Whole-Genome Sequencing Analysis of Serially Isolated Multi-Drug and Extensively Drug Resistant Mycobacterium tuberculosis from Thai Patients.

Kiatichai Faksri,Yik-Ying Teo,Eryu Xia,Jun Hao Tan,Rick Twee-Hee Ong,Chiea Chuen Khor,Areeya Disratthakit,Prapat Suriyaphol,Angkana Chaiprasert,Therdsak Prammananan,Srinand Sreevatsan

doi:10.1371/journal.pone.0160992

Kiatichai Faksri, Yik-Ying Teo + Show 9 more

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https://doi.org/10.1371/journal.pone.0160992

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Abstract

Multi-drug and extensively drug-resistant tuberculosis (MDR and XDR-TB) are problems that threaten public health worldwide. Only some genetic markers associated with drug-resistant TB are known. Whole-genome sequencing (WGS) is a promising tool for distinguishing between re-infection and persistent infection in isolates taken at different times from a single patient, but has not yet been applied in MDR and XDR-TB. We aim to detect genetic markers associated with drug resistance and distinguish between reinfection and persistent infection from MDR and XDR-TB patients based on WGS analysis. Samples of Mycobacterium tuberculosis (n = 7), serially isolated from 2 MDR cases and 1 XDR-TB case, were retrieved from Siriraj Hospital, Bangkok. The WGS analysis used an Illumina Miseq sequencer. In cases of persistent infection, MDR-TB isolates differed at an average of 2 SNPs across the span of 2–9 months whereas in the case of reinfection, isolates differed at 61 SNPs across 2 years. Known genetic markers associated with resistance were detected from strains susceptible to streptomycin (2/7 isolates), p-aminosalicylic acid (3/7 isolates) and fluoroquinolone drugs. Among fluoroquinolone drugs, ofloxacin had the highest phenotype-genotype concordance (6/7 isolates), whereas gatifloxcain had the lowest (3/7 isolates). A putative candidate SNP in Rv2477c associated with kanamycin and amikacin resistance was suggested for further validation. WGS provided comprehensive results regarding molecular epidemiology, distinguishing between persistent infection and reinfection in M/XDR-TB and potentially can be used for detection of novel mutations associated with drug resistance.

Highlights

Drug resistant tuberculosis (DR-TB), especially multi-drug and extensively drug resistant TB (MDR and XDR-TB), is a serious public health problem causing higher cost of treatment, lower treatment success rates, higher rate of persistent infection and higher mortality rates in TB patients.Genetic analysis of DR-TB provides rapid information for early management of TB patients
We confirm that Whole-genome sequencing (WGS) analysis provides rapid and comprehensive results for molecular epidemiology and detection of drug-resistant TB
The robust single nucleotide polymorphisms (SNPs)-barcode typing [35] showed that the strains WBB259 (DS16220) and WBB260 (DS16780) from Patient I belong to lineage 2.1 (nonBeijing in the East Asian (EA) lineage that normally lacks the RD105 deletion), yet had the RD105 deletion

Summary

Introduction

Genetic analysis of DR-TB provides rapid information for early management of TB patients. Random mutations may occur, comparisons between drug-resistant and susceptible cohorts in large Mtb populations provide statistically convincing identification of genetic markers for DR-TB [2]. Isolated Mtb samples from a single patient showing gradual increase of drug resistance provides raw material for the study of genetics associated with drug resistance [3,4,5]. The genetic backbone of the Mtb lineage derived from the same clone diminishes the risk of assuming that random mutations are associated with drug resistance. Additional WGS analysis of serially isolated Mtb samples from different settings and drug resistance profiles are required to provide the lacking information

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