The prevalence of latent tuberculosis infection (LTBI) in the various populations of New York City (NYC), a city with a high density of non-US-born persons, is unknown. We examined the prevalence of TST positivity in patients who received a tuberculin skin test (TST) between 1/2002 and 8/2004 at any of 10 NYC health department chest centers. A positive TST was defined as an induration reaction to tuberculin of ≥10 mm. In the study population of 41,022 individuals, prevalence of TST positivity was 24.4% (95%CI = 24.0, 24.8); four times higher among non-US-born persons than US-born (39.5% vs. 8.8%, Prevalence ratio (PR) = 4.5; 95%CI = 4.4, 4.6). Prevalence of TST positivity increased with age in both US and non-US-born persons. Persons from countries with a TB case rate >100/100,000 population had higher prevalence of TST positivity (47% vs. ≤39%), even after controlling for BCG (PR = 1.3, 95%CI = 1.2, 1.4). These findings provide insight into current prevalence of TST positivity in many immigrant populations and will help both clinicians and health departments to target patients for LTBI treatment.