Epstein-Barr virus (EBV) has been implicated in a variety of central nervous system syndromes. In a few well-studied patients, EBV has been detected by viral isolation or EBV DNA has been found by Southern hybridization analysis. Using polymerase chain reaction, we evaluated brain biopsy specimens from 24 patients for the presence of EBV genomes. EBV DNA was found in brain specimens from 18 patients in whom presence of the virus in the brain was suspected clinically or on the basis of serological tests. Six patients had acquired immunodeficiency syndrome; 2 were kidney transplant recipients. Brain specimens from 4 patients with encephalitis due to other herpes group viruses and from a patient with metabolic encephalopathy were negative for EBV DNA as determined by polymerase chain reaction. The findings indicate a need to evaluate the role of EBV in diverse neurological syndromes, especially those occurring in immunodeficient hosts.