We have begun a systematic study of human papillomavirus (HPV) infection in colposcopically and/or morphologically normal epithelium of the uterine cervix. Paired biopsies were taken from the lesions (cervical intraepithelial neoplasia [CIN], condyloma, invasive carcinoma) and from the normal-appearing adjacent epithelium 3 to 5 mm from the edge of the lesion. Myometrium or ectocervical epithelium from patients who had undergone hysterectomy for reasons other than genital dysplasia or malignancy served as controls. One biopsy was examined histologically. DNA from the second biopsy was digested with Pst I, and the presence or absence of HPV was determined by Southern blotting using HPV-16 DNA as a probe. HPV was not detected in any of the 12 control samples. Of 30 patients with CIN and/or condyloma, five of 18 who were HPV-positive had either HPV-16 (three cases) or virus resembling HPV-31 (two cases) in the lesion and adjacent epithelium. Of seven patients with invasive carcinoma, four had HPV in the lesion and adjacent epithelium; two of these four patients had typical HPV-16. Such infection of apparently normal epithelium has major implications for our understanding of the pathogenesis, treatment, and follow-up of patients with cervical neoplasia.