title: Early detection of Mycobacterium tuberculosis complex in BACTEC MGIT cultures using nucleic acid amplification.
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.
authors.affiliation: Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, 100 Tzyou 1st Road, Kaohsiung, Taiwan.|Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: S Y
authors.initials: SY
authors.lastname: Lin
authors.affiliation: Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.|Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: S C
authors.initials: SC
authors.lastname: Hwang
authors.affiliation: Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Y C
authors.initials: YC
authors.lastname: Yang
authors.affiliation: Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: C F
authors.initials: CF
authors.lastname: Wang
authors.affiliation: Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, 100 Tzyou 1st Road, Kaohsiung, Taiwan.|Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Y H
authors.initials: YH
authors.lastname: Chen
authors.affiliation: Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, 100 Tzyou 1st Road, Kaohsiung, Taiwan.|Kaohsiung Municipal Ta-Tung Hospital, Kaohsiung Medical University Hospital, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan.
authors.forename: T C
authors.initials: TC
authors.lastname: Chen
authors.affiliation: Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, 100 Tzyou 1st Road, Kaohsiung, Taiwan. idpaul@gmail.com.|Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. idpaul@gmail.com.|School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan. idpaul@gmail.com.
authors.forename: P L
authors.initials: PL
authors.lastname: Lu
mesh_terms: D015373:Bacterial Typing Techniques; D042241:Early Diagnosis; D005798:Genes, Bacterial; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000145:classification / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014376:Tuberculosis / Q000175:diagnosis* / Q000382:microbiology*