Speeding up drug susceptibility testing in Mycobacterium tuberculosis using RNA biomarkers
Abstract
BACKGROUND: Efficient management of drug-resistant tuberculosis relies on fast diagnostics. To accelerate phenotypic drug susceptibility testing [pDST] for Mycobacterium tuberculosis [TB], we introduce TRACeR-TB, a test that infers drug resistance from antibiotic-specific mRNA biomarkers.
METHODS: To develop TRACeR-TB, target genes were first identified through RNA sequencing experiments conducted on two drug-exposed, susceptible strains for four antitubercular drugs. Based on these findings, we designed drug-specific multiplex Quantigene panels to quantify mRNA levels of 8-9 biomarkers per drug (class), directly from crude cell lysates. The performance of TRACeR-TB was compared to the widely used Mycobacteria Growth Indicator Tube [MGIT] pDST by subjecting 238 strains with diverse drug resistance profiles to both methods, and aligning results to genotypic data. Furthermore, we explored TRACeR-TB's potential for evaluating molecules that enhance antibiotic efficacy, and investigated its applicability in macrophage models to assess Mtb's intracellular stress responses to drugs.
FINDINGS: Antituberculosis drugs trigger distinct transcriptional stress responses in susceptible, but not resistant bacilli, enabling a differentiation of the antibiotic phenotype in only 6 h. Validation on 238 strains showed TRACeR-TB had 100% (95% CI: 93·1-100%) sensitivity and 89·5% (95% CI: 74·7-97·2%) specificity compared to, respectively, 82·3% (95% CI: 69·2%-91·5%) and 94·8% (95% CI:…
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