Use of Whole-Genome Sequencing to Predict<i>Mycobacterium tuberculosis</i>Complex Drug Resistance from Early Positive Liquid Cultures

Wu, Xiaodong; Tan, Guangkun; Wang, Sha; Liu, Haican; Yang, Jinghui; Guo, Yinjuan; Shen, Xin; Wu, Zheyuan; Shen, Hongbo; Yu, Fangyou

doi:10.1128/spectrum.02516-21

Cited by 12 publications

(15 citation statements)

References 50 publications

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“…Tuberculosis (TB) disease is a chronic infectious disease caused by Mycobacterium tuberculosis (MTB). Globally, there were an estimated 9.9 million new cases of MTB in 2020 [1], and about half a million of these cases were multidrug-resistant TB (MDR-TB) [2]. The emergency of drug-resistant TB (DR-TB) has threatened global public health efforts to control TB [3,4].…”

Section: Introductionmentioning

confidence: 99%

Drug Resistance and Molecular Characteristics of Mycobacterium tuberculosis: A Single Center Experience

Chen

Feng

et al. 2022

JPM

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In recent years, the incidence of tuberculosis (TB) and mortality caused by the disease have been decreasing. However, the number of drug-resistant tuberculosis patients is increasing rapidly year by year. Here, a total of 380 Mycobacterium tuberculosis (MTB)-positive formalin-fixed and paraffin-embedded tissue (FFPE) specimens diagnosed in the Department of Pathology of the Eighth Medical Center, Chinese PLA General Hospital were collected. Among 380 cases of MTB, 85 (22.37%) were susceptible to four anti-TB drugs and the remaining 295 (77.63%) were resistant to one or more drugs. The rate of MDR-TB was higher in previously treated cases (52.53%) than in new cases [(36.65%), p < 0.05]. Of previously treated cases, the rate of drug resistance was higher in females than in males (p < 0.05). Among specimens obtained from males, the rate of drug resistance was higher in new cases than in previously treated cases (p < 0.05). Of mutation in drug resistance-related genes, the majority (53/380, 13.95%) of rpoB gene carried the D516V mutation, and 13.42% (51/380) featured mutations in both the katG and inhA genes. Among the total specimens, 18.68% (71/380) carried the 88 M mutation in the rpsL gene, and the embB gene focused on the 306 M2 mutation with a mutation rate of 19.74%. Among the resistant INH, the mutation rate of −15 M was higher in resistance to more than one drug than in monodrug-resistant (p < 0.05). In conclusion, the drug resistance of MTB is still very severe and the timely detection of drug resistance is conducive to the precise treatment of TB.

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Section: Introductionmentioning

confidence: 99%

Drug Resistance and Molecular Characteristics of Mycobacterium tuberculosis: A Single Center Experience

Chen

Feng

et al. 2022

JPM

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“…Nowadays, with the help of advanced molecular biology techniques, difficult-to-culture microorganisms can also be sequenced using WGS. For example, the sequencing of Mycobacterium tuberculosis from liquid media using the Mycobacteria Growth Indicator Tube (MGIT) [ 22 ] has been established, although with a slower turnaround time compared to targeted DNA enrichment directly from sputum [ 23 ]. Viruses or intracellular bacteria can be amplified or enriched using (i) pathogen-specific PCRs (e.g., for SARS-CoV-2 or Influenza) [ 24 , 25 ] or (ii) bead-based DNA pull-down assays, e.g., for Chlamydia trachomatis [ 26 ], human papillomaviruses [ 27 ], or Noroviruses [ 28 ].…”

Section: Focus On Individual Microorganismsmentioning

confidence: 99%

Combination of Whole Genome Sequencing and Metagenomics for Microbiological Diagnostics

Purushothaman

Meola

Egli

2022

IJMS

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Whole genome sequencing (WGS) provides the highest resolution for genome-based species identification and can provide insight into the antimicrobial resistance and virulence potential of a single microbiological isolate during the diagnostic process. In contrast, metagenomic sequencing allows the analysis of DNA segments from multiple microorganisms within a community, either using an amplicon- or shotgun-based approach. However, WGS and shotgun metagenomic data are rarely combined, although such an approach may generate additive or synergistic information, critical for, e.g., patient management, infection control, and pathogen surveillance. To produce a combined workflow with actionable outputs, we need to understand the pre-to-post analytical process of both technologies. This will require specific databases storing interlinked sequencing and metadata, and also involves customized bioinformatic analytical pipelines. This review article will provide an overview of the critical steps and potential clinical application of combining WGS and metagenomics together for microbiological diagnosis.

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“…In addition, multiplexing in most library preparation protocols can reduce the detection cost. The constraint for using metagenomic sequencing in the routine clinical diagnostic workflow is, however, a high level of human DNA contamination, as expected in clinical sputum specimens, as well as low concentration of TB in metagenomic samples 7 – 9 . Also, a robust and reproducible complementary bioinformatics workflow is required for fast and accurate diagnosis.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of Mycobacterium tuberculosis enrichment in metagenomic samples using ONT adaptive sequencing and amplicon sequencing for identification and variant calling

Lee

et al. 2023

Sci Rep

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Sensitive detection of Mycobacterium tuberculosis (TB) in small percentages in metagenomic samples is essential for microbial classification and drug resistance prediction. However, traditional methods, such as bacterial culture and microscopy, are time-consuming and sometimes have limited TB detection sensitivity. Oxford nanopore technologies (ONT) MinION sequencing allows rapid and simple sample preparation for sequencing. Its recently developed adaptive sequencing selects reads from targets while allowing real-time base-calling to achieve sequence enrichment or depletion during sequencing. Another common enrichment method is PCR amplification of the target TB genes. In this study, we compared both methods using ONT MinION sequencing for TB detection and variant calling in metagenomic samples using both simulation runs and those with synthetic and patient samples. We found that both methods effectively enrich TB reads from a high percentage of human (95%) and other microbial DNA. Adaptive sequencing with readfish and UNCALLDE achieved a 3.9-fold and 2.2-fold enrichment compared to the control run. We provide a simple automatic analysis framework to support the detection of TB for clinical use, openly available at https://github.com/HKU-BAL/ONT-TB-NF. Depending on the patient's medical condition and sample type, we recommend users evaluate and optimize their workflow for different clinical specimens to improve the detection limit.

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Use of Whole-Genome Sequencing to PredictMycobacterium tuberculosisComplex Drug Resistance from Early Positive Liquid Cultures

Cited by 12 publications

References 50 publications

Drug Resistance and Molecular Characteristics of Mycobacterium tuberculosis: A Single Center Experience

Drug Resistance and Molecular Characteristics of Mycobacterium tuberculosis: A Single Center Experience

Combination of Whole Genome Sequencing and Metagenomics for Microbiological Diagnostics

Evaluation of Mycobacterium tuberculosis enrichment in metagenomic samples using ONT adaptive sequencing and amplicon sequencing for identification and variant calling

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