Simian immunodeficiency virus (SIVcpz) from the chimpanzee subspecies Pan troglodytes troglodytes has been linked phylogenetically to the origin of HIV-1. Related but distinct SIVcpz strains have also been found in P. t. schweinfurthii , suggesting that SIVcpz may have coevolved among the four chimpanzee subspecies. However, SIVcpz strains from P. t. verus and P. t. vellerosus have not yet been identified. To better understand the epidemiology and natural history of SIVcpz among chimpanzees, we tested serum samples from 1415 chimpanzees housed at eight U.S. research centers and six zoos. Records indicated that 264 (18.6%) of the chimpanzees were African-born. Subspecies identities for 161 chimpanzees, based on analysis of mitochondrial DNA sequences, were found to be P. t. troglodytes (n = 14), P. t. schweinfurthii (n = 3), P. t. verus (n = 143), and P. t. vellerosus (n = 1). All samples were screened for HIV/SIV antibodies by using an HIV-1/2 peptide- based enzyme immunoassay (EIA). Reactive samples were tested further by Western blot (WB). Eight sera (0.57%) were EIA reactive, but none was HIV-1/2 WB positive. Two samples were HIV-1 WB indeterminate. Both samples tested negative for SIVcpz and HIV-1 sequences by reverse transcriptase PCR, suggesting an absence of infection. We also tested sera available from 8 male sexual partners, 6 offspring, and 12 cage mates of a known SIVcpz-infected chimpanzee. All samples were negative, suggesting that SIVcpz may not be easily transmitted to close contacts. Our data show that this large population of chimpanzees is not infected with SIVcpz. The absence of SIVcpz infection in P. t. verus suggests that SIVcpz may not be endemic to this subspecies and implies that SIVcpz may have been introduced more recently into the chimpanzee subspecies following divergence.