Deep amplicon sequencing for culture-free prediction of susceptibility or resistance to 13 anti-tuberculous drugs

Agathe Jouet, Cyril Gaudin, Nelly Badalato, Caroline Allix-Béguec, Stéphanie Duthoy, Alice Ferré, Maren Diels, Yannick Laurent, Sandy Contreras, Silke Feuerriegel, Stefan Niemann, Emmanuel André, Michel K Kaswa, Elisa Tagliani, Andrea Cabibbe, Vanessa Mathys, Daniela Cirillo, Bouke C de Jong, Leen Rigouts, Philip Supply

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Conventional molecular tests for detecting Mycobacterium tuberculosis complex (MTBC) drug resistance on clinical samples cover a limited set of mutations. Whole-genome sequencing (WGS) typically requires culture.Here, we evaluated the Deeplex Myc-TB targeted deep-sequencing assay for prediction of resistance to 13 anti-tuberculous drugs/drug classes, directly applicable on sputum.With MTBC DNA tests, the limit of detection was 100-1000 genome copies for fixed resistance mutations. Deeplex Myc-TB captured in silico 97.1-99.3% of resistance phenotypes correctly predicted by WGS from 3651 MTBC genomes. On 429 isolates, the assay predicted 92.2% of 2369 first- and second-line phenotypes, with a sensitivity of 95.3% and a specificity of 97.4%. 56 out of 69 (81.2%) residual discrepancies with phenotypic results involved pyrazinamide, ethambutol and ethionamide, and low-level rifampicin or isoniazid resistance mutations, all notoriously prone to phenotypic testing variability. Only two out of 91 (2.2%) resistance phenotypes undetected by Deeplex Myc-TB had known resistance-associated mutations by WGS analysis outside Deeplex Myc-TB targets. Phenotype predictions from Deeplex Myc-TB analysis directly on 109 sputa from a Djibouti survey matched those of MTBSeq/PhyResSE/Mykrobe, fed with WGS data from subsequent cultures, with a sensitivity of 93.5/98.5/93.1% and a specificity of 98.5/97.2/95.3%, respectively. Most residual discordances involved gene deletions/indels and 3-12% heteroresistant calls undetected by WGS analysis or natural pyrazinamide resistance of globally rare " Mycobacterium canettii" strains then unreported by Deeplex Myc-TB. On 1494 arduous sputa from a Democratic Republic of the Congo survey, 14 902 out of 19 422 (76.7%) possible susceptible or resistance phenotypes could be predicted culture-free.Deeplex Myc-TB may enable fast, tailored tuberculosis treatment.

Original languageEnglish
Article number2002338
JournalEuropean Respiratory Journal
Issue number3
Number of pages15
Publication statusPublished - 2021


  • Antitubercular Agents/pharmacology
  • Humans
  • Microbial Sensitivity Tests
  • Mutation
  • Mycobacterium tuberculosis/genetics
  • Pharmaceutical Preparations
  • Tuberculosis, Multidrug-Resistant/diagnosis
  • Tuberculosis/drug therapy


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