Single-cell transcriptomic profiling of the splenic and peripheral immune landscape in rhesus macaques during CHIKV infection.

Chikungunya virus (CHIKV) is a re-emerging arbovirus causing acute febrile illness and chronic debilitating arthritis, thereby imposing significant global public health and economic burdens. While robust immune responses involving inflammatory cytokines and immune cell infiltration characterize acute infection, the cellular mechanisms underlying pathogenesis and chronicity remain incompletely defined. In this study, we employed single-cell RNA sequencing (scRNA-seq) to comprehensively profile splenic and peripheral blood mononuclear cells (PBMCs) immune responses in rhesus macaques at day 7 post CHIKV infection, an acute phase. Splenic neutrophils marked recruitment, upregulation of S100A8/S100A9, downregulation of interferon-stimulated genes (ISGs), and functional activation marked by degranulation, enhanced anti-apoptotic pathways, and neutrophil extracellular traps (NETs) formation, as visualized by multiplex immunofluorescence. Pseudotime trajectories delineated progressive states transitioning from proliferation to effector functions. Splenic T and B cells showed increased abundance with innate-to-adaptive transition signature. Splenic CD4+ and CD8+ T cell subsets and B cell subsets exhibited enrichment for innate immune pathways and translational machinery. In PBMCs, CD8+ T cell subsets and B cell subsets exhibited activation of adhesion, cytokine signaling, and protein homeostasis pathways in infection group. Notably, CHIKV infection induced skewing of T cells toward states enriched for Th1 and Th17 differentiation (marked by NXPE3, LEF1, NFKBIZ expression) in PBMCs. These results delineate the spatiotemporal immune landscape during CHIKV acute phase, in this non-human primate model, identifying neutrophil-mediated inflammation, lymphocyte transcriptional adaptation, and Th1/Th17 polarization as hallmarks of acute phase, offering a detailed cellular map that may inform future investigations into CHIKV pathogenesis and therapeutic strategies.