CONTEXT: Low-grade squamous intraepithelial lesions (LSILs) have been described as a benign cytological consequence of active human papillomavirus (HPV) replication. Several studies have reported that certain behavioral and biological risks exist for LSIL, suggesting that HPV alone is not sufficient for the development of LSIL. However, because most of these studies have been cross-sectional, it is not known whether behavioral and biological risks are simply risks for HPV infection itself. OBJECTIVE: To prospectively examine risks of incident HPV infection in HPV-negative females and of incident LSIL development in females with HPV infection. DESIGN: Prospective cohort study conducted between 1990-2000, with a median follow-up of 50 months. SETTING AND PARTICIPANTS: Females aged 13 to 21 years who attended 2 family planning clinics in the San Francisco bay area; 496 had prevalent HPV infection and 105 were HPV-negative. MAIN OUTCOME MEASURE: Incident development of HPV infection and LSIL, analyzed by various demographic, behavioral, and clinical risk factors. RESULTS: Fifty-four incident HPV infections occurred in the 105 females who were HPV-negative at study entry (median duration of follow-up for those who remained HPV-negative was 26 months). Multivariable analysis showed that risks of HPV included sexual behavior (relative hazard [RH], 10.10; 95% confidence interval [CI], 3.24-31.50 per new partner per month), history of herpes simplex virus (RH, 3.54; 95% CI, 1.37-9.10), and history of vulvar warts (RH, 2.73; 95% CI, 1.27-5.87). Current use of oral contraceptives had a significantly protective effect (RH, 0.49; 95% CI, 0.28-0.86). Among the 496 individuals who were HPV-positive at baseline or in follow-up, there were 109 incident cases of LSIL during the follow-up interval, with a median follow-up time of 60 months for those who never developed LSIL. Human papillomavirus infection was the most significant risk factor for development of LSIL. The multivariable model showed the following risks for LSIL: HPV infection for less than 1 year (RH, 7.40; 95% CI, 4.74-11.57); HPV infection for 1 to 2 years (RH, 10.27; 95% CI, 5.64-18.69); HPV infection for 2 to 3 years (RH, 6.11; 95% CI, 1.86-20.06); and daily cigarette smoking (RH, 1.67; 95% CI, 1.12-2.48). CONCLUSION: Our results indicate distinct risks for HPV and LSIL. In addition, most women with HPV infection in our study did not develop LSIL within a median follow-up period of 60 months. These findings underscore the hypothesis that certain biological risks thought to be associated with LSIL are, in fact, risks for acquisition of HPV. Cigarette smoking was a risk specific to LSIL, supporting the role of tobacco in neoplastic development.