We have cloned and sequenced the glycoprotein B genes from five strains of BaCMV, isolated from three subspecies of cynocephalus baboons (olive, yellow and chacma). Primers were designed using conserved DNA regions of the gB gene to allow DNA amplification from all strains of BaCMV. These regions differ sufficiently from human CMV that HCMV strains are not amplified, thus allowing differentiation of BaCMV from HCMV. These diagnostic primers were used to test crude nucleic acid extracts from 27 strains of BaCMV and detected 26 of them. Overall, the sensitivity and specificity of this assay are 96.7 and 100%, respectively. BaCMV strains isolated from yellow and olive baboons were very similar and could be discriminated from strains isolated from chacma baboons using a second set of PCR primers. Phylogenetic analysis of the gB genes supported the inferred close relationship of strains isolated from olive and yellow baboons.