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Early detection of Mycobacterium tuberculosis complex in BACTEC MGIT cultures using nucleic acid amplification

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Abstract

We evaluated the application of nucleic acid amplification (NAA) in liquid cultures for the early detection of Mycobacterium tuberculosis. The Cobas TaqMan MTB test, IS6110 real-time PCR, and hsp65 PCR-restriction fragment length polymorphism (RFLP) analysis were used to detect BACTEC MGIT 960 (MGIT) cultures on days 3, 5, 7, and 14. The procedure was initially tested with a reference strain, H37Rv (ATCC 27294). Subsequently, 200 clinical specimens, including 150 Acid Fast bacillus (AFB) smear-positive and 50 AFB smear-negative samples, were examined. The Cobas TaqMan MTB test and IS6110-based PCR analysis were able to detect M. tuberculosis after 1 day when the inoculum of H37Rv was >3 x 10−2 CFU/ml. After a 5-day incubation in the MGIT system, all three NAA assays had a positive detection regardless of the inoculum size. After a 1-day incubation of the clinical specimens in the MGIT system, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for the Cobas TaqMan MTB assay were 70.2%, 100%, 100%, and 82.3% respectively. For IS6110-based PCR analysis, these values were 63.1%, 100%, 100%, and 78.9%, and were 88.1%, 100%, 100%, and 92.1% respectively for hsp65 PCR-RFLP analysis. After a 3-day incubation, the specificity and PPV were 100% for all three NAA tests; the Cobas TaqMan MTB assay had the best sensitivity (97.6%) and NPV (98.3%). The sensitivity, specificity, PPV, and NPV for conventional culture analysis were 98.8%, 100%, 100%, and 99.1%. Thus, NAA may be useful for the early detection of M. tuberculosis after 3 days in MGIT.

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Acknowledgments

The authors thank colleagues at the Mycobacteriology Section of Clinical Microbiology Laboratory, Kaohsiung Medical University Hospital, for their support

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Authors and Affiliations

  1. Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, 100 Tzyou 1st Road, Kaohsiung, Taiwan

    S. Y. Lin, Y. H. Chen, T. C. Chen & P. L. Lu

  2. Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan

    S. C. Hwang, Y. C. Yang, C. F. Wang & P. L. Lu

  3. Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan

    S. Y. Lin, S. C. Hwang & Y. H. Chen

  4. School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan

    P. L. Lu

  5. Kaohsiung Municipal Ta-Tung Hospital, Kaohsiung Medical University Hospital, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    T. C. Chen

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  1. S. Y. Lin
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  2. S. C. Hwang
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  4. C. F. Wang
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  7. P. L. Lu
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Correspondence to P. L. Lu.

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Conflict of Interest

The authors declared no conflict of interest.

Funding

This work is supported by Kaohsiung Medical University Hospital (KMUH101-1R13).

Ethical approval

This study was approved by the Institutional Review Board (IRB) of Kaohsiung Medical University Chung-Ho Memorial Hospital, Kaohsiung, Taiwan (KMUH-IRB-970369).

Informed consent

The study subjects were mycobacterial culture, and the written consent given by the patients was waived by the approving IRB.

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Lin, S.Y., Hwang, S.C., Yang, Y.C. et al. Early detection of Mycobacterium tuberculosis complex in BACTEC MGIT cultures using nucleic acid amplification. Eur J Clin Microbiol Infect Dis 35, 977–984 (2016). https://doi.org/10.1007/s10096-016-2625-9

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  • DOI: https://doi.org/10.1007/s10096-016-2625-9

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