Design of Multidrug-Resistant Tuberculosis Treatment Regimens Based on DNA Sequencing

Abstract

Background: Comprehensive and reliable drug susceptibility testing (DST) is urgently needed to provide adequate treatment regimens for patients with multidrug-resistant/rifampicin-resistant tuberculosis (MDR/RR-TB). We determined whether next-generation sequencing (NGS) analysis of Mycobacterium tuberculosis complex isolates and genes implicated in drug resistance can guide the design of effective MDR/RR-TB treatment regimens.

Methods: NGS-based genomic DST predictions of M. tuberculosis complex isolates from MDR/RR-TB patients admitted to a TB reference center in Germany between 1 January 2015 and 30 April 2019 were compared with phenotypic DST results of mycobacteria growth indicator tubes (MGIT). Standardized treatment algorithms were applied to design individualized therapies based on either genomic or phenotypic DST results, and discrepancies were further evaluated by determination of minimal inhibitory drug concentrations (MICs) using Sensititre MYCOTBI and UKMYC microtiter plates.

Results: In 70 patients with MDR/RR-TB, agreement among 1048 pairwise comparisons of genomic and phenotypic DST was 86.3%; 76 (7.2%) results were discordant, and 68 (6.5%) could not be evaluated due to the presence of polymorphisms with yet unknown implications for drug resistance. Importantly, 549 of 561 (97.9%) predictions of drug susceptibility were phenotypically confirmed in MGIT, and 27 of 64 (42.2%) false-positive results were linked to previously described mutations mediating a low or moderate MIC increase. Virtually all drugs (99.0%) used in combination therapies that were inferred from genomic DST were confirmed to be susceptible by phenotypic DST.

Conclusions: NGS-based genomic DST can reliably guide the design of effective MDR/RR-TB treatment regimens.

Keywords: DST; MDR-TB; NGS; tuberculosis.

Figure 1.

Patients flow chart. Flow chart showing the patients evaluated in this study. Abbreviations: NGS, next-generation sequencing; MDR-TB = multidrug-resistant tuberculosis; RIF = rifampicin; MIC = minimum inhibitory concentration; MGIT, mycobacteria growth indicator tube; RR, rifampicin resistance; TB, tuberculosis.

Figure 2.

Phenotypic and genomic drug susceptibility testing (DST) data. Results of genomic prediction of DST by NGS, phenotypic DST in the MGIT 960 system, and phenotypic DST by broth microdilution assays (Sensititre MYCOTBI and/or UKMYC plates ; MGIT for pyrazinamide) for 70 patients. Each row represents a patient, and each column represents a drug. Resistant test results are shown in red, and susceptible results are represented by green boxes. In the case of Sensititre MYCOTBI/UKMYC, MICs 1 level above the cutoff are displayed in light red, and MICs at the cutoff are displayed in light green. Polymorphisms without clear association to drug resistance are displayed in gray. In cases where no result was available, a white box was inserted. Abbreviations: CC, critical concentration; MGIT, mycobacteria growth indicator tube; MIC, minimal inhibitory concentration; NGS, next-generation sequencing; PAS, para-aminosalicylic acid.

Figure 3.

Distribution of drug susceptibility test (DST) results by genomic NGS and phenotypic drug susceptibility testing (MGIT). A, Concordant and discordant test results and results of phenotypic testing with unclassified NGS results. Red box: false gDST predictions of susceptibility B, Overview of discrepant test results between DST prediction by NGS and phenotypic DST by MGIT. Susceptible test results are shown in green, and resistant test results are shown in red. Mutations of unknown relevance are displayed in gray. Abbreviations: MGIT, mycobacteria growth indicator tube; NGS, next-generation sequencing.

Figure 4.

Algorithm-derived treatment regimens based on different methods of drug susceptibility testing. Regimens were based on respective results of NGS, MGIT, and minimal inhibitory concentration (by Sensititre MYCOTBI and/or UKMYC; MGIT for pyrazinamide). Differences in the resulting therapy regimes compared with MGIT are highlighted by black frames. Red frames indicate treatment with a drug that tested resistant in MGIT. Columns indicate data for 16 drugs for each patient Meropenem was selected as per treatment algorith, irrespective of the unavailability of DST. Abbreviations: MGIT, mycobacteria growth indicator tube; NGS, next-generation sequencing; para-aminosalicylic acid.