The purpose of this retrospective cohort study was to identify individual-level demographic and community-level socioeconomic and health care resource factors associated with late diagnosis of HIV in rural and urban areas of Florida. Multilevel modeling was conducted with linked 2007-2011 Florida HIV surveillance, American Community Survey, Area Health Resource File, and state counseling and testing data. Late diagnosis (defined as AIDS diagnosis within 3 months of HIV diagnosis) was more common in rural than urban areas (35.8% vs. 27.4%) (p<0.0001). This difference persisted after controlling for age, sex, race/ethnicity, HIV transmission mode, country of birth, and diagnosis year (adjusted OR 1.39; 95% CI 1.17-1.66). In rural areas, older age and male sex were associated with late HIV diagnosis; zip code-level socioeconomic and county level health care resource variables were not associated with late diagnosis in rural areas. In urban areas only, Hispanic and non-Hispanic black race/ethnicity, foreign birth, and heterosexual mode of transmission were additionally associated with late HIV diagnosis. These findings suggest that, in rural areas, enhanced efforts are needed to target older individuals and men in screening programs and that studies of psychosocial and structural barriers to HIV testing in rural and urban areas be pursued.