Genetically distinct lentiviruses constitute a quasispecies population that can evolve in response to selective forces. To move beyond characterization of the population as a whole to the behavior of individual members, we devised an in situ hybridization approach that uses genotype- specific probes. We used probes that detect simian immunodeficiency viruses (SIV) that differ in sequence in the V1 region of the surface envelope glycoprotein (env) gene to investigate the replication and cellular tropisms of four viral variants in the tissues of infected rhesus macaques. We found that the V1 genotypic variants replicated in spatially defined patterns and to different extents at each anatomic site. The two variants that replicated most extensively in animals with AIDS were detected in both macrophages and T lymphocytes in tissues. By extension of this approach, it will be possible to investigate the role of individual lentiviruses in a quasispecies in pathogenesis and to evaluate the effects of antiviral or immunotherapeutic treatment on select members of a quasispecies.