activates immune signaling networks from structural cells to myeloid and lymphoid populations. * Bite-induced fibroblast-expressed LRRC15 restricts viral transmission and attenuates disease severity.