Ureaplasmas from six animal species were inoculated into cell cultures derived from human, simian and murine hosts. All ureaplasmas infected at least two of the cell cultures tested. Supplemental medium urea was not a requirement for cell culture infection, and Ureaplasma growth in cell cultures occurred in as little urea as 4 micrograms/ml, similar to findings in broth-propagated organisms. These findings are of potential significance due to an observation (HM Jernigan, Jr: Exp Eye Res 37:551, 1983) that certain tissues produce urea locally. This may explain why ureaplasmas can be isolated from tissues outside the urinary tract where large amounts of urea are not routinely encountered. Ureaplasmas generally increased from titers of approximately 10(2) to 10(4) to ones of 10(5) to 10(6), but occasionally 10(7), color changing-units/ml within 2 days. Apparently a cell culture product is required for Ureaplasma growth. In several systems, Ureaplasma infection persisted and Ureaplasma-infected cell cultures could be passaged.