During the development of adenovirus (Ad) vaccines in the 1950s, Ad strains 1-5 and 7 recombined with SV40 during adaptation to growth in rhesus monkey kidney cells. The recombination events between the Ad and SV40 genomes produced hybrid viruses that contained, within the Ad genome, either portions of the early region of the SV40 genome or single or multiple copies of the entire SV40 genome inserted in a configuration that permitted the generation of SV40 progeny. When portions of the SV40 early region were inserted into the Ad2 E3 region, a region non-essential for viral replication, the resulting Ad-SV40 hybrids were non-defective in that they were capable of independent replication. Such hybrids have no known selective advantage for replication in human tissues; however, through inadvertent human exposure it is theoretically possibly that they could be induced to spread in the environment. Because of deletions in the Ad genome, the Ad-SV40 hybrids that contain infectious SV40 DNA were defective and were not capable of replication without a helper virus. Due to the low frequency with which cells in an infected individual could be co-infected with both defective hybrid and helper virions, it is unlikely that such defective viruses could be established in the population. Based on the Ad-SV40 model, it is theoretically possible that SV40 could recombine with other DNA viruses that infect humans. The introduction of Ad-SV40 hybrids or SV40-other virus hybrids into the environment could contribute to establishing SV40 as a human polyomavirus and to the SV40 DNA sequences that are being detected in human tissues.