The objective of this study was to clarify the longitudinal epidemiology of Chlamydia trachomatis serovars in Japan. A total of 339 endocervical swab specimens obtained from female patients who attended the Department of Obstetrics and Gynecology, Saitama Medical School, were used. Positive specimens of either transport medium of IDEIA Chlamydia (1st group, from 1999 to 2001), or DNA extract of Cobas Amplicor STD-1 Chlamydia trachomatis (2nd group, from 2003 to 2005) were used for serotyping. Typing of C. trachomatis serovars in DNA extracts was performed by polymerase chain reaction-restriction fragment length polymorphism. Ten serovars, A, B, D, E, F, G, H, I, J and K, were identified in the 1st group, and serovar E was most frequently identified (27.6%). In the 2nd group, nine serovars, B, C, D, E, F, G, H, I and K, were identified, and serovar D was most frequently identified (24.7%). Serovars B and Ba were significantly more common around 2000 and the mid-1990s (from 1993 to 1996), respectively. Numbers of serovar I increased significantly during the research period. In addition, serovar I was more frequent in the 2nd group than in the 1st group in women aged 20-29 years. There were no significant differences of serovar distribution between pregnant and non-pregnant women.