Detection of low-level mixed-population drug resistance in Mycobacterium tuberculosis using high fidelity amplicon sequencing

Rebecca E. Colman, James M. Schupp, Nathan D. Hicks, David E. Smith, Jordan L. Buchhagen, Faramarz Valafar, Valeriu Crudu, Elena Romancenco, Ecaterina Noroc, Lynn Jackson, Donald G. Catanzaro, Timothy C. Rodwell, Antonino Catanzaro, Paul S Keim, David M. Engelthaler

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Undetected and untreated, low-levels of drug resistant (DR) subpopulations in clinical Mycobacterium tuberculosis (Mtb) infections may lead to development of DR-tuberculosis, potentially resulting in treatment failure. Current phenotypic DR susceptibility testing has a theoretical potential for 1% sensitivity, is not quantitative, and requires several weeks to complete. The use of "single molecule-overlapping reads" (SMOR) analysis with next generation DNA sequencing for determination of ultra-rare target alleles in complex mixtures provides increased sensitivity over standard DNA sequencing. Ligation free amplicon sequencing with SMOR analysis enables the detection of resistant allele subpopulations at ≥0.1% of the total Mtb population in near real-time analysis. We describe the method using standardized mixtures of DNA from resistant and susceptible Mtb isolates and the assay's performance for detecting ultra-rare DR subpopulations in DNA extracted directly from clinical sputum samples. SMOR analysis enables rapid near real-time detection and tracking of previously undetectable DR sub-populations in clinical samples allowing for the evaluation of the clinical relevance of low-level DR subpopulations. This will provide insights into interventions aimed at suppressing minor DR subpopulations before they become clinically significant.

Original languageEnglish (US)
Article numbere0126626
JournalPLoS One
Volume10
Issue number5
DOIs
StatePublished - May 13 2015

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drug resistance
Mycobacterium tuberculosis
Drug Resistance
drugs
Pharmaceutical Preparations
Population
DNA Sequence Analysis
DNA
Molecules
Alleles
sequence analysis
Multidrug-Resistant Tuberculosis
Mycobacterium Infections
alleles
Sputum
Treatment Failure
Complex Mixtures
Ligation
tuberculosis
Assays

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

Colman, R. E., Schupp, J. M., Hicks, N. D., Smith, D. E., Buchhagen, J. L., Valafar, F., ... Engelthaler, D. M. (2015). Detection of low-level mixed-population drug resistance in Mycobacterium tuberculosis using high fidelity amplicon sequencing. PLoS One, 10(5), [e0126626]. https://doi.org/10.1371/journal.pone.0126626

Detection of low-level mixed-population drug resistance in Mycobacterium tuberculosis using high fidelity amplicon sequencing. / Colman, Rebecca E.; Schupp, James M.; Hicks, Nathan D.; Smith, David E.; Buchhagen, Jordan L.; Valafar, Faramarz; Crudu, Valeriu; Romancenco, Elena; Noroc, Ecaterina; Jackson, Lynn; Catanzaro, Donald G.; Rodwell, Timothy C.; Catanzaro, Antonino; Keim, Paul S; Engelthaler, David M.

In: PLoS One, Vol. 10, No. 5, e0126626, 13.05.2015.

Research output: Contribution to journalArticle

Colman, RE, Schupp, JM, Hicks, ND, Smith, DE, Buchhagen, JL, Valafar, F, Crudu, V, Romancenco, E, Noroc, E, Jackson, L, Catanzaro, DG, Rodwell, TC, Catanzaro, A, Keim, PS & Engelthaler, DM 2015, 'Detection of low-level mixed-population drug resistance in Mycobacterium tuberculosis using high fidelity amplicon sequencing', PLoS One, vol. 10, no. 5, e0126626. https://doi.org/10.1371/journal.pone.0126626
Colman, Rebecca E. ; Schupp, James M. ; Hicks, Nathan D. ; Smith, David E. ; Buchhagen, Jordan L. ; Valafar, Faramarz ; Crudu, Valeriu ; Romancenco, Elena ; Noroc, Ecaterina ; Jackson, Lynn ; Catanzaro, Donald G. ; Rodwell, Timothy C. ; Catanzaro, Antonino ; Keim, Paul S ; Engelthaler, David M. / Detection of low-level mixed-population drug resistance in Mycobacterium tuberculosis using high fidelity amplicon sequencing. In: PLoS One. 2015 ; Vol. 10, No. 5.
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