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A cross-sectional study was conducted to assess the prevalence of human immunodeficiency virus (HIV), hepatitis B virus (HBV), and their coinfection among blood donors at the National Blood Transfusion Center in Bamako, Mali, from November 2001 to July 2002. Enzyme-linked immunosorbent assay techniques with reagents from Bio-Rad (France) were used to test the blood samples. 11 592 blood donors were tested for HIV and HBV surface antigens. The prevalence of HIV was 4.5% and the prevalence of HBV was 14.9%. The HIV/HBV coinfection rate was only 1.13% in this population. The coinfection rate was unexpectedly low in this blood donor population where monoinfection with HIV or HBV prevalence was high.