Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system

Elisa Schena, Lubov Nedialkova, Emanuele Borroni, Simone Battaglia, Andrea Maurizio Cabibbe, Stefan Niemann, Christian Utpatel, Matthias Merker, Alberto Trovato, Sabine Hofmann-Thiel, Harald Hoffmann, Daniela Maria Cirillo

Research output: Contribution to journalArticle

Abstract

Objectives: The objective of this study was to develop standardized protocols for rapid delamanid drug susceptibility testing (DST) using the colorimetric resazurin microtitre assay (REMA) and semi-automated BACTEC™ MGIT™ 960 system (MGIT) by establishing breakpoints that accurately discriminate between susceptibility and resistance of Mycobacterium tuberculosis to delamanid. Methods: MICs of delamanid were determined by the MGIT, the REMA and the solid agar method for 19 pre-characterized strains. The MIC distribution of delamanid was then established for a panel of clinical strains never exposed to the drug and characterized by different geographical origins and susceptibility patterns.WGSwas used to investigate genetic polymorphisms in five genes (ddn, fgd1, fbiA, fbiB and fbiC) involved in intracellular delamanid activation. Results: We demonstrated that the REMA and MGIT can both be used for the rapid and accurate determination of delamanid MIC, showing excellent concordance with the solid agar reference method, as well as high reproducibility and repeatability.We propose the tentative breakpoint of 0.125 mg/L for the REMA and MGIT, allowing reliable discrimination between M. tuberculosis susceptible and resistant to delamanid. Stop codon mutations in ddn (Trp-88→STOP) and fbiA (Lys-250→STOP) have only been observed in strains resistant to delamanid. Conclusions: We established protocols for DST of delamanid in the MGIT and REMA, confirming their feasibility in routine TB diagnostics, utilizing the same discriminative concentration for both methods. Moreover, taking advantage of WGS analysis, we identified polymorphisms potentially associated with resistance in two genes involved in delamanid activation.

Original languageEnglish
Pages (from-to)1532-1539
Number of pages8
JournalJournal of Antimicrobial Chemotherapy
Volume71
Issue number6
DOIs
Publication statusPublished - Jun 13 2016

Fingerprint

Mycobacterium tuberculosis
Agar
OPC-67683
resazurin
Pharmaceutical Preparations
Terminator Codon
Genetic Polymorphisms
Genes
Mutation

ASJC Scopus subject areas

  • Pharmacology
  • Pharmacology (medical)
  • Infectious Diseases

Cite this

Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system. / Schena, Elisa; Nedialkova, Lubov; Borroni, Emanuele; Battaglia, Simone; Cabibbe, Andrea Maurizio; Niemann, Stefan; Utpatel, Christian; Merker, Matthias; Trovato, Alberto; Hofmann-Thiel, Sabine; Hoffmann, Harald; Cirillo, Daniela Maria.

In: Journal of Antimicrobial Chemotherapy, Vol. 71, No. 6, 13.06.2016, p. 1532-1539.

Research output: Contribution to journalArticle

Schena, E, Nedialkova, L, Borroni, E, Battaglia, S, Cabibbe, AM, Niemann, S, Utpatel, C, Merker, M, Trovato, A, Hofmann-Thiel, S, Hoffmann, H & Cirillo, DM 2016, 'Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system', Journal of Antimicrobial Chemotherapy, vol. 71, no. 6, pp. 1532-1539. https://doi.org/10.1093/jac/dkw044
Schena, Elisa ; Nedialkova, Lubov ; Borroni, Emanuele ; Battaglia, Simone ; Cabibbe, Andrea Maurizio ; Niemann, Stefan ; Utpatel, Christian ; Merker, Matthias ; Trovato, Alberto ; Hofmann-Thiel, Sabine ; Hoffmann, Harald ; Cirillo, Daniela Maria. / Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system. In: Journal of Antimicrobial Chemotherapy. 2016 ; Vol. 71, No. 6. pp. 1532-1539.
@article{eec58f31d75f4b439ef706507e8b3041,
title = "Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system",
abstract = "Objectives: The objective of this study was to develop standardized protocols for rapid delamanid drug susceptibility testing (DST) using the colorimetric resazurin microtitre assay (REMA) and semi-automated BACTEC™ MGIT™ 960 system (MGIT) by establishing breakpoints that accurately discriminate between susceptibility and resistance of Mycobacterium tuberculosis to delamanid. Methods: MICs of delamanid were determined by the MGIT, the REMA and the solid agar method for 19 pre-characterized strains. The MIC distribution of delamanid was then established for a panel of clinical strains never exposed to the drug and characterized by different geographical origins and susceptibility patterns.WGSwas used to investigate genetic polymorphisms in five genes (ddn, fgd1, fbiA, fbiB and fbiC) involved in intracellular delamanid activation. Results: We demonstrated that the REMA and MGIT can both be used for the rapid and accurate determination of delamanid MIC, showing excellent concordance with the solid agar reference method, as well as high reproducibility and repeatability.We propose the tentative breakpoint of 0.125 mg/L for the REMA and MGIT, allowing reliable discrimination between M. tuberculosis susceptible and resistant to delamanid. Stop codon mutations in ddn (Trp-88→STOP) and fbiA (Lys-250→STOP) have only been observed in strains resistant to delamanid. Conclusions: We established protocols for DST of delamanid in the MGIT and REMA, confirming their feasibility in routine TB diagnostics, utilizing the same discriminative concentration for both methods. Moreover, taking advantage of WGS analysis, we identified polymorphisms potentially associated with resistance in two genes involved in delamanid activation.",
author = "Elisa Schena and Lubov Nedialkova and Emanuele Borroni and Simone Battaglia and Cabibbe, {Andrea Maurizio} and Stefan Niemann and Christian Utpatel and Matthias Merker and Alberto Trovato and Sabine Hofmann-Thiel and Harald Hoffmann and Cirillo, {Daniela Maria}",
year = "2016",
month = "6",
day = "13",
doi = "10.1093/jac/dkw044",
language = "English",
volume = "71",
pages = "1532--1539",
journal = "Journal of Antimicrobial Chemotherapy",
issn = "0305-7453",
publisher = "Oxford University Press",
number = "6",

}

TY - JOUR

T1 - Delamanid susceptibility testing of Mycobacterium tuberculosis using the resazurin microtitre assay and the BACTEC™ MGIT™ 960 system

AU - Schena, Elisa

AU - Nedialkova, Lubov

AU - Borroni, Emanuele

AU - Battaglia, Simone

AU - Cabibbe, Andrea Maurizio

AU - Niemann, Stefan

AU - Utpatel, Christian

AU - Merker, Matthias

AU - Trovato, Alberto

AU - Hofmann-Thiel, Sabine

AU - Hoffmann, Harald

AU - Cirillo, Daniela Maria

PY - 2016/6/13

Y1 - 2016/6/13

N2 - Objectives: The objective of this study was to develop standardized protocols for rapid delamanid drug susceptibility testing (DST) using the colorimetric resazurin microtitre assay (REMA) and semi-automated BACTEC™ MGIT™ 960 system (MGIT) by establishing breakpoints that accurately discriminate between susceptibility and resistance of Mycobacterium tuberculosis to delamanid. Methods: MICs of delamanid were determined by the MGIT, the REMA and the solid agar method for 19 pre-characterized strains. The MIC distribution of delamanid was then established for a panel of clinical strains never exposed to the drug and characterized by different geographical origins and susceptibility patterns.WGSwas used to investigate genetic polymorphisms in five genes (ddn, fgd1, fbiA, fbiB and fbiC) involved in intracellular delamanid activation. Results: We demonstrated that the REMA and MGIT can both be used for the rapid and accurate determination of delamanid MIC, showing excellent concordance with the solid agar reference method, as well as high reproducibility and repeatability.We propose the tentative breakpoint of 0.125 mg/L for the REMA and MGIT, allowing reliable discrimination between M. tuberculosis susceptible and resistant to delamanid. Stop codon mutations in ddn (Trp-88→STOP) and fbiA (Lys-250→STOP) have only been observed in strains resistant to delamanid. Conclusions: We established protocols for DST of delamanid in the MGIT and REMA, confirming their feasibility in routine TB diagnostics, utilizing the same discriminative concentration for both methods. Moreover, taking advantage of WGS analysis, we identified polymorphisms potentially associated with resistance in two genes involved in delamanid activation.

AB - Objectives: The objective of this study was to develop standardized protocols for rapid delamanid drug susceptibility testing (DST) using the colorimetric resazurin microtitre assay (REMA) and semi-automated BACTEC™ MGIT™ 960 system (MGIT) by establishing breakpoints that accurately discriminate between susceptibility and resistance of Mycobacterium tuberculosis to delamanid. Methods: MICs of delamanid were determined by the MGIT, the REMA and the solid agar method for 19 pre-characterized strains. The MIC distribution of delamanid was then established for a panel of clinical strains never exposed to the drug and characterized by different geographical origins and susceptibility patterns.WGSwas used to investigate genetic polymorphisms in five genes (ddn, fgd1, fbiA, fbiB and fbiC) involved in intracellular delamanid activation. Results: We demonstrated that the REMA and MGIT can both be used for the rapid and accurate determination of delamanid MIC, showing excellent concordance with the solid agar reference method, as well as high reproducibility and repeatability.We propose the tentative breakpoint of 0.125 mg/L for the REMA and MGIT, allowing reliable discrimination between M. tuberculosis susceptible and resistant to delamanid. Stop codon mutations in ddn (Trp-88→STOP) and fbiA (Lys-250→STOP) have only been observed in strains resistant to delamanid. Conclusions: We established protocols for DST of delamanid in the MGIT and REMA, confirming their feasibility in routine TB diagnostics, utilizing the same discriminative concentration for both methods. Moreover, taking advantage of WGS analysis, we identified polymorphisms potentially associated with resistance in two genes involved in delamanid activation.

UR - http://www.scopus.com/inward/record.url?scp=84973370236&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84973370236&partnerID=8YFLogxK

U2 - 10.1093/jac/dkw044

DO - 10.1093/jac/dkw044

M3 - Article

AN - SCOPUS:84973370236

VL - 71

SP - 1532

EP - 1539

JO - Journal of Antimicrobial Chemotherapy

JF - Journal of Antimicrobial Chemotherapy

SN - 0305-7453

IS - 6

ER -