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The time course of persistence of Mycobacterium tuberculosis as measured by detection of rRNA, acid-fast bacillus (AFB) smear, and culture was determined for pulmonary tuberculosis patients during antimicrobial therapy. Twenty-three patients who were initially AFB smear positive and who subsequently completed a course of antimicrobial therapy were selected for the study. Sequential specimens were tested by AFB smear, culture, and rRNA amplification (Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test [MTD]). The initial diagnostic specimens of all patients were positive by culture; those of 22 patients (96%) also were positive by MTD. Overall, MTD results remained positive longer than both smear and culture results. The median times to the last positive test result were 9 days for AFB smear, 26 days for culture, and 30 days for MTD. The last positive test result was the AFB smear result in 4% of cases, the culture result in 22%, and the MTD result in 52%. Fifty-six percent of patients had a period of shedding of noncultivable M. tuberculosis which was detected by MTD after culture results had converted to negative. This noncultivable period lasted 7 to 245 days. All three tests became reproducibly negative before the end of therapy and remained negative during follow-up for up to 1 year. These results indicate that during successful antimicrobial therapy, M. tuberculosis is eliminated in sputum samples as measured by amplification of rRNA, as well as by AFB smear and culture. No long-term rRNA carrier state was detected. While the time course of clearance of M. tuberculosis measured by rRNA overall was longer than with the two traditional tests, the rRNA test results allow sensitive and precise measurement of the clearance of noncultivable M. tuberculosis from respiratory specimens. This attribute may allow rRNA testing to be useful in clarifying patient response to antimicrobial therapy.

Seven hundred fifty-eight processed sputum sediments received for the diagnosis of tuberculosis or other mycobacterial infections were tested by utilizing a rRNA target amplification assay and traditional culture techniques. The results from the rRNA target amplification assay (Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test), available in 5 h, were compared with the results from standard culture techniques held for 6 weeks. A total of 119 specimens (16%) were culture positive for Mycobacterium tuberculosis. Overall sensitivity, specificity, positive predictive value, and negative predictive value were 82, 99, 97, and 96%, respectively, for the Gen-Probe assay; 88, 100, 100, and 97%, respectively, for culture; and 53, 99.8, 99.6, and 91%, respectively, for fluorochrome stain. The Gen-Probe assay employs the isothermal enzymatic amplification of M. tuberculosis complex rRNA followed by detection of the amplicon with an acridinium ester-labeled DNA probe. This assay has the potential of reducing the time for diagnosis of tuberculosis to 1 day.