Genital ulcers (GU) have been rarely described in temporal association with acute Epstein-Barr Virus (EBV) infection, but a cause-and-effect relationship has not been established. A previously healthy 14-year-old girl with no history of sexual activity developed fever, sore throat, and low back pain; a day later, five painful ulcers appeared over her external genitalia. A hemogram was normal, herpes simplex virus (HSV) culture of the lesions was negative, and bacterial GU cultures grew normal flora. Her fever worsened(40.2°C) and the genital ulcers continued to enlarge. Examination on day 5 of GU showed deep, tender, hemorrhagic, coalesced ulcers involving both labia majora and minora, 2 mm aphthous ulcers on her buccal mucosa, no lymphadenopathy, and the hymen was estrogenized and without interruption. HSV cultures, ANA, VDRL, and HIV antibodies were negative. She was empirically treated with Augmentin®(2d) and then metronidazole (7d), as well as oral methylprednisolone (5d). On day 9 of GU, she was afebrile and the lesions were smaller and not hemorrhagic. A tentative diagnosis of Behcet syndrome was made. On day 15 of GU, she developed sore throat and dysphagia, but no fever. Examination revealed exudative tonsillitis and cervical lymphadenopathy; the ulcers were granulating and less tender. Her Monospot, EBV VCA IgG and IgM, and anti-EA (1:40) were positive; EBNA was negative. Whole blood, serum, and buffy coat obtained on day 16 of GU and the genital culture specimen obtained on day 5 (10 days before primary EBV infection was diagnosed) were assayed for EBV DNA using the polymerase chain reaction (PCR). The PCR assay was used to amplify a 177 bp region in the LMP-2A gene of EBV; the sensitivity of this assay was 8 EBV copies per 105 cells. All specimens were positive for EBV DNA. Conclusion: EBV DNA was found in genital ulcer specimens obtained 10 days before symptoms of EBV-related infectious mononucleosis were apparent. EBV should be considered as an etiologic agent of acute, non-venereal genital ulcers.