In a blind study, DNA from 40 clinical isolates of herpes simplex viruses was analyzed by restriction endonucleases to determine whether serial isolates from an individual patient could be identified and whether exogenous reinfection occurred within this population. Five of the 40 isolates served as controls. Based on restriction patterns obtained following Bam H₁ cleavage of DNA, 35 isolates were assigned to 15 patients. Isolates from two patients displayed variation in the electrophoretic mobility of certain Bam H₁ fragments. However, the isolates from one patient were identical when digested with four additional enzymes. One of three sequential isolates from a patient, when cleaved with Eco R₁ and Hind III, showed variable fragments in the terminal and joint regions of the S segment of the genome. The appearance of the fragments was not due to the addition of a known restriction site. We conclude that exogenous reinfection with a genetically distinct strain of herpes simplex virus type 2 (HSV-2) did not occur among these 15 patients with recurrent genital HSV infections.