Sequence variation in the gp116 component of cytomegalovirus envelope glycoprotein B was examined in 11 clinical strains and compared with variation in gp55. The peptide variation in gp116 was found to be strongly clustered at codons 27-67, 440-460, and to a lesser extent at codons 181-194, 311-317, and 387-397. Strains adopted one of three to four peptide configurations at these loci, usually consistent with their gp55 sequence configuration. Two instances were observed of a sequence variation arising from recombination within gB. The limited, largely group-specific nature of variation in both gp116 and gp55 facilitates functional and immunologic analyses.