The utility of polymerase chain reaction (PCR) in the diagnosis of pulmonary tuberculosis

Department of Microbiology, University Hospital and Medical School, Valencia, Spain.
Chest (Impact Factor: 7.48). 07/1995; 107(6):1631-5. DOI: 10.1378/chest.107.6.1631
Source: PubMed


A fragment of DNA of 123 bp belonging to insertion sequence IS6110, specific of Mycobacterium tuberculosis complex, was amplified by polymerase chain reaction (PCR) of respiratory samples, for the diagnosis of pulmonary tuberculosis. A total of 314 samples (286 sputum and 28 bronchoalveolar lavages) from 242 patients were evaluated by PCR, and the results were compared with the those obtained by acid-fast-stained smears, culture, and clinical diagnosis. Mycobacterium tuberculosis was detected by PCR in 102 of 105 patients with clinical diagnosis of pulmonary tuberculosis. All smear and culture-positive samples were PCR positive. The sensitivity of PCR, culture, and staining was 97%, 88%, and 65%, respectively, and the specificity was 100% in all cases. In ten patients with old residual lesions, but no active disease, M tuberculosis genome was detected by PCR. In our experience, PCR proved to be a useful method for the rapid diagnosis of pulmonary tuberculosis.

Full-text preview

Available from:
  • Source
    • "Kocagoz et al. (1993) recorded that sensitivity, specificity and positive and negative predictive values of ZN smear were respectively 68, 100, 100 and 70 %. Similar results were reported by Querol et al. (1995), who found a microscopic sensitivity of 65 % and specificity of 100 %. Also, Herrera & Segovia (1996) recorded that, considering clinical diagnosis as the 'gold standard', the sensitivity, specificity and positive and negative predictive values for ZN smear were respectively 69, 87, 84 and 74 %. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Rapid, sensitive and low-cost methods are needed urgently for the detection of Mycobacterium tuberculosis in clinical samples, especially in developing countries. To this end, the clinical performance of FASTPlaqueTB(TM) (a bacteriophage-based method) has been studied in parallel with microscopy, standard microbiological culture and in-house IS6110-based PCR methods. A total of 64 samples, including 42 sputum samples and 22 urine samples, were tested in this study. The sensitivity, specificity and overall accuracy values for the FASTPlaqueTB assay relative to that of culture were respectively 76.5, 95 and 90 %. The corresponding values for the in-house IS6110-based PCR assay were 88, 91 and 90 % and, for Ziehl-Neelsen staining, were 59, 95 and 85 %. FASTPlaqueTB gave better clinical performance with urine samples than with sputum samples (sensitivity, specificity and overall accuracy were 100 % with urine samples and 64, 93 and 84 % with sputum samples). The 100 % sensitivity of FASTPlaqueTB was higher than that of the corresponding values for PCR (67 %) with urine samples. In conclusion, FASTPlaqueTB proved to be sensitive, cheap relative to the PCR and rapid. It is able to detect M. tuberculosis in clinical samples within 1 day, reducing the time to diagnosis in comparison with culture.
    Journal of Medical Microbiology 05/2003; 52(Pt 4):331-5. · 2.25 Impact Factor
  • Source
    • "Noordhoek et al. (1995) observed that phenol extraction of DNA removes inhibiting substances from those samples in which inhibitors were present even after DNA extraction with guanidinium thiocyanate (GuSCN) and silica particles. Querol et al. (1995) achieved 97% PCR positivity by using phenol:chloroform:iso amyl alcohol extraction followed by isopropanol precipitation of DNA. Similarly, some workers (Sjobring et al., 1990; Boddinghaus et al., 1990) reported that the use of phenol and chloroform for extraction and ethanol and/or isopropanol for precipitation of DNA surely improves the yield of the purified target DNA, which finally results in increased sensitivity of PCR. "
    [Show abstract] [Hide abstract]
    ABSTRACT: We have compared the efficacy of various reported protocols of mycobacterial DNA extraction for detection of mycobacterial DNA by PCR assay. Seven DNA extraction protocols were tested for their quantitative as well as qualitative yield of mycobacterial DNA in 15 known positive sputum samples having occasional acid fast bacilli (AFB). DNA samples obtained by various methods were amplified in uniform standard conditions and analysed on 3% agarose gel. Protocol 6 and 7 showed 100% detection sensitivity with strong bands on agarose gel. Protocols 1-5 were found to be unsatisfactory because they yielded either low quantity or poor quality of DNA or were unable to remove inhibitors of DNA amplification. We conclude that a strong physical treatment, use of a detergent and enzyme for lysis, treatment with proteinase K, DNA purification step with or without phenol and DNA precipitation in ethanol or isopropanol are essential steps for extraction of mycobacterial DNA from clinical samples. Protocol 6 is standard in our laboratory and we have found reproducible results with this method. Purification with phenol followed by chloroform treatment was not found to have any inhibitory effect on amplification. A more extensive evaluation of this protocol in samples with lower bacterial load may be necessary.
    Molecular Biology Today 01/2002; 3(2).
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Tuberculosis, caused by Mycobacterium tuberculosis, is responsible for over two million deaths per year worldwide. Due to its long doubling time (18 h), the microbiological detection of M. tuberculosis by conventional methods takes up to one month, unless the number of bacilli in the biological sample is high enough. Thus, drug resistance assessment requires at least one month for obtaining the primary culture and another month to determine its susceptibility to antimycobacterial drugs. Moreover, for a long time, the lack of genetic tools for mycobacteria has been a barrier for undertaking studies aimed at understanding the mechanisms of drug resistance and drug target identification, being all these topics of utmost importance considering the increase in the number of drug-resistant clones and the few therapeutic options available. Mycobacteriophages are promising as a novel source of genetic elements for mycobacteria manipulation, as well as for the development of versatile, simple, fast and cheap methods for drug resistance assessment of M. tuberculosis clinical isolates. We herein describe the background related to the use of mycobacteriophages, with emphasis placed on their utilization for drug resistance analysis in our country.
    Revista Argentina de microbiología 01/2009; 41(1):45-55. · 0.80 Impact Factor
Show more