PROJECT SUMMARY Chikungunya virus (CHIKV) is a mosquito-transmitted, positive-strand enveloped alphavirus that causes global disease in humans. At present, no antiviral agents or licensed vaccines exist for the treatment or prevention of any alphavirus infections. While age, immune status, and pre-existing chronic illness are associated with increased risk of severe CHIKV infection, the role of acquired factors in disease progression is poorly understood. Our preliminary data suggests that the microbiota regulates CHIKV infection, dissemination, and musculoskeletal inflammation and disease through a previously undefined axis by which signals from gut bacteria and bile acids instruct innate immune cell responses to control CHIKV infection of monocytes in circulation and monocyte migration to affected joint tissues. We hypothesize that specific gut bacteria and their microbial constituents modulate CHIKV pathogenesis by regulating antiviral type I IFN and inflammatory responses in pDCs and monocytes. In the absence of these microbial signals, CHIKV disseminates widely, and arthritis ensues after joint infiltration by immune cells. This proposal combines investigators with expertise in alphavirus pathogenesis and immunity (Diamond) and the study of the gut microbiota in disease (Handley, Stappenbeck, and Fischbach). Using a suite of transgenic mice and microbiome reconstitution experiments paired with detailed virological and immunological analyses, we will address the following key questions: (a) which immune cells coordinate the rapid systemic IFN response following CHIKV infection (e.g., pDCs) (b) what immune cues limit viral infection in circulating immune cells (e.g., monocytes)? (c) how does the gut microbiota regulate pDC IFN production and trafficking of circulating immune cells? and (d) which constituents (e.g., metabolites) of the microbiota regulate antiviral and inflammatory responses? Through these detailed mechanistic studies, we expect to link the microbe-derived constituents of specific commensal bacteria with innate antiviral responses that modulate alphavirus infection, dissemination, joint disease, and possibly transmission. Beyond enhancing our understanding of acquired determinants of alphavirus pathogenesis, the findings of this proposal could inform more generally our understanding of how the gut microbiota shapes innate immune responses to limit infection and pathogenesis of other viruses.