To investigate the involvement of S. anginosus infection in head and neck cancer in the extra-oropharyngeal cavity, we analyzed 3 DNA samples prepared from squamous cell carcinoma of the external auditory canal and one from squamous cell carcinoma of the skin using polymerase chain reaction (PCR) analysis and Southern blot analysis to detect the DNA sequence of S. anginosus. We also examined these four specimens by Gram's stain to detect the streptococcal bacterial bodies. By PCR analysis, the DNA sequence of S. anginosus was found in 4 out of 4 (100%) DNA samples obtained from these tumors. By Southern blot analysis, positive bands were detected in one out of the 3 (33%) samples from the tumor taken from the external auditory canal. We detected streptococcal bacterial bodies in one of the three specimens from the tumor obtained from cancer of the external auditory canal and in the one specimen from the skin cancer by the method of Gram's stain. Contrary to our expectations, these bacterial bodies were located in the middle of the tumor. Since S. anginosus is thought to exist in the mouth as a normal flora and to be located mainly in the gingiva and dental plaque, these data strongly indicate that S. anginosus infection is implicated in the carcinogenesis of head and neck squamous cell carcinoma.