Coinfection by multiple human cytomegalovirus (HCMV) strains was investigated in immunocompetent individuals and AIDS patients. Thirty HCMV maternal and fetal or newborn isolate pairs from 9 cases of congenital HCMV infection as well as 36 HCMV isolates and 2 PCR-HCMV-positive CSF samples from 13 AIDS patients were tested by restriction fragment length polymorphism analysis of multiple genome regions. Results from the group of congenital infections showed that: i) all the 9 women with primary HCMV infection presumably harboured a single HCMV strain; ii) all strains were genetically unrelated; iii) isolates from infected fetuses or newborns consisted of a single strain apparently indistinguishable from the maternal strain; iv) no strain variations were observed in isolates from different body sites or in sequential isolates from newborns up to 8 months after birth. Results from the AIDS patient group demonstrated that; i) all patients were infected by unrelated strains; ii) 6/13 (46.1%) patients were coinfected by 2 or more HCMV strains; iii) in a single patient two different HCMV strains were detected in blood and urine, respectively, whereas a mixture of the two was found in the pharynx; iv) 4 patients showed the sequential appearance of a mixed virus population or different strains suggesting sequential reinfections.