High titre replication of human cytomegalovirus (HCMV) in cell culture is restricted to primary human fibroblasts. During acute infection in vivo, HCMV nucleic acids and antigens have been found in various organs. Using only morphological criteria, inconsistent data have been reported about the cell types that can be infected by HCMV. In particular, the role of fibroblasts in organ infections has remained unclear. To define accurately the target cells of HCMV in vivo, tissue sections from lung and gastrointestinal tract of patients suffering from acute HCMV infection were investigated using immunohistochemical double-labelling analyses. Monoclonal antibodies with defined specificity against immediate early (IE), early (E) and late (L) viral antigens and antibodies directed against cell marker proteins were employed to identify infected cells. The results demonstrated that a broad spectrum of cells was infected by HCMV in vivo. Consistent with their susceptibility in culture, fibroblasts formed a major population of HCMV-infected cells. In contrast, haemopoietic cells were only infrequently stained with virus-specific antibodies. Fibroblasts, epithelial cells, endothelial cells, smooth muscle cells and macrophages appeared to be permissive for HCMV replication. Contrary to this, polymorphonuclear cells showed only IE gene expression, indicating that these cells were abortively infected. The analysis of the distribution of infected cells in tissue supported the hypothesis that endothelial cells and monocytes/macrophages may play a crucial role in the haematogenous spread of HCMV; in contrast, fibroblasts, smooth muscle cells and epithelial cells may form the cell populations important for the multiplication and spread of the virus in infected tissues.