In contrast to simian immunodeficiency viruses (SIVs), which induce immunodeficiency over a 1- to 3-year period, highly pathogenic simian-human immunodeficiency viruses (SHIVs) cause a complete, irreversible, and systemic depletion of CD4⁺ T lymphocytes in rhesus monkeys within weeks of infection. By using small-molecule competitors specific for CCR5 and CXCR4 in ex vivo assays, we found that highly pathogenic SHIV_DH12R exclusively uses CXCR4 for infection of rhesus peripheral blood mononuclear cells, whereas SIV_mac239 and SIV_smE543 use CCR5 for entry into the same cells. During the period of peak virus production in SHIV_DH12R- or SHIV_89.6P-infected rhesus monkeys, massive elimination of CXCR4⁺ naïve CD4⁺ T cells occurred. In contrast, circulating CCR5⁺ memory CD4⁺ T cells were selectively depleted in rapidly progressing SIV-infected monkeys. At the time of their death, two SIV rapid progressors had experienced a nearly complete loss of the memory CD4⁺ T cell subset from the blood and mesenteric lymph nodes. Thus, pathogenic SHIVs and SIVs target different subsets of CD4⁺ T cells in vivo, with the pattern of CD4⁺ T lymphocyte depletion being inextricably linked to chemokine receptor use. In the context of developing an effective prophylactic vaccine, which must potently control virus replication during the primary infection, regimens that suppress SHIVs might not protect monkeys against SIV or humans against HIV-1.