This is the first study to investigate vertical transmission of Chlamydia trachomatis in Chongqing China. For this study, 300 cervical swab samples from pregnant women and 305 nasopharygeal swab samples from their babies (605 specimens) were collected for nest polymerase chain reaction (nPCR) of the ompl gene, which encodes the major outer membrane protein (MOMP) and typed C. trachomatis using Cleavase fragment-length polymorphism (CFLP) labeled with digoxin. From these samples, 11% (33/300) of pregnant women samples were successfully amplified. The vertical transmission rate of C. trachomatis from mother to baby was 24% (8/33). The vertical transmission rates were 66.7% (6/9) for mothers with vaginal delivery and 8.3% (2/24) for those with cesarean section. The incidence of premature membrane rupture among C. trachomatis-positive pregnant women was 30.3% (10/33), which was greater than among those who were C. trachomatis-negative (13.5%, 36/267; chi(2) = 4.2; p < 0.05). Four genotypes including type E (3 pairs), type F (2 pairs), type H (2 pairs), and type D (1 pair) were observed by CFLP assay labeled with digoxin and confirmed by DNA sequencing in the 16 C. trachomatis-positive samples from eight pregnant women and their eight infants. Each pair of matched maternal-infantile samples showed identical CFLP. This study showed the incidence of C. trachomatis infection in pregnant women, the vertical transmission rate for C. trachomatis, and the genotypes of C. trachomatis in Chongqing, China. The CFLP assay labeled at the 5' end of the forward primer with digoxin was first used successfully to genotype of C. trachomatis. As a promising method for C. trachomatis genotyping, CFLP had good sensitivity, reproducibility, and simplicity and no radioactive contamination.