DIAGNOSIS OF SMEAR-NEGATIVE PULMONARY TUBERCULOSIS USING SEQUENCE CAPTURE POLYMERASE CHAIN-REACTION

Techniques based on the polymerase chain reaction (PCR) can he used to rapidly identify DNA from Mycobacterium tuberculosis in clinical samp les from patients with tuberculosis, but prior studies evaluating this approach in the diagnosis of paucibacillary forms of pulmonary tuberc ulosis have reported poor sensitivity and/or specificity. We have deve loped a procedure in which mycobacterial DNA in crude samples is speci fically captured prior to amplification, thereby concentrating the tar get sequences and removing irrelevant DNA and other inhibitors of the amplification reaction (sequence capture PCR). To evaluate the usefuln ess of this approach in the diagnosis of paucibacillary forms of pulmo nary tuberculosis, sequence capture PCR was performed prospectively on samples of bronchoalveolar lavage fluid from consecutive patients sus pected of having pulmonary tuberculosis but for whom three consecutive samples of respiratory secretions were smear negative. Of the 27 pati ents evaluated, active tuberculosis was diagnosed in nine; sequence ca pture PCR was positive for ail of these patients, including the three for whom all specimens submitted for culture were negative. No positiv e results were obtained for lavage fluid from the 18 patients for whom the diagnosis of active tuberculosis was subsequently excluded or 25 additional patients undergoing bronchoalveolar lavage for evaluation o f other pulmonary problems, even though many of these patients had a h istory of prior tuberculosis or radiographic evidence of prior tubercu lous infection. Paucibacillary forms of pulmonary tuberculosis can be rapidly identified with high sensitivity and specificity using sequenc e capture PCR performed on samples obtained by bronchoalveolar lavage.