The central demyelinating disease progressive multifocal leukoencephalopathy (PML) is caused by the human polyomavirus JC virus (JCV). JCV evolved as geographically based genotypes of which Type 3 is an African variant first characterized in HIV-1 positive patients from Tanzania. This study reports the complete sequence of five JCV Type 3 strains. The entire JCV genome was PCR amplified from urine specimens of three African and two African-American individuals. The African consensus sequence was compared to the Type 1 and Type 2 prototype strains, JCV (Mad-1) and JCV(GS/B), respectively. Type 3 differed in 2.2% of its coding region genome from JCV (Mad-1) and in 1.3% from JCV(GS/B). Within the coding region the sequence variation among the three types was higher in the capsid protein VP1 and in the regulatory protein large T antigen than in the agnoprotein or in VP2/3. Notable Type 3-specific changes were located at sites adjacent to the zinc finger motif and near the major donor and acceptor splice junctions of large T antigen. Four of the five urinary Type 3 strains had an unrearranged, archetypal regulatory region. African strain #309 showed a 10-bp deletion at a location similar to that previously described for #307 from Tanzania. The African-American Type 3 strain #312 was closely related to the African consensus sequence. The complete genome of a urinary JCV strain from another African-American male, previously reported as a possible Type 5, showed a sequence difference of only 0.52% from the Tanzanian consensus and has been reclassified as a subtype of Type 3.