Mycobacteriology
Evaluation of the BDProbeTec ET system as screening tool in the direct detection of mycobacterium tuberculosis complex in respiratory specimens

https://doi.org/10.1016/S0732-8893(03)00163-9Get rights and content

Abstract

We evaluated the BDProbeTec ET System (Becton Dickinson) for the routine detection of Mycobacterium tuberculosis complex (MTC) in respiratory specimens and pleural fluids, comparing with microscopy (Ziehl Neelsen stain, ZN) and culture in liquid (BACTEC MGIT 960, MGIT) and solid (Löwenstein Jensen, LJ) media. Five hundred and two specimens, collected from 266 patients, of which 257 with suspected tuberculosis and 9 receiving anti-tuberculosis treatment, were investigated. Thirty-nine specimens were positive by any method, including false positives. Mycobacteria were isolated from 33 specimens (32 Mycobacterium tuberculosis and 1 Mycobacterium chelonae). Thirty-six specimens were BDProbeTec ET positive, 33 specimens were MGIT positive, 27 were LJ positive and 22 were ZN positive. With BDProbeTec ET, 2 specimens were false negative (culture positive), and 2 specimens from non-treated patients were false positive (culture negative). The overall sensitivity, specificity, and positive and negative predictive values for BDProbeTec ET compared to culture were 93.7, 98.7, 83.3, and 99.5%, respectively, while with smear-positive and smear-negative specimens the sensitivities were 100% and 81.5% respectively. In five treated patients the disappearance of MTC could be monitored using BDProbeTec ET in parallel with culture. The overall inhibition rate was 0.2%. BDProbeTec ET can be very useful for rapid detection of MTC, especially in smear-negative respiratory specimens.

Introduction

Tuberculosis (TB) is an increasing health problem worldwide, especially in developing countries. The spread of HIV/AIDS and the emergence of multidrug-resistant TB are contributing to the worsening impact of this disease. Up to one third of the world's population is estimated by the World Health Organization (WHO) to be infected with Mycobacterium tuberculosis and it is estimated that between 2002 and 2020, approximately 1 billion people will be newly infected, over 150 million people will develop the disease, and 36 million people will die of TB if control is not further strengthened (World Health Organization, 2002).

TB control is based on early detection by acid-fast bacilli (AFB) through AFB stain and culture of mycobacteria using liquid and/or solid media. AFB smear results are available in hours or less but the technique has poor sensitivity and can not distinguish among different species of mycobacteria (Peterson et al., 1999). Although culture is the gold standard with excellent sensitivity, while the liquid media have significantly reduced the detection time, it requires on the average 2-3 weeks to obtain results for slow growing mycobacteria and up to 6 weeks to finalize negative specimens. Because of the low sensitivity of AFB microscopy it is possible that in some cases tuberculosis will not be early detected, which can cause the patient to become an important source of transmission, particularly if culture is not performed. Tuberculosis transmission from smear negative patients is estimated to be approximately 17% (Behr et al., 1999).

Methods based on direct nucleic acid testing (NAT) are more sensitive than microscopy and have reduced the time to detection of Mycobacterium tuberculosis complex (MTC). Polymerase chain reaction (PCR), transcript-mediated assay (TMA), and ligase chain reaction (LCR) were used as molecular tools in the diagnosis of tuberculosis Bergmann and Woods, 1996, Dalovisio et al., 1996, Ichiyama et al., 1996, Moore and Curry, 1995, Wobeser et al., 1996, Bergmann et al., 1999, Gamboa et al., 1998, Piersimoni et al., 1998, Bennedsen et al., 1996, Jonas et al., 1993, Jungkind et al., 1996, Lindbrathen et al., 1997, Pfyffer et al., 1996. Strand Displacement Amplification (SDA) is an isothermal method for the detection of IS6110, a specific target for MTC Ichiyama et al., 1997, Pfyffer et al., 1999. The BD ProbeTec ET system (Becton Dickinson) uses SDA in combination with real time fluorescence detection of amplified product, thus making results available in a few hours (Little et al., 1999). The BD ProbeTec ET system is approved for direct detection of MTC DNA (DTB test) from decontaminated, digested clinical respiratory specimens such as sputa, induced sputa, bronchial washings, and other respiratory specimens. The DTB test is not available in the USA but is commercially available for diagnostic purposes in Europe and recently in Asia. We have evaluated the BD ProbeTec ET System for the direct detection of MTC in respiratory specimens from patients with suspected TB.

Section snippets

Materials and methods

A total of 502 clinical specimens (435 sputa, 39 broncheoalveolar lavages (BAL), 27 pleural fluids and one pulmonar aspirate) from 266 patients (257 with suspected tuberculosis and 9 receiving anti-tuberculosis treatment), were included in the study. A variable number of specimens per patient was received: more than three per patient in 13 cases, three in 72 cases, two in 43 cases, and only one in 138 cases. Specimens were tested at the Clinical Microbiology Laboratory, Puerto Real University

Results

Thirty-nine from 502 respiratory specimens collected from 23 patients were positive with any method (ZN microscopy, culture on LJ slant, culture in MGIT broth, and/or BDProbeTec ET). Only one specimen, which was negative in all methods, was inhibited in BDProbeTec ET. Results are summarized in Table 1. Twenty-two specimens were ZN positive. Mycobacteria grew from 33 specimens (32 Mycobacterium tuberculosis and 1 Mycobacterium chelonae); 32 MTC isolates grew in MGIT liquid medium and 26 grew on

Discussion

SDA is a molecular technique based on isothermal amplification of DNA, using a two-enzyme system (restriction enzyme and DNA polymerase) (Walker et al., 1992). More recently, new thermophilic restriction endonuclease (BsoBI) and DNA polymerase (exo'Bca) were incorporated (Spargo et al., 1996), as well as fluorescence polarization detection (Walker et al., 1996) which was applied to the diagnosis of Mycobacterium tuberculosis using the IS6110 insertion sequence (Walker & Linn, 1996). Down et al.

Acknowledgements

We are indebted to André De Bock, Yolanda Gonzalo, and Fernando García (Becton Dickinson) for their valuable help in the preparation of the manuscript. We thank Becton Dickinson (Madrid, Spain) for providing reagents and instrumentation. This study was partially presented at the 42nd ICAAC, San Diego, CA, 2002.

References (35)

  • J.R. Dalovisio et al.

    Comparison of the amplified Mycobacterium tuberculosis (M. T.B) direct test, Amplicor M. T.B. P. C.R., and IS6110-P. C.R. for detection of MTB in respiratory specimens

    Clin Infect Dis

    (1996)
  • J.A. Down et al.

    Detection of Mycobacterium tuberculosis in respiratory specimens by strand displacement amplification of DNA

    J Clin Microbiol

    (1996)
  • F. Gamboa et al.

    Comparative evaluation of initial and new versions of the Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for direct detection of Mycobacterium tuberculosis in respiratory and nonrespiratory specimens

    J Clin Microbiol

    (1998)
  • S. Ichiyama et al.

    Evaluation of Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test and Roche PCR-microwell plate hybridization method (AMPLICOR MYCOBACTERIUM) for direct detection of mycobacteria

    J Clin Microbiol

    (1996)
  • S. Ichiyama et al.

    Diagnostic value of the strand displacement amplification method compared to those of Roche Amplicor PCR and culture for detecting mycobacteria in sputum samples

    J Clin Microbiol

    (1997)
  • I.S. Johansen et al.

    Evaluation of a new commercial assay for diagnosis of pulmonary and nonpulmonary tuberculosis

    Eur J Clin Microbiol Infect Dis

    (2002)
  • V. Jonas et al.

    Detection and identification of Mycobacterium tuberculosis directly from sputum sediments by amplification of rRNA

    J Clin Microbiol

    (1993)
  • Cited by (10)

    • Performance of the BD ProbeTec ET direct detection assay for the analysis of Mycobacterium tuberculosis in respiratory and non-respiratory clinical specimens

      2017, Journal of Taibah University Medical Sciences
      Citation Excerpt :

      Sensitivity of DTB as low as 63.2% has also been reported14 which appears to be lower than a number of other studies reporting sensitivity of the DTB ranging from 82.7% to 100% for the detection of MTBC in respiratory specimens.15,16 The findings of this study are consistent with previous reports of DTB being a highly sensitive and specific test with a higher negative predictive value of 99.5%,17 making it a powerful tool for diagnosis or exclusion of tuberculosis in respiratory specimens. Despite its excellent performance in respiratory samples, the performance of the DTB in extra-pulmonary specimens, particularly in tissue samples from extra-pulmonary sites was not comparable.

    View all citing articles on Scopus
    View full text