The objective of this work was to address the problem of tuberculosis (TB) infection among Health Care Workers (HCW), based on an experience in a TB Prevention and Treatment Programme at a General Acute Hospital in Buenos Aires City. The objectives of the programme were to provide health education and periodic screening of HCW, to detect TB cases and to implement control measures. TB infection was defined by a 10 mm reaction to 2 UT PPD. Two prevalence studies of TB infection were performed. In 1998 infection prevalence was 31.7% (n = 533), and in 2008 it was 35% (n = 670). Nurses presented the highest percentage. An increase in TB infection prevalence was observed in 2008 among physicians (p = 0.02). A sample of 45 HCW integrating the first prevalence study group was analysed by applying 2 UT PPD three month later. Eight tuberculin conversions were detected (17%), all of them in nurses; 163 HCW who had contact with other eight HCW who developed TB were evaluated, four of whom were nurses. PPD reactions were positive in 39.9% with no any new case of TB detected among them. In nurses, five tuberculin conversions were observed. Other 127 HCW from low risk areas were also evaluated, 25.9% of whom had positive reactions to 2 UT PPD, while this percentage was 39.9% in the previous group (p = 0.01).Despite the scarcity of human resources that limit the planning of TB incidence studies, personal protective measures, teaching HCW about TB and chemoprophylaxis might reduce the risk of TB in this group.