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A new simian-human immunodeficiency virus (SHIV) stock (SHIV 89.6-PD), derived from plasma of a rhesus macaque used for in vivo serial passage of virulence-attenuated SHIV 89.6, produces systemic infection after intravenous, intravaginal, or intrarectal inoculation of rhesus macaques. Infection with this virus results in high levels of viral antigen in plasma, a precipitous decline in CD4+ T-cell counts, and a disease syndrome that is characteristic of AIDS. Rapid progression to disease was associated with failure to seroconvert to viral antigens, whereas longer survival was associated with production of antiviral antibodies. In Intravenously inoculated animals, peak antigenemia occurred at 7 days postinjection (PI) and severe CD4+ depletion occurred at 14 days PI. In mucosally infected animals, peak antigenemia occurred at 14 days PI and severe CD4+ depletion was not evident until 21 days PI. The 1-week delay in both viral antigenemia and CD4+ T-cell decline in mucosally infected animals is consistent with the hypothesis that, following vaginal inoculation, virus dissemination proceeds in a stepwise manner from the mucosal surface to the draining lymph nodes and subsequently to the bloodstream. This animal model can be used to test the ability of HIV-1 envelope-based vaccines to prevent infection or disease after challenge by the three major routes of HIV transmission.