Purpose This study has two aims: determine the prevalence of CIN3 + in patients with discordant cotesting, defined as negative cytology and positive human papillomavirus (HPV) testing, and identify factors (including HPV strain) associated with CIN3 + , defined as cervical intraepithelial neoplasia (CIN) 3 or cancer within this population. Methods We conducted a retrospective chart review of women age 30-65 with intact cervices who had discordant cotesting results between January 1, 2013 and September 1, 2018, at an academic medical center. We used the t test for continuous variables and the Chi-square test for categorical variables to compare women with and without CIN3 + . To identify factors associated with CIN3 + , we performed univariate and multivariate logistic regression. Results The primary outcome was the prevalence of CIN3 + based on pathologic diagnosis following biopsy or excisional procedure. Among 375 patients with discordant co-testing, the mean age was 43.8 years, 58.4% were parous, and 84.8% were white. Overall, 43/375 (12.0%) had CIN3 + and 7/375 (1.9%) had AIS. On logistic regression, only parity >= 1 (p = 0.04, adjusted OR = 2.23, CI = 1.06-4.68) was significantly associated with CIN3 + . HPV-18 was less likely to be associated with CIN3 + (p = 0.02, adjusted OR 0.08, CI 0.01-0.65) but was present in 43% of AIS cases. HPV16 and other HR-HPV strains were highly associated with CIN3 + . Conclusion Women with discordant cotesting are at significant risk for CIN3 + . We recommend that biopsy be performed at the time of indicated colposcopy for all patients with discordant cotesting to assess for high-grade dysplasia.