Evaluation of the Rapid MGIT TBc Identification Test for Culture Confirmation of Mycobacterium tuberculosis Complex Strain Detection

Abstract

A culture confirmation test for the detection of Mycobacterium tuberculosis complex strains that uses a lateral-flow immunochromatographic assay to detect the MPB64 antigen, the MGIT TBc identification (TBc ID) test, has been developed. We evaluated the performance of the TBc ID test in the detection of the M. tuberculosis complex in 222 primary-positive liquid cultures. We compared these results to those of nucleic acid-based identification and conventional biochemical tests. The validity of the TBc ID test was determined, and all of the nontuberculous mycobacteria (NTM) and Nocardia species tested were found to be negative. The detection limit of the TBc ID test was 5 ؋ 10 5 CFU/ml, and for IS6110 real-time PCR it was 5 CFU/ml. All of the M. tuberculosis and M. africanum cultures were found to be positive, while M. bovis and M. bovis BCG cultures were negative. With the exception of 1 contaminated culture, the 221 culture-positive isolates contained 171 (77.5%) M. tuberculosis isolates, 39 (17.6%) NTM species, and 11 (5.0%) unidentified species. Two culture-positive isolates harbored a 63-bp deletion at position 196 of the mpb64 gene. The sensitivity, specificity, positive predictive values, and negative predictive values of the TBc ID test were 98.8, 100, 100, and 95.1%, respectively. Furthermore, the approximate turnaround time for real-time PCR was 4 h (including buffer and sample preparation), while for the TBc ID test it was less than 1 h. We suggest an algorithm for the primary identification of M. tuberculosis in liquid culture using the TBc ID test as an alternative to conventional subculture followed by identification using biochemical methods.In 2007, the World Health Organization (WHO) adopted a policy that recommended the use of liquid culture methods for culture and drug susceptibility tests as a standard for tuberculosis (TB) diagnosis and case management (28). The Taiwan Centers for Disease Control (CDC) recommended that liquid and solid media be used simultaneously for mycobacterial culture (7), and approximately 90% of clinical mycobacteriology laboratories in Taiwan use a liquid culture system for the isolation of the Mycobacterium tuberculosis complex from clinical specimens. Acid-fast bacillin (AFB) smear tests then are performed on positive cultures to dismiss contamination (4, 20). The turnaround time (TAT) for the recovery of the M. tuberculosis complex thus is reduced to 10 to 14 days (8, 18). Although the recovery of mycobacteria can be accelerated by using liquid culture systems, this practice provides only partial benefits if it is not accompanied by a rapid species identification test (16). Differentiating M. tuberculosis from nontuberculous mycobacteria (NTM) as soon as possible is important, particularly in situations in which NTM strains represent a considerable share of the clinical isolates.The identification of M. tuberculosis is time-consuming using conventional biochemical methods. The subculturing of isolated mycobacteria from liquid cultures onto solid media and their subs...

Performance of the MGIT TBc Identification Test and Meta-Analysis of MPT64 Assays for Identification of the Mycobacterium tuberculosis Complex in Liquid Culture

“…Meta-analyses of the data, including data from the current study, demonstrated high sensitivity and specificity for all 3 tests, with high positive and negative predictive values for clinical isolates tested (Table 3). There was no evidence that sensitivities and specificities differed between tests even when we confined the meta-analysis to studies that used 16S rRNA sequencing or two or more independent, validated reference methods (4, 12, 16-18 (4)(5)(6)33). The mpb64 gene was sequenced for 27 of 53 (51%) clinically independent isolates for which the MPT64 ICT gave a false-negative result, and a mutation was demonstrated in 23 (85%) of those 27 isolates (12,16,17,20,22,33 Our data and meta-analysis demonstrate noninferiority of MGIT TBcID to LPA for MTBC detection and similar high sensitivities and specificities of 3 commercial MPT64 assays across a range of geographical locations.…”

“…There was no evidence that sensitivities and specificities differed between tests even when we confined the meta-analysis to studies that used 16S rRNA sequencing or two or more independent, validated reference methods (4, 12, 16-18 (4)(5)(6)33). The mpb64 gene was sequenced for 27 of 53 (51%) clinically independent isolates for which the MPT64 ICT gave a false-negative result, and a mutation was demonstrated in 23 (85%) of those 27 isolates (12,16,17,20,22,33 Our data and meta-analysis demonstrate noninferiority of MGIT TBcID to LPA for MTBC detection and similar high sensitivities and specificities of 3 commercial MPT64 assays across a range of geographical locations. A small minority of MTBC isolates are not detected by MPT64 assays due to deletion or mutation of the mpb64 gene (absent from some M. bovis BCG strains) (1,12,16,17,20,22,33) or to low MPT64 concentrations in early cultures (10,18) or mixed cultures (10,18).…”

Seek TB, a Two-Stage Multiplex Real-Time-PCR-Based Method for Differentiation of the Mycobacterium tuberculosis Complex

“…Both molecular methods demonstrated 100% agreement, identifying 84 isolates as M. tuberculosis isolates and 41 isolates as non-MTC isolates. The results of TBc ID (with a reported specificity and sensitivity of 92.4 to 100% and 94.9 to 100%, respectively [18,25,29]) and the molecular methods were concordant for 79 MTC isolates and 37 non-MTC isolates. The ssrA genes of the 9 discordant samples were sequenced to consolidate results.…”

Clinical Laboratory Diagnostics for Mycobacterium tuberculosis

“…These tests are rapid requiring only minutes to perform after growth of the organism but literature reports indicate variable performance (Martin et al, 2011;Said et al, 2011;Steingart et al, 2011;Yu et al, 2011) 6. Direct identification of M. tuberculosis from clinical specimens…”