Targeted-Sequencing Workflows for Comprehensive Drug Resistance Profiling of Mycobacterium tuberculosis Cultures Using Two Commercial Sequencing Platforms: Comparison of Analytical and Diagnostic Performance, Turnaround Time, and Cost

Ketema Tafess, Timothy Ting Leung Ng, Hiu Yin Lao, Kenneth Siu Sing Leung, Kingsley King Gee Tam, Rahim Rajwani, Sarah Tsz Yan Tam, Lily Pui Ki Ho, Corey Mang Kiu Chu, Dimitri Gonzalez, Chalom Sayada, Oliver Chiu Kit Ma, Belete Haile Nega, Gobena Ameni, Wing Cheong Yam, Gilman Kit Hang Siu

Research output: Journal article publicationJournal articleAcademic researchpeer-review

33 Citations (Scopus)

Abstract

BACKGROUND: The emergence of Mycobacterium tuberculosis with complex drug resistance profiles necessitates a rapid and comprehensive drug susceptibility test for guidance of patient treatment. We developed two targeted-sequencing workflows based on Illumina MiSeq and Nanopore MinION for the prediction of drug resistance in M. tuberculosis toward 12 antibiotics. METHODS: A total of 163 M. tuberculosis isolates collected from Hong Kong and Ethiopia were subjected to a multiplex PCR for simultaneous amplification of 19 drug resistance-associated genetic regions. The amplicons were then barcoded and sequenced in parallel on MiSeq and MinION in respective batch sizes of 24 and 12 samples. A web-based bioinformatics pipeline, BacterioChek-TB, was developed to translate the raw datasets into clinician-friendly reports. RESULTS: Both platforms successfully sequenced all samples with mean read depths of 1,127× and 1,649×, respectively. The variant calling by MiSeq and MinION could achieve 100% agreement if variants with an allele frequency of <40% reported by MinION were excluded. Both workflows achieved a mean clinical sensitivity of 94.8% and clinical specificity of 98.0% when compared with phenotypic drug susceptibility test (pDST). Turnaround times for the MiSeq and MinION workflows were 38 and 15 h, facilitating the delivery of treatment guidance at least 17-18 days earlier than pDST, respectively. The higher cost per sample on the MinION platform ($71.56) versus the MiSeq platform ($67.83) was attributed to differences in batching capabilities. CONCLUSION: Our study demonstrates the interchangeability of MiSeq and MinION platforms for generation of accurate and actionable results for the treatment of tuberculosis.

Original languageEnglish
Pages (from-to)809-820
Number of pages12
JournalClinical Chemistry
Volume66
Issue number6
DOIs
Publication statusPublished - 1 Jun 2020

Keywords

  • Mycobacterium tuberculosis
  • drug-resistant TB
  • Illumina MiSeq
  • Nanopore MinION
  • next-generation sequencing
  • targeted sequencing

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Biochemistry, medical

Fingerprint

Dive into the research topics of 'Targeted-Sequencing Workflows for Comprehensive Drug Resistance Profiling of Mycobacterium tuberculosis Cultures Using Two Commercial Sequencing Platforms: Comparison of Analytical and Diagnostic Performance, Turnaround Time, and Cost'. Together they form a unique fingerprint.

Cite this