Rapid detection of Mycobacterium tuberculosis and rifampin resistance by use of on-demand, near-patient technology

Danica Helb, Martin Jones, Elizabeth Story, Catharina Boehme, Ellen Wallace, Ken Ho, Joann Kop, Michelle R. Owens, Richard Rodgers, Padmapriya Banada, Hassan Safi, Robert Blakemore, N. T. Ngoc Lan, Edward C. Jones-López, Michael Levi, Michele Burday, Irene Ayakaka, Roy D. Mugerwa, Bill McMillan, Emily Winn-DeenLee Christel, Peter Dailey, Mark D. Perkins, David H. Persing, David Alland

Research output: Contribution to journalArticle

543 Scopus citations

Abstract

Current nucleic acid amplification methods to detect Mycobacterium tuberculosis are complex, labor-intensive, and technically challenging. We developed and performed the first analysis of the Cepheid Gene Xpert System's MTB/RIF assay, an integrated hands-free sputum-processing and real-time PCR system with rapid on-demand, near-patient technology, to simultaneously detect M. tuberculosis and rifampin resistance. Analytic tests of M. tuberculosis DNA demonstrated a limit of detection (LOD) of 4.5 genomes per reaction. Studies using sputum spiked with known numbers of M. tuberculosis CFU predicted a clinical LOD of 131 CFU/ml. Killing studies showed that the assay's buffer decreased M. tuberculosis viability by at least 8 logs, substantially reducing biohazards. Tests of 23 different commonly occurring rifampin resistance mutations demonstrated that all 23 (100%) would be identified as rifampin resistant. An analysis of 20 nontuberculosis mycobacteria species confirmed high assay specificity. A small clinical validation study of 107 clinical sputum samples from suspected tuberculosis cases in Vietnam detected 29/29 (100%) smear-positive culture-positive cases and 33/39 (84.6%) or 38/53 (71.7%) smear-negative culture-positive cases, as determined by growth on solid medium or on both solid and liquid media, respectively. M. tuberculosis was not detected in 25/25 (100%) of the culture-negative samples. A study of 64 smear-positive culture-positive sputa from retreatment tuberculosis cases in Uganda detected 63/64 (98.4%) culture-positive cases and 9/9 (100%) cases of rifampin resistance. Rifampin resistance was excluded in 54/55 (98.2%) susceptible cases. Specificity rose to 100% after correcting for a conventional susceptibility test error. In conclusion, this highly sensitive and simple-to-use system can detect M. tuberculosis directly from sputum in less than 2 h.

Original languageEnglish (US)
Pages (from-to)229-237
Number of pages9
JournalJournal of Clinical Microbiology
Volume48
Issue number1
DOIs
StatePublished - Jan 1 2010

ASJC Scopus subject areas

  • Microbiology (medical)

Fingerprint Dive into the research topics of 'Rapid detection of Mycobacterium tuberculosis and rifampin resistance by use of on-demand, near-patient technology'. Together they form a unique fingerprint.

  • Cite this

    Helb, D., Jones, M., Story, E., Boehme, C., Wallace, E., Ho, K., Kop, J., Owens, M. R., Rodgers, R., Banada, P., Safi, H., Blakemore, R., Ngoc Lan, N. T., Jones-López, E. C., Levi, M., Burday, M., Ayakaka, I., Mugerwa, R. D., McMillan, B., ... Alland, D. (2010). Rapid detection of Mycobacterium tuberculosis and rifampin resistance by use of on-demand, near-patient technology. Journal of Clinical Microbiology, 48(1), 229-237. https://doi.org/10.1128/JCM.01463-09