Arginine is necessary for the development of the cytopathogenic effect of human cytomegalovirus in human embryonic fibroblasts. It is also required, though in greater concentrations, for the production of infective virions, the requirement being at an early stage of replication. Inhibitor studies suggested that this block in replication caused by arginine deficiency was prior to the formation of viral DNA. Withdrawal of arginine from the medium 24 or 48 hours after infection resulted in a decline in virus production indicating that the continued presence of the amino acid is necessary for constant virus production. Infected cultures deprived of arginine could be stimulated to produce cytopathic effects and infective virions by replacement of the amino acid even eight days after inoculation, demonstrating that the information for cytomegalovirus replication remains intact within the cell. This establishment of latency in vitro may be related to the ability of the virus to establish a similar state in vivo.