We investigated the prevalence of human papillomavirus types 16, 18 and 33 deoxyribonucleic acid (DNA) by polymerase chain reaction and the localization of human papillomavirus DNA by in situ hybridization using formalin-fixed, paraffin-embedded tissue specimens of penile carcinoma in Japan. Of 111 untreated penile carcinomas 70 (63.1%) were positive for human papillomavirus DNA by the polymerase chain reaction method. Human papillomavirus type 16 was identified in 68 penile carcinoma cases and type 18 in 2, whereas type 33 could not be detected. Of 12 treated penile carcinomas 2 (16.7%) were positive for human papillomavirus type 16. These data indicate that human papillomavirus type 16 DNA is the type most commonly associated with penile carcinoma. Human papillomavirus type 16 was also detected in lymph node metastasis of penile carcinoma, and that was the same as the human papillomavirus DNA type in the primary carcinoma. The in situ hybridization analysis found the human papillomavirus DNA to be localized in the nuclei of carcinoma cells.