The highly fibroblast‐passaged AD169, Towne, and Davis strains of cytomegalovirus (CMV) were found to have a restricted capacity to infect endothelial cells in vitro. Although such replication could be increased by a combination of low speed centrifugation and sodium butyrate treatment, the extracellular virus produced was infectious for fibroblasts but not for endothelial cells. In contrast, the low passage Toledo strain, and a low passage fibroblast‐grown clinical isolate of CMV, C1F, could be continually passaged in endothelial cells, giving the strains C1FE and Toledo.E. Whilst, using the conditions described above, initial infection of endothelial cells with AD169 or C1F resulted in similar titres of extracellular virus as assayed on fibroblasts, only the virus from the C1F strain was infectious for endothelial cells. Passage of C1F in fibroblasts decreased its ability to infect endothelial cells, whilst retaining equal ability to infect fibroblasts. Although endothelial‐cell‐passaged cell‐free C1FE virus was endothelial cell‐tropic, it was still much more infectious for fibroblasts than for endothelial cells. It is concluded that the C1F and Toledo strains, but not the AD169, Towne, or Davis strains, contained endothelial cell tropic variants, which could be lost on passage through fibroblasts, but retained on passage through endothelial cells. Furthermore, virus in an ex vivo source of CMV, a blood specimen, was found to be more tropic for fibroblasts than for endothelial cells, suggesting that in vivo CMV exists as quasi strains with different cell tropism, some of which might be lost in vitro by passage in an inappropriate cell type. J. Med. Virol. 57:298–307, 1999. © 1999 Wiley‐Liss, Inc.