Introduction:Tuberculosis (TB) is an airborne communicable disease caused by Mycobacterium tuberculosis. Infection is most likely to occur when an individual is exposed to someone with pulmonary TB on a day-to-day basis, such as by living or working in close quarters with someone who has an active disease. This study was undertaken using reliable screening and investigation tools for assessing the incident latent tuberculosis infection (LTBI) among healthcare workers (HCWs).Methodology:This descriptive longitudinal study was conducted among 600 HCWs at a tertiary care hospital in Bengaluru city for a period of 1 year 4 months (March 2016–June 2017). All HCWs were stratified according to their work profile as doctors, nurses, laboratory technicians, nursing aides, pharmacists, laundry workers (only those who come in contact with patients), dietary workers (only those who come in contact with patients), and workers from housekeeping department. Study subjects were enrolled in proportion to their population size. The data were collected by administering TB screening questionnaire and performing tuberculin skin test (TST). All the subjects who had negative TST reaction were followed up after 1 year from March 2017 to June 2017.Results:The study subjects consisted of 600 HCWs with a mean age of 27.8 ± 7.1 years. Initially, 600 study subjects underwent TST, 2 of whom were not contactable between 48 and 72 h of test administration. Of the remaining 598 subjects, 120 (20.1%) had a positive TST reaction. After 1 year, of the 478 subjects who tested negative, 345 study subjects underwent the repeat TST and 67 (19.4%) of the study subjects had a positive TST reaction.Conclusion:The prevalence of LTBI was 20.1% and it was significantly associated with age, place of residence, education, work commute time, and mode of transport. The incidence of LTBI was 19.4% and it was significantly associated with gender, place of residence, education, work commute time, and marital status.