HPV 18 is associated with 2 divergent phenotypes: (i) aggressive cervical cancer and a preponderance of cancer relative to cervical intra-epithelial neoplasia (CIN) and (ii) benign warty lesions of the cervix. The E2 gene of HPV 18 encodes a regulatory protein that represses viral oncogene transcription and is involved in viral replication. Variation within the E2 gene of HPV 18 and its correlation with the morphologic grade of associated lesions were analyzed in a sample of 20 HPV 18-positive cervical specimens representing a spectrum of pathology from low-grade CIN to cervical cancer. An amplifiable HPV 18 E2 gene was present in 3 of 5 cancers, indicating that E2 disruption was not required for cancer development. Single-strand conformation polymorphism and PCR analyses revealed a high degree of polymorphism throughout the E2 gene. Direct DNA sequencing of both strands of a 154-bp fragment revealed a variability of 5.8%. Six intra-epithelial lesions contained alterations in common that account for 3.9% of the variation and appear to constitute a subtype. Within the 154-bp region, 2 of 3 cervical cancers and 0 of 12 intra-epithelial lesions were identical to the published HPV 18 sequence. DNA sequence analysis of a region extending into the E5 open reading frame revealed deletions in the E2/E5 intragenic region that were present in 50% of the members of the subtype. Our data demonstrate significant sequence variation within the E2 gene and suggest the presence of an HPV 18 subtype with decreased oncogenic potential. © 1995 Wiley-Liss, Inc.