Title: Early detection of emergent extensively drug resistant tuberculosis (XDR-TB) by flow cytometry-based phenotyping and whole genome sequencing
Authors: O'Donnell MR, Larsen MH, Brown TS, Jain P, Munsamy V, Wolf A, Uccellini L, Karim F, de Oliveira T, Mathema B, Jacobs WR, Pym A.
Journal: Antimicrob Agents Chemother., pii: AAC.01834-18.:doi: 10.1128/AAC.01834-18 (2019)


Background A critical gap in tuberculosis (TB) treatment is detection of emergent drug resistance. We hypothesized that advanced phenotyping with whole genome sequencing (WGS) will detect low frequency Mycobacterium tuberculosis (MTB) drug-resistance.

Methods We assessed a reporter mycobacteriophage (?2GFP10) in vitro to detect drug-resistant subpopulations and predict MTB bactericidal activity in this pilot study. Subsequently, we prospectively studied 20 TB patients with serial ?2GFP10, Xpert MTB/RIF, and MTB culture through end of treatment. WGS was performed and single nucleotide polymorphisms (SNPs) were examined to detect mixed infection in selected MTB isolates.

Results Resistant MTB were detected at 1:100,000 and changes in cytometry gated events were predictive of in vitro MTB bactericidal activity using the ?2GFP10 assay. Emergent drug-resistance was detected in one patient by ?2GFP10 at three weeks but not by conventional testing (MTB culture and GeneXpert). WGS revealed a phylogeographically distinct extensively drug-resistant tuberculosis (XDR-TB) genome, identical to an XDR-TB isolate from the patient's spouse. Variant lineage-specific SNPs were present early, suggesting mixed infection as the etiology of emergent resistance with temporal trends providing evidence for selection during treatment.

Conclusions ?2GFP10 can detect low frequency drug-resistant MTB and with WGS characterize emergent MTB resistance. In areas of high TB transmission and drug resistance, rapid screening for heteroresistance should be considered.