Objective: To ascertain the likelihood that perivascular leukocyte infiltrates are sites for replication and dissemination of HIV-1. Design and methods: We measured the ability of HIV patients' peripheral blood mononuclear leukocytes to migrate through confluent endothelial monolayers in vitro and infect phytohemagglutinin-stimulated allogeneic lymphoblasts. We also measured the ability of migratory leukocytes to transmit virus to uninfected leukocytes that have localized outside an endothelial barrier, and the subsequent ability of these newly infected cells to reverse-migrate back across the endothelial barrier - a process that might facilitate reentry of infected cells into the circulation and dissemination of the virus to distant sites. Results: Leukocytes from 27 out of 63 unselected patients spontaneously carried infectious virus across endothelial barriers. On follow-up, these 27 patients were frequently observed to develop uncontrolled viremia, despite treatment, and be hospitalized for secondary infections. Migratory leukocytes transmitted HIV to both T lymphocytes and non-T cells that had previously crossed the endothelial barrier. Either cell type could subsequently reverse-migrate carrying virus back across this barrier. Conclusions: Reverse-migration of HIV-1 infected leukocytes out of perivascular reservoirs may provide a way to disseminate HIV-1 and expand the body burden of virus in some patients receiving highly active antiretroviral therapy. © 2002 Lippincott Williams & Wilkins.