The objective of this study was to assess the distribution of Chlamydia trachomatis (C. trachomatis) genovars in male sexually transmitted disease (STD) clinic patients from 1 province in eastern China (Jiangsu) and 3 provinces in southern China (Guangdong, Guangxi, and Hainan). Urine specimens (n = 140) were collected from 46 public STD clinics in the 4 provinces. Specimens that tested positive for C. trachomatis by polymerase chain reaction were genotyped for the infecting C. trachomatis strain by amplifying and sequencing the genovar-specific ompA gene, which encodes the chlamydial major outer membrane protein. Nine C. trachomatis genovars were identified from 129 specimens; they consisted of the F (25.6%, n = 33), E (17.1%, n = 22), J (16.3%, n = 21), D (15.5%, n = 20), G (11.6%, n = 15), K (5.4%, n = 7), H (3.9%, n = 5), I (1.6%, n = 2), and B (0.8%, n = 1) genovars. Nine genovars were found in specimens from Guangxi province, 6 were found in specimens from each of Guangdong and Hainan provinces, but only 5 were found in specimens from Jiangsu province. Significant differences were observed in C. trachomatis genovar distributions between different provinces: G/Ga was absent among male STD clinic patients (MSPs) from the eastern province of Jiangsu (Fisher exact test = 0.036), while being prevalent in the 3 southern provinces.