Abstract Body

Current ART has the capacity to drastically suppress but not abolish previously infected cells which can persist for the life of the host. This persistent rebound competent virus is the primary barrier to HIV cure. This long-term stable virus resides in multiple subsets of CD4+ T cells, particularly CD4+ memory T cells, and possibly macrophages that are widely distributed throughout the body. To mediate post-ART viral rebound, viruses must be genetically intact and replication competent, but viral rebound is not a wholesale resumption of viral replication from all replication-competent genomes but rather reflects an initial oligofocal outgrowth of virus from a limited subset of the total infected cell population that subsequently spreads throughout the body. Despite the importance of understanding the clonal origins of post-ART viral rebound, virtually nothing is known about these early local dynamic events, due to challenges with sampling, variability in time to rebound, and limited genetic diversity between individual rebounding lineages with previous work studying viral rebound only at later viremic stages or in limited numbers of tissues. To overcome these challenges, we developed and validated a barcoded virus model (SIVmac239M) that contains a short, unique nucleotide insert allowing for genetic discrimination of over 10,000 viral lineages. We used viral barcode analysis from a comprehensive collection of tissue samples in a large cohort of SIV-infected, ART suppressed rhesus macaque during both on-ART and post-ART cessation, to identify the initial sites of rebound and patterns of subsequent viral spread. Our results indicate that it is possible to identify the earliest stages of local tissue-level post-ART rebound and that this early rebound preferentially occurs in gut and gut-draining lymph nodes. Pinpointing the anatomic sites of viral rebound can facilitate the identification of the mechanisms that specifically mediate these reactivation events in these specific tissues. Together, these observations may have potential therapeutic implications for interventions designed to prevent or control viral rebound.