As a first step in understanding the role that health care providers may play in observed gender disparities in HIV care in the United States, we sought to examine whether HIV providers' perceptions of and attitudes toward female and male patients differ. We used data from the Enhancing Communication to Improve HIV Outcomes (ECHO) study, a multisite, cross-sectional study focused on the role of the patient-provider relationship in disparities in HIV care conducted from October 2006 to June 2007. Using separate scales, we assessed HIV providers' perceptions about their patients (e.g., intelligence, compliance, responsibility) as well as providers' attitudes toward their patients (e.g., like, respect, frustrate). We used multivariable linear regression with generalized estimating equations to compare provider scores for female and male patients. Our sample comprised 37 HIV providers and 317 patients. Compared with male patients, HIV-infected females were less likely to be highly educated or employed, and more likely to report nonadherence to antiretroviral medications and depressive symptoms. In unadjusted and adjusted analyses, there was a significant difference in providers' perceptions of female and male patients, with providers having more negative perceptions of female patients. However, there was no significant difference in HIV providers' attitudes toward female and male patients in unadjusted or adjusted analyses. Further study is needed to elucidate the role of providers' perceptions and attitudes about female and male patients in observed gender disparities in HIV care.