Comparison of Quantitative Techniques including Xpert MTB/RIF to Evaluate Mycobacterial Burden
Open Access
- 22 December 2011
- journal article
- research article
- Published by Public Library of Science (PLoS) in PLOS ONE
- Vol. 6 (12), e28815
- https://doi.org/10.1371/journal.pone.0028815
Abstract
Accurate quantification of mycobacterial load is important for the evaluation of patient infectiousness, disease severity and monitoring treatment response in human and in-vitro laboratory models of disease. We hypothesized that newer techniques would perform as well as solid media culture to quantify mycobacterial burden in laboratory specimens. We compared the turn-around-time, detection-threshold, dynamic range, reproducibility, relative discriminative ability, of 4 mycobacterial load determination techniques: automated liquid culture (BACTEC-MGIT-960), [3H]-uracil incorporation assays, luciferase-reporter construct bioluminescence, and quantitative PCR(Xpert -MTB/RIF) using serial dilutions of Mycobacterium bovis and Mycobacterium tuberculosis H37RV. Mycobacterial colony-forming-units(CFU) using 7H10-Middlebrook solid media served as the reference standard. All 4 assays correlated well with the reference standard, however, bioluminescence and uracil assays had a detection threshold ≥1×103 organisms. By contrast, BACTEC-MGIT-960 liquid culture, although only providing results in days, was user-friendly, had the lowest detection threshold (<10 organisms), the greatest discriminative ability (1 vs. 10 organisms; p = 0.02), and the best reproducibility (coefficient of variance of 2% vs. 38% compared to uracil incorporation; p = 0.02). Xpert-MTB/RIF correlated well with mycobacterial load, had a rapid turn-around-time (<2 hours), was user friendly, but had a detection limit of ∼100 organisms. Choosing a technique to quantify mycobacterial burden for laboratory or clinical research depends on availability of resources and the question being addressed. Automated liquid culture has good discriminative ability and low detection threshold but results are only obtained in days. Xpert MTB/RIF provides rapid quantification of mycobacterial burden, but has a poorer discrimination and detection threshold.Keywords
This publication has 24 references indexed in Scilit:
- Evaluation of the Xpert MTB/RIF Assay for the Diagnosis of Pulmonary Tuberculosis in a High HIV Prevalence SettingAmerican Journal of Respiratory and Critical Care Medicine, 2011
- Using Bioluminescence To Monitor Treatment Response in Real Time in Mice with Mycobacterium ulcerans InfectionAntimicrobial Agents and Chemotherapy, 2011
- Rapid Molecular Detection of Tuberculosis and Rifampin ResistanceNew England Journal of Medicine, 2010
- Evaluation of the Analytical Performance of the Xpert MTB/RIF AssayJournal of Clinical Microbiology, 2010
- Rapid Detection of Mycobacterium tuberculosis and Rifampin Resistance by Use of On-Demand, Near-Patient TechnologyJournal of Clinical Microbiology, 2010
- Diagnostic Accuracy of In-House PCR for Pulmonary Tuberculosis in Smear-Positive Patients: Meta-Analysis and MetaregressionJournal of Clinical Microbiology, 2009
- New recommendations for duration of respiratory isolation based on time to detect Mycobacterium tuberculosis in liquid cultureEuropean Respiratory Journal, 2007
- The implications of using an inappropriate reference gene for real-time reverse transcription PCR data normalizationAnalytical Biochemistry, 2005
- Evaluation of Human Antimycobacterial Immunity Using Recombinant Reporter MycobacteriaThe Journal of Infectious Diseases, 2000
- Strand Displacement Amplification and the Polymerase Chain Reaction for Monitoring Response to Treatment in Patients with Pulmonary TuberculosisThe Journal of Infectious Diseases, 1996