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J Clin Microbiol. 1993 August; 31(8): 2049-2056

Large-scale use of polymerase chain reaction for detection of Mycobacterium tuberculosis in a routine mycobacteriology laboratory.

J E Clarridge 3rd, R M Shawar, T M Shinnick and B B Plikaytis

Laboratory Service Veterans Affairs Medical Center, Houston, Texas.

ABSTRACT

We investigated the use of DNA amplification by the polymerase chain reaction reaction (PCR) for detection of Mycobacterium tuberculosis from clinical specimens. Two-thirds of each sample was processed for smear and culture by standard methods, and one-third was submitted for DNA extraction, amplification of a 317-bp segment within the insertion element IS6110, and detection by agarose gel electrophoresis, hybridization, or both. DNA was prepared from over 5,000 samples, with 623 samples being culture positive for acid-fast bacilli. Of 218 specimens that were identified as M. tuberculosis, 181 (85%) were positive by PCR. In the M. tuberculosis culture-positive group, PCR was positive for 136 of 145 (94%) and 45 of 73 (62%) of the fluorochrome smear-positive and -negative specimens, respectively. Of 948 specimens that were either culture positive for mycobacteria other than M. tuberculosis or culture negative, 937 specimens were negative by PCR and 11 (1%) specimens initially appeared to be false positive for M. tuberculosis. The reason for discrepant results varied; some errors were traced to the presence of an inhibitor in the specimen (7.3% in unselected specimens), nucleic acid contamination, low numbers of organisms in the specimen antituberculosis therapy, and possible low-level nonspecific hybridization. In comparison with culture, the sensitivity, specificity, and positive predictive value were 83.5, 99.0, and 94.2%, respectively, for PCR. When PCR was corrected for DNA contamination, the presence of inhibitor, and culture-negative disease, the values became 86.1, 99.7, and 98.4%, respectively. If the results for multiple specimens submitted from the same patient are considered, no patient who had three of more sputum specimens tested would have been misdiagnosed.

J Clin Microbiol. 1993 August; 31(8): 2049-2056




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