Human papillomavirus (HPV) is detected in nearly all cervical cancers and approximately half of vaginal cancers. However, vaginal cancer is an order of magnitude less common than cervical cancer, not only in the general population but also among women with HIV/AIDS. It is interesting therefore that recent studies found that HPV was common in both normal vaginal and cervical tissue, with higher prevalence of nononcogenic HPV types in the vagina. In our investigation, we prospectively examined HPV infection in 86 HIV-positive and 17 HIV-negative women who underwent hysterectomy during follow-up in a longitudinal cohort. Cervicovaginal lavage specimens were obtained semi-annually and tested for HPV DNA by polymerase chain reaction. To address possible selection biases associated with having a hysterectomy, subjects acted as their own comparison group—before versus after hysterectomy. The average HPV prevalence was higher in HIV-positive than HIV-negative women both before (59% vs. 12%; p < 0.001) and after hysterectomy (56% vs. 6%; p < 0.001). Multivariate random effects models (within-individual comparisons) demonstrated significantly lower HPV prevalence [odds ratio (OR) = 0.71; 95% confidence interval (CI) = 0.59–0.85) after hysterectomy. The association of HPV prevalence with hysterectomy was similar among HIV-positive and HIV-negative women. However, hysterectomy had greater effects on oncogenic (OR = 0.48; 95% CI = 0.35–0.66) than nononcogenic HPV types (OR = 0.89; 95% CI = 0.71–1.11; Pinteraction = 0.002). Overall, we observed greater reductions in oncogenic than nononcogenic HPV prevalence after hysterectomy. If correct, these data could suggest that oncogenic HPV have greater tropism for cervical compared to vaginal epithelium, consistent with the lower incidence of vaginal than cervical cancer.