Initial sites of SIV rebound after antiretroviral treatment cessation in rhesus macaques

The tissue origin(s) and the earliest viral dynamics of HIV rebound after antiretroviral therapy (ART) remain unclear. Here, using barcoded SIVmac239 in rhesus macaques ( n  = 24), we defined the distribution of barcode-specific viral RNA expression in tissues during ART ( n  = 6) and then assessed initial clonal rebound 5 and 7 days after ART cessation by identifying barcodes in individual tissues that exceeded the 99th percentile of the on-ART distribution (‘outliers’). In 4 of 11 aviraemic and 6 of 7 viraemic animals, 32 such outlier barcodes were identified. Sixteen of these barcodes were also identified in rebound viraemia, confirming specific tissues as rebound origin and early amplification sites. Overall, 27 of the 32 outlier barcodes were determined to reflect rebound origins, of which 96% were in the gastrointestinal tract (26%) or gastrointestinal tract-associated lymphoid tissues (70%). These results indicate that distinct tissue sites differentially support post-ART viral rebound, with potential therapeutic implications for interventions designed to prevent or control these events.