BACKGROUND: The intestinal mucosa contains many cells targeted by human immunodeficiency virus type 1 (HIV-1), and high levels of HIV-1 DNA persist in this compartment under antiretroviral therapy (ART). While CD4+ T cells are the best-characterized reservoir of HIV-1, the role of long-lived intestinal macrophages in HIV-1 persistence on ART remains controversial. METHODS: We collected duodenal and colonic biopsies from 12 people with HIV (PWH) on suppressive ART, enrolled in the ARNS EP61 GALT study. Total, integrated, intact and defective HIV-1 proviruses were quantified from sorted T cells and monocytes/macrophages. HIV-1 env quasispecies were analyzed by single-molecule next-generation sequencing and env-pseudotyped viruses were constructed to assess macrophage versus T-tropism. RESULTS: Total HIV-1 DNA levels in intestinal T cells were significantly higher than those in monocytes/macrophages (P <
.0001). Unintegrated HIV-1 DNA was detected in one-third of T-cell and monocyte/macrophage-positive samples. Intact HIV-1 proviruses were detected using the intact proviral DNA assay in 4 of 16 T-cell samples and 1 of 6 monocyte/macrophage samples with detectable HIV-1 DNA. HIV-1 sequences were compartmentalized between intestinal monocytes/macrophages and T cells in some subjects. Phenotypic analysis of pseudotyped viruses expressing HIV-1 envelopes from colonic monocytes/macrophages revealed T-tropism rather than M-tropism. CONCLUSIONS: Our results show that monocytes/macrophages from the intestinal mucosa of PWH on ART can contain HIV-1 DNA, including intact or unintegrated forms, but at much lower levels than those found in T cells, and with some compartmentalization, although they do not exhibit a specific macrophage tropism, raising the question of the mechanisms of infection involved.