DETECTION OF MYCOBACTERIUM-TUBERCULOSIS IN RESPIRATORY SPECIMENS BY STRAND DISPLACEMENT AMPLIFICATION OF DNA

A total of 294 clinical respiratory specimens, including 75 with cultu re-positive results, were tested for the presence of Mycobacterium tub erculosis by strand displacement amplification (SDA) of DNA. A region of the IS6110 insertion element and an internal control sequence were amplified and then detected by a chemiluminescence assay. Receiver ope rator-characteristic curves were used to evaluate three methods for de claring specimens positive for M. tuberculosis. By the preferred metho d, SDA chemiluminescence results were converted to theoretical numbers of M. tuberculosis organisms. A positive threshold (PT) value, above which 95% of the SDA results were judged to be M. tuberculosis positiv e (sensitivity = 95%), was found to be 2.4 M. tuberculosis organisms p er SDA reaction. The analogous PT value for 95% sensitivity on smear-p ositive specimens was 3.6 M. tuberculosis organisms per reaction. The PT of 2.4 M. tuberculosis organisms per reaction detected 100% of cult ure-positive, smear-positive specimens (sensitivity = 100%), while 95% sensitivity was achieved with a PT of 15.5 M. tuberculosis organisms per reaction, Specificities, which were calculated with respect to cul ture- and smear-negative specimens, ranged from 96% at a PT of 15.5 M. tuberculosis organisms to 84% at a PT of 2.4 M. tuberculosis organism s per reaction. The M. tuberculosis-negative specimens were also segre gated according to whether the patients received antituberculosis chem otherapy. SDA specificity ranged from 90% (PT = 2.4 M. tuberculosis or ganisms) to 98% (PT = 15.5 M. tuberculosis organisms) for the M. tuber culosis-negative specimens from patients who had not received chemothe rapy. SDA specificity in the M. tuberculosis-negative specimens from p atients who received chemotherapy was lower (85 to 94%). This study re presents the first large-scale demonstration of M. tuberculosis detect ion in clinical sputum specimens by isothermal DNA amplification with SDA.