Background: Expansion of the health care workforce in Peru to combat tuberculosis (TB) includes both professional health care providers (HCPs) such as doctors and nurses, and non-professional HCPs such as community health workers (CHWs). We describe the knowledge and attitudes of these HCPs, and identify modifiable barriers to appropriate anti-tuberculosis treatment. Methodology: We surveyed HCPs practicing in 30 clinical settings (hospitals, community health centers, and health posts) in the San Juan de Lurigancho district of Eastern Lima, Peru. Multiple-choice questions were used to assess knowledge of TB. A five-item Likert scale was created to assess attitudes toward the community, patients, and clinics. Linear regression was used to identify predictors of mean knowledge score, and analysis of variance was used to test differences in HCP score. Results: Of the 73 HCPs surveyed, 15% were professionals (doctors or nurses). The remaining 85% were health technicians, community health workers (CHWs) or students. The mean knowledge score was 10.0 ± 1.9 (maximum 14) with professional HCPs scoring higher than other HCPs (11.7 ± 1.1 vs. 9.7 ± 1.9), p < .01). Knowledge gaps included identification of patients at high risk for TB, assessment of treatment outcomes, and consequences of treatment failure. The most commonly cited modifiable barriers were structural, including laboratory facilities and staffing of TB clinics, with 52.1% and 62.5% of HCPs, respectively, citing these as problematic. Conclusions: Efforts to improve knowledge of TB HCPs in Peru should focus on the specific gaps we have identified. Further research is needed to evaluate whether these knowledge gaps correlate with TB control. Copyright © 2009 Kiefer et al.