In resource-limited countries, nosocomial transmission of bloodborne pathogens is a major public health concern. After a major outbreak of human immunodeficiency virus (HIV) infection in approximately 400 children in 1998 in Libya, we tested HIV, hepatitis C virus (HCV), and hepatitis B virus (HBV) markers in 148 children and collected epidemiological data in a subgroup of 37 children and 46 parents. HIV infection was detected in all children but one, with HCV or HBV coinfection in 47% and 33%, respectively. Vertical transmission was ruled out by analysis of parents' serology. The children visited the same hospital 1-6 times; at each visit, invasive procedures with potential blood transmission of virus were performed. HIV and HCV genotypic analyses identified a HIV monophyletic group, whereas 4 clusters of HCV sequences were identified. To our knowledge, this is the largest documented outbreak of nosocomial HIV transmission.