Objective: To examine the prevalence of and risk for anal human papillomavirus (HPV) infection and abnormal anal cytology in sexually active adolescents. Design: Prevalence data from adolescents aged 13-18 years with and without HIV infection and with a history of high-risk sexual behavior. Methods: HPV DNA was detected using amplification techniques. Abnormal anal cytology was defined as atypical squamous cell of undetermined significance or worse. Results: Prevalence of anal HPV infection was similar in HIV-infected [28/58 (48%)] and uninfected [9/25 (36%)] boys (P = 0.3). but greater in HIV-infected [59/183 (59%)] than in uninfected [11/82 (13%)] girls (P < 0.001). Perianal warts were a risk for anal HPV in both boys [odds ratio (OR), 15.5; 95% confidence interval (CI), 1.6-149] and girls (OR, 9.9; 95% CI, 1.9-51.3). In subjects without anal warts, HIV infection was significant for girls (OR, 2.3; 95% CI, 1.1-4.9) and homosexual/bisexual orientation was significant for boys (OR, 5.2; 95% CI, 1.3-20.6). Abnormal anal cytology was more common among boys [32/77 (41.6%)] than girls [38/230 (16.5%)] (P < 0.001) and in addition to anal HPV, independent risk factors were positive HIV status in boys (OR, 6.5; 95% CI, 1.5-11.9) and number of partners within the past 3 months in girls (OR, 4.2; 95% CI, 1.5-11.9). Conclusions: Strong risk factors for abnormal anal cytology were HIV infection and anal HPV in boys and anal HPV and higher number of sexual partners for girls. The results suggest that anal cytology screening should be considered in HIV infected homosexual/bisexual males. © 2003 Lippincott Williams & Wilkins.