A prospective evaluation of 146 children with fever of unknown origin (FUO) and prolonged fever was performed from 1990 to 1996. FUO was defined as a documented daily temperature of > or = 38 degrees C for at least 14 days without diagnostic signs or symptoms. Prolonged fever was defined as fever for at least 14 days and no diagnosis at the time of referral for evaluation. An established diagnosis was made for 84 (57.5%) of 146 patients. The most common infectious disease diagnoses were Epstein-Barr virus infection (22 [15.1%] of 146), osteomyelitis (14 [9.6%] of 146), bartonellosis (7 [4.8%] of 146), and urinary tract infection (6 [4.1%] of 146). Three of seven patients with confirmed Bartonella henselae infection presented with FUO and no ultrasonographic findings compatible with hepatosplenic involvement; two patients presented with FUO and hepatosplenic involvement. The relatively common finding of acute bartonellosis in this population suggests that FUO and prolonged fever in children are other presentations of infection with B. henselae.