Ketema Tafess 1,2 , Timothy Ting Leung Ng 1 , Hiu Yin Lao 1 , Kenneth Siu Sing Leung 3 , Kingsley King Gee Tam 3 , Rahim Rajwani 1 , Sarah Tsz Yan Tam 1 , Lily Pui Ki Ho 1 , Corey Mang Kiu Chu 1 , Dimitri Gonzalez 4 , Chalom Sayada 4 , Oliver Chiu Kit Ma 5 , Belete Haile Nega 6 , Gobena Ameni 6 , Wing Cheong Yam 2 , Gilman Kit Hang Siu 1 Show Affiliations »
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. © American Association for Clinical Chemistry 2020. All rights reserved. For permissions, please email: journals.permissions@oup.com.
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 . © American Association for Clinical Chemistry 2020. All rights reserved. For permissions, please email: journals.permissions@oup.com.
Entities: Disease
Species
Keywords:
zzm321990 Mycobacterium tuberculosiszzm321990 ; Illumina MiSeq; Nanopore MinION; drug-resistant TB; next-generation sequencing; targeted sequencing
Year: 2020
PMID： 32402055 DOI： 10.1093/clinchem/hvaa092
Source DB: PubMed Journal: Clin Chem ISSN： 0009-9147 Impact factor: 8.327