OBJECTIVE Human papillomavirus (HPV) is the necessary cause of cervical cancer and Chlamydia trachomatis (CT) is considered a potential cofactor in the development of cervical intraepithelial neoplasia (CIN). The objective of this pilot study was to determine the association of CT infection with HPV, other risk factors for cervical cancer, and CIN in symptomatic women. METHODS A total of 600 consecutively selected women aged 30-74 yr with persistent vaginal discharge, intermenstrual/postcoital bleeding or unhealthy cervix underwent conventional Pap smear, Hybrid Capture 2 (HC2) testing for HPV and CT DNA and colposcopy, with directed biopsy of all lesions. RESULTS HPV DNA was positive in 108 (18.0%) women, CT DNA in 29 (4.8%) women. HPV/CT co-infection was observed in only four (0.7%) women. Of the 127 (21.2%) women with Pap >ASCUS, 60 (47.2%) were HPV positive and four (3.1%) were CT positive. Of the 41 women with CIN1 lesions, 11 (26.8%) were HPV positive, while two were CT positive. Of the 46 women with CIN2+ on histopathology, 41 (89.1%) were HPV positive, two (4.3%) were CT positive and one was positive for both. The risk of CIN2+ disease was significantly increased (P<0.05) by the following factors: age <18 yr at first coitus, HPV infection and a positive Pap smear. Older age (>35 yr), higher parity, use of oral contraceptives or smoking did not show any significant association with HPV or abnormal histopathology. Parity >5 was the only risk factor positivity associated with CT infection (P<0.05). CONCLUSIONS Our findings showed that CT infection was not significantly associated with CIN, and most of its risk factors, including HPV infection, in symptomatic women. Longitudinal studies with carefully selected study sample would be able to answer these questions.