To substantiate the hypothesis that multiple infection by human herpesvirus 8 (HHV-8) and by cytomegalovirus (CMV) commonly occurs, intraperson nucleotide variation in DNA amplified from hypervariable subgenomic domains of HHV-8 and CMV was evaluated in a group of people living in Malawi, where both viruses circulate hyperendemically. Mouth rinses, throat gargles, palatal exfoliates and blood were sampled from 89 people (age range: 4 m to 45 y). In 24 people (27%), HHV-8 DNA could be amplified in >1 sample 9 (38%) were seropositive for human immunodeficiency virus-1 and 7 (29%) exhibited Kaposi's sarcoma. Sequence variation was sought in DNA segments derived from: the domain in open reading frame (ORF) 73 that encodes latent nuclear antigen the Bam330 segment of ORF 26 and the VI region of ORF Kl. Restriction fragment-length polymorphism analysis, nucleotide sequencing, PCR cloning and denaturing gel gradient electrophoresis were applied to study the sequence diversity of these segments. Significant intraperson/inter- and intra-sample sequence polymorphisms could be found in 15 people (62.5%). Variation in urine-derived VI sequences could, in addition, be evaluated: in 5 people, the sequences were monotypic, and in 2, urinary and oral sequences were genotypically different. Variation in hypervariable domains in the gO and gN regions of CMV was studied in urine and saliva samples of 77 people. In 41 individuals (53%), DNA could be amplified from at least one domain, and, in 14 (18%), from both domains. In 4 individuals (5%), urinary and oral sequences were genotypically different. The extent of inter- and intra-person variation in the 2 CMV domains studied was significantly less than inKWl of HHV-8. Multiple infection by HHV-8 and by CMV is common. It would be important to determine if such infection reflects coinfection occurring during initial transmission or superinfection, as the latter implies that vaccination might be ineffective to prevent and control the spread of the viruses.