Identifying persons infected with both human immunodeficiency virus (HIV) and Mycobacterium tuberculosis (MTB) is often difficult because of the reduced sensitivity of tuberculin skin testing in HIV-infected persons. To determine the value of two-step tuberculin skin testing (TTST) as a method of increasing the sensitivity of tuberculin skin testing in HIV-infected persons, a consecutive sample of 58 HIV-infected persons being screened for a TB preventive therapy trial in Uganda with an initial purified protein derivative (PPD) response < 5 mm completed two-step tuberculin and candida skin testing. The mean change in PPD size between the two tests, placed a mean of 8 d apart, was +2.1 mm (SD 4.4 mm, range -4 to +16 mm). Seventeen subjects (29%) had a boosted response (PPD1 < 5 and PPD2 > or = 5). In a multiple logistic regression model, boosted responses were independently associated with a CD4 count between 200 and 500 microl(-1) (p = 0.02) and a higher body mass index (p = 0.05). TTST may be valuable in identifying MTB infection and in preventing misclassification of boosted responses as skin test conversions in HIV-infected persons, especially persons with CD4 counts between 200 and 500 microl(-1) from areas with a high prevalence of MTB infection or from areas with a low prevalence of MTB infection who have other risk factors for MTB infection.