Whole-genome sequencing of presumptive MDR-TB isolates from a tertiary healthcare setting in Mumbai

Zade, Amrutraj; Shah, Sanchi; Hirani, Nilma; Kondabagil, Kiran; Joshi, Ameeta; Chatterjee, Anirvan

doi:10.1016/j.jgar.2022.10.004

Cited by 4 publications

(2 citation statements)

References 32 publications

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“…These observations potentially allow the replacement of phenotypic DST with WGS analysis for both medications. Regarding linezolid, there are known mutations in rplC and rrl genes which were associated with the resistance [ 31 , 32 ]. However, we did not identify any linezolid-resistant Mtb isolates, thus, in this study, the clinically-sound conclusions regarding the interchangeability of phenotypic DST and WGS for linezolid could not be drawn.…”

Section: Discussionmentioning

confidence: 99%

Genotypic and phenotypic comparison of drug resistance profiles of clinical multidrug-resistant Mycobacterium tuberculosis isolates using whole genome sequencing in Latvia

Vīksna,

Sadovska,

Berge

et al. 2023

BMC Infect Dis

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Background Multidrug-resistant tuberculosis (MDR–TB) remains a major public health problem in many high tuberculosis (TB) burden countries. Phenotypic drug susceptibility testing (DST) take several weeks or months to result, but line probe assays and Xpert/Rif Ultra assay detect a limited number of resistance conferring gene mutations. Whole genome sequencing (WGS) is an advanced molecular testing method which theoretically can predict the resistance of M. tuberculosis (Mtb) isolates to all anti-TB agents through a single analysis. Methods Here, we aimed to identify the level of concordance between the phenotypic and WGS-based genotypic drug susceptibility (DS) patterns of MDR–TB isolates. Overall, data for 12 anti-TB medications were analyzed. Results In total, 63 MDR–TB Mtb isolates were included in the analysis, representing 27.4% of the total number of MDR–TB cases in Latvia in 2012–2014. Among them, five different sublineages were detected, and 2.2.1 (Beijing group) and 4.3.3 (Latin American-Mediterranean group) were the most abundant. There were 100% agreement between phenotypic and genotypic DS pattern for isoniazid, rifampicin, and linezolid. High concordance rate (> 90%) between phenotypic and genotypic DST results was detected for ofloxacin (93.7%), pyrazinamide (93.7%) and streptomycin (95.4%). Phenotypic and genotypic DS patterns were poorly correlated for ethionamide (agreement 56.4%), ethambutol (85.7%), amikacin (82.5%), capreomycin (81.0%), kanamycin (85.4%), and moxifloxacin (77.8%). For capreomycin, resistance conferring mutations were not identified in several phenotypically resistant isolates, and, in contrary, for ethionamide, ethambutol, amikacin, kanamycin, and moxifloxacin the resistance-related mutations were identified in several phenotypically sensitive isolates. Conclusions WGS is a valuable tool for rapid genotypic DST for all anti-TB agents. For isoniazid and rifampicin phenotypic DST potentially can be replaced by genotypic DST based on 100% agreement between the tests. However, discrepant results for other anti-TB agents limit their prescription based solely on WGS data. For clinical decision, at the current level of knowledge, there is a need for combination of genotypic DST with modern, validated phenotypic DST methodologies for those medications which did not showed 100% agreement between the methods.

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Section: Discussionmentioning

confidence: 99%

Genotypic and phenotypic comparison of drug resistance profiles of clinical multidrug-resistant Mycobacterium tuberculosis isolates using whole genome sequencing in Latvia

Vīksna,

Sadovska,

Berge

et al. 2023

BMC Infect Dis

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“…At the Upper Myanmar TB Laboratory, genomic DNA was extracted from sputum culture isolates. MoBio Microbial DNA Isolation Kits, (Qigen DNEasy Ultraclean Microbial DNA extraction kit, Cat No: 12224-50) was used [14][15][16][17]. Both first-and second-line line probe assay (LPA) were tested by the MTB DRplus kit and the MTB DRsl kit.…”

Section: Extraction Of Genomic Dnamentioning

confidence: 99%

Genomic Sequencing Profiles of Mycobacterium tuberculosis in Mandalay Region, Myanmar

et al. 2023

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This study aimed to characterize whole-genome sequencing (WGS) information of Mycobacterium tuberculosis (Mtb) in the Mandalay region of Myanmar. It was a cross-sectional study conducted with 151 Mtb isolates obtained from the fourth nationwide anti-tuberculosis (TB) drug-resistance survey. Frequency of lineages 1, 2, 3, and 4 were 55, 65, 9, and 22, respectively. The most common sublineage was L1.1.3.1 (n = 31). Respective multi-drug resistant tuberculosis (MDR-TB) frequencies were 1, 1, 0, and 0. Four clusters of 3 (L2), 2 (L4), 2 (L1), and 2 (L2) isolates defined by a 20-single-nucleotide variant (SNV) cutoff were detected. Simpson’s index for sublineages was 0.0709. Such high diversity suggests that the area probably had imported Mtb from many geographical sources. Relatively few genetic clusters and MDR-TB suggest there is a chance the future control will succeed if it is carried out properly.

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An integrated method for targeted Oxford Nanopore sequencing and automated bioinformatics for the simultaneous detection of bacteria, fungi, and ARG

Kuruwa,

Zade,

Shah

et al. 2024

Journal of Applied Microbiology

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Aims The use of metagenomics for pathogen identification in clinical practice has been limited. Here we describe a workflow to encourage the clinical utility and potential of NGS for the screening of bacteria, fungi, and antimicrobial resistance genes (ARGs). Methods and results The method includes target enrichment, long-read sequencing, and automated bioinformatics. Evaluation of several tools and databases was undertaken across standard organisms (n = 12), clinical isolates (n = 114), and blood samples from patients with suspected bloodstream infections (n = 33). The strategy used could offset the presence of host background DNA, error rates of long-read sequencing, and provide accurate and reproducible detection of pathogens. Eleven targets could be successfully tested in a single assay. Organisms could be confidently identified considering ≥60% of best hits of a BLAST-based threshold of e-value 0.001 and a percent identity of >80%. For ARGs, reads with percent identity of >90% and >60% overlap of the complete gene could be confidently annotated. A kappa of 0.83 was observed compared to standard diagnostic methods. Thus, a workflow for the direct-from-sample, on-site sequencing combined with automated genomics was demonstrated to be reproducible. Conclusion NGS-based technologies overcome several limitations of current day diagnostics. Highly sensitive and comprehensive methods of pathogen screening are the need of the hour. We developed a framework for reliable, on-site, screening of pathogens.

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Whole-genome sequencing of presumptive MDR-TB isolates from a tertiary healthcare setting in Mumbai

Cited by 4 publications

References 32 publications

Genotypic and phenotypic comparison of drug resistance profiles of clinical multidrug-resistant Mycobacterium tuberculosis isolates using whole genome sequencing in Latvia

Genotypic and phenotypic comparison of drug resistance profiles of clinical multidrug-resistant Mycobacterium tuberculosis isolates using whole genome sequencing in Latvia

Genomic Sequencing Profiles of Mycobacterium tuberculosis in Mandalay Region, Myanmar

An integrated method for targeted Oxford Nanopore sequencing and automated bioinformatics for the simultaneous detection of bacteria, fungi, and ARG

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