Early Detection of Emergent Extensively Drug-Resistant Tuberculosis by Flow Cytometry-Based Phenotyping and Whole-Genome Sequencing.

Antimicrobial agents and chemotherapy, Volume: 63, Issue: 4
April 27, 2019
Max R O'Donnell MR, Michelle H Larsen MH, Tyler S Brown TS, Paras Jain P, Vanisha Munsamy V, Allison Wolf A, Lorenzo Uccellini L, Farina Karim F, Tulio de Oliveira T, Barun Mathema B, William R Jacobs WR, Alexander Pym A

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 drug resistance. We assessed a reporter mycobacteriophage (ΦGFP10) to detect drug-resistant subpopulations and predict bactericidal activity in this pilot study. Subsequently, we prospectively studied 20 TB patients with serial ΦGFP10, Xpert MTB/RIF, and culture through end of treatment. WGS was performed, and single nucleotide polymorphisms (SNPs) were examined to detect mixed infection in selected isolates. Resistant isolates were detected at 1:100,000, and changes in cytometry-gated events were predictive of bactericidal activity using the ΦGFP10 assay. Emergent drug resistance was detected in one patient by ΦGFP10 at 3 weeks but not by conventional testing ( 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. ΦGFP10 can detect low-frequency drug-resistant and with WGS characterize emergent resistance. In areas of high TB transmission and drug resistance, rapid screening for heteroresistance should be considered.

Courtesy of the U.S. National Library of Medicine