[Detection of Mycobacterium tuberculosis in sputum specimens of pulmonary tuberculosis by DNA amplification]

The PCR was used to detect M. tuberculosis DNA sequences in uncultured clinical specimens. Two oligonucleotide primers with 20 bp each amplified target template DNA of M. tuberculosis. Amplified DNA product was 245 bp which was identified by agarose gel electrophoresis. The sensitivity of detection of M. tuberculosis genomic DNA and bacteria suspension by PCR was lpg and 13 viable bacteria cell/ml, respectively. In specificity experiments, only M. tuberculosis, M. bovis and BCG were positive by PCR, but all other 14 Mycobacterium tested, including streptomyces lividans and E. coli plasmid PUC19 were negative. The sensitivity of detection of M. tuberculosis by PCR was determined by comparing the fast-acid staining and culture on total 54 sputum specimens of pulmonary tuberculosis and 12 nontuberculosis lung disease. The positive rate of PCR in pulmonary tuberculosis were 37.0%, culture method showed only 14.8%, fast-acid staining were 16.7%. Nontuberculosis lung disease were negative. The results show that DNA amplification is a superior method with high degree of sensitivity and specificity for rapid diagnosis of pulmonary tuberculosis.