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Healthcare workers (HCWs) have a higher risks of contracting tuberculosis (TB) than general population. International and national policies recommend routine screening of latent tuberculosis infection (LTBI) as an essential component in the control and prevention of TB in healthcare facilities.From January 2008 to December 2016, 1759 hospital employees were screened for LTBI. Symptom assessment and chest X-ray were conducted to exclude active TB, and tuberculin skin test (TST) and/or QuantiFERON-TB Gold test (QFT) were performed to diagnose LTBI.At the end of 2016, 1054 active workers were screened one or more times, totalizing 1810 screenings; 81.5% were female and 18.5% were male; mean age was 42 years. None were found to have active TB. LTBI prevalence in the screened population was 17.7% (n=187): 101 individuals had a QTF positive test and 86 didn’t perform QTF test but had a TST ≥15 mm. The majority were positive for LTBI at the first screening (n=110; 58.8%). Among the screened HCWs, medical aid assistants had the highest prevalence of LTBI (21.7%), followed by nurses (19.4%), administrative and supportive staff (14.4%), while physicians had the lowest prevalence (12.4%) of LTBI. QFT was negative in 47.3% of the individuals with TST ≥15 mm (n=61, which 41 submitted to repetitive TST testing), and in 76.5% of the cases with TST ≥10 mm but <15 mm (n=65, which 37 submitted to repetitive TST testing).Since 2015, Portugal has been a low-incidence country regarding TB. The prevalence of LTBI in HCWs is relatively high as far as 17,7%. As a result, active screening for TB and LTBI is needed for these workers. Screening with TST and QTF is a cost-effective approach as high numbers of discordant TST positive/QTF negative results are probably caused by BCG vaccination or boosting due to repetitive TST testing.