Please use this identifier to cite or link to this item: http://hdl.handle.net/10397/79822
Title: Comparative genomic analysis of two clonally related multidrug resistant mycobacterium tuberculosis by single molecule real time sequencing
Authors: Leung, KSS
Siu, GKH 
Tam, KKG
To, SWC
Rajwani, R 
Ho, PL
Wong, SSY
Zhao, WW
Ma, OCK
Yam, WC
Keywords: Multidrug resistance
Mycobacterium tuberculosis
PacBio sequencing
Growth rate
Comparative genomic analysis
Issue Date: 2017
Publisher: Frontiers Research Foundation
Source: Frontiers in cellular and infection microbiology, 15 Nov. 2017, v. 7, 478 How to cite?
Journal: Frontiers in cellular and infection microbiology 
Abstract: Background : Multidrug-resistant tuberculosis (MDR-TB) is posing a major threat to global TB control. In this study, we focused on two consecutive MDR-TB isolated from the same patient before and after the initiation of anti-TB treatment. To better understand the genomic characteristics of MDR-TB, Single Molecule Real-Time (SMRT) Sequencing and comparative genomic analyses was performed to identify mutations that contributed to the stepwise development of drug resistance and growth fitness in MDR-TB under in vivo challenge of anti-TB drugs.
Result : Both pre-treatment and post-treatment strain demonstrated concordant phenotypic and genotypic susceptibility profiles toward rifampicin, pyrazinamide, streptomycin, fluoroquinolones, aminoglycosides, cycloserine, ethionamide, and para-aminosalicylic acid. However, although both strains carried identical missense mutations at rpoB S531L, inhA C-15T, and embB M306V, MYCOTB Sensititre assay showed that the post-treatment strain had 16-, 8-, and 4-fold elevation in the minimum inhibitory concentrations (MICs) toward rifabutin, isoniazid, and ethambutol respectively. The results have indicated the presence of additional resistant-related mutations governing the stepwise development of MDR-TB. Further comparative genomic analyses have identified three additional polymorphisms between the clinical isolates. These include a single nucleotide deletion at nucleotide position 360 of rv0888 in pre-treatment strain, and a missense mutation at rv3303c (lpdA) V44I and a 6-bp inframe deletion at codon 67-68 in rv2071c (cobM) in the post-treatment strain. Multiple sequence alignment showed that these mutations were occurring at highly conserved regions among pathogenic mycobacteria. Using structural-based and sequence-based algorithms, we further predicted that the mutations potentially have deleterious effect on protein function.
Conclusion : This is the first study that compared the full genomes of two clonally-related MDR-TB clinical isolates during the course of anti-TB treatment. Our work has demonstrated the robustness of SMRT Sequencing in identifying mutations among MDR-TB clinical isolates. Comparative genome analysis also suggested novel mutations at rv0888, lpdA, and cobM that might explain the difference in antibiotic resistance and growth pattern between the two MDR-TB strains.
URI: http://hdl.handle.net/10397/79822
ISSN: 2235-2988
DOI: 10.3389/fcimb.2017.00478
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