Chikungunya virus persists in joint-associated macrophages and promotes chronic disease.
Arthritogenic alphaviruses including chikungunya, Mayaro, and Ross River viruses cause long-lasting musculoskeletal pain and inflammation. However, mechanisms driving chronic disease remain poorly understood. Here, we investigated joint-associated tissues in alphavirus-infected mice at a late stage of infection. Utilizing scRNA-seq, spatial transcriptomics, and flow cytometry we identified an accumulation of inflammatory macrophages in joint-associated tissues with elevated Tnf, Nlrp3, Il1b, and H2-Aa expression, and these cells harbored CHIKV RNA. Moreover, we identified accumulation of CD4+ T cells in joint-associated tissues, which express Ifng. Depletion of CD4+ T cells diminished MHC-II expression on joint macrophages, highlighting their potential role in inflammation. In addition, treatment with a small molecule inhibitor of CHIKV replication during chronic disease reduced viral RNA and joint inflammation, suggesting that viral RNA replication promotes chronic joint disease. Our data suggest that macrophages harbor replicating viral RNA and contribute to the sustained joint inflammation associated with chronic alphavirus disease.