scholarly journals Revisiting Activation of and Mechanism of Resistance to Compound IQG-607 in Mycobacterium tuberculosis

2017 ◽  
Vol 62 (2) ◽  
Author(s):  
Bruno L. Abbadi ◽  
Anne D. Villela ◽  
Valnês S. Rodrigues-Junior ◽  
Fernanda T. Subtil ◽  
Pedro F. Dalberto ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Metal Complex ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Isolates ◽  
Whole Genome ◽  
Content Type ◽  
Mechanism Of Resistance ◽  
Resistant Strains ◽  
Spontaneous Mutants

ABSTRACT IQG-607 is a metal complex previously reported as a promising anti-tuberculosis (TB) drug against isoniazid (INH)-resistant strains of Mycobacterium tuberculosis. Unexpectedly, we found that INH-resistant clinical isolates were resistant to IQG-607. Spontaneous mutants resistant to IQG-607 were subjected to whole-genome sequencing, and all sequenced colonies carried alterations in the katG gene. The katG(S315T) mutation was sufficient to confer resistance to IQG-607 in both MIC assays and inside macrophages. Moreover, overexpression of the InhA(S94A) protein caused IQG-607's resistance.

scholarly journals Tracing Melioidosis Back to the Source: Using Whole-Genome Sequencing To Investigate an Outbreak Originating from a Contaminated Domestic Water Supply

2015 ◽  
Vol 53 (4) ◽  
pp. 1144-1148 ◽  
Author(s):  
Evan McRobb ◽  
Derek S. Sarovich ◽  
Erin P. Price ◽  
Mirjam Kaestli ◽  
Mark Mayo ◽  
...  
Keyword(s):  
Water Supply ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Isolates ◽  
Whole Genome ◽  
Northern Australia ◽  
Source Attribution ◽  
Environmental Strain ◽  
Content Type ◽  
Groundwater Supply

Melioidosis, a disease of public health importance in Southeast Asia and northern Australia, is caused by the Gram-negative soil bacillusBurkholderia pseudomallei. Melioidosis is typically acquired through environmental exposure, and case clusters are rare, even in regions where the disease is endemic.B. pseudomalleiis classed as a tier 1 select agent by the Centers for Disease Control and Prevention; from a biodefense perspective, source attribution is vital in an outbreak scenario to rule out a deliberate release. Two cases of melioidosis within a 3-month period at a residence in rural northern Australia prompted an investigation to determine the source of exposure.B. pseudomalleiisolates from the property's groundwater supply matched the multilocus sequence type of the clinical isolates. Whole-genome sequencing confirmed the water supply as the probable source of infection in both cases, with the clinical isolates differing from the likely infecting environmental strain by just one single nucleotide polymorphism (SNP) each. For the first time, we report a phylogenetic analysis of genomewide insertion/deletion (indel) data, an approach conventionally viewed as problematic due to high mutation rates and homoplasy. Our whole-genome indel analysis was concordant with the SNP phylogeny, and these two combined data sets provided greater resolution and a better fit with our epidemiological chronology of events. Collectively, this investigation represents a highly accurate account of source attribution in a melioidosis outbreak and gives further insight into a frequently overlooked reservoir ofB. pseudomallei. Our methods and findings have important implications for outbreak source tracing of this bacterium and other highly recombinogenic pathogens.

scholarly journals Comprehensive Whole-Genome Sequencing and Reporting of Drug Resistance Profiles on Clinical Cases of Mycobacterium tuberculosis in New York State

2017 ◽  
Vol 55 (6) ◽  
pp. 1871-1882 ◽  
Author(s):  
Joseph Shea ◽  
Tanya A. Halse ◽  
Pascal Lapierre ◽  
Matthew Shudt ◽  
Donna Kohlerschmidt ◽  
...  
Keyword(s):  
New York ◽  
Drug Resistance ◽  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Predictive Value ◽  
New York State ◽  
Whole Genome ◽  
Content Type ◽  
York State

ABSTRACTWhole-genome sequencing (WGS) is a newer alternative for tuberculosis (TB) diagnostics and is capable of providing rapid drug resistance profiles while performing species identification and capturing the data necessary for genotyping. Our laboratory developed and validated a comprehensive and sensitive WGS assay to characterizeMycobacterium tuberculosisand otherM. tuberculosiscomplex (MTBC) strains, composed of a novel DNA extraction, optimized library preparation, paired-end WGS, and an in-house-developed bioinformatics pipeline. This new assay was assessed using 608 MTBC isolates, with 146 isolates during the validation portion of this study and 462 samples received prospectively. In February 2016, this assay was implemented to test all clinical cases of MTBC in New York State, including isolates and early positive Bactec mycobacterial growth indicator tube (MGIT) 960 cultures from primary specimens. Since the inception of the assay, we have assessed the accuracy of identification of MTBC strains to the species level, concordance with culture-based drug susceptibility testing (DST), and turnaround time. Species identification by WGS was determined to be 99% accurate. Concordance between drug resistance profiles generated by WGS and culture-based DST methods was 96% for eight drugs, with an average resistance-predictive value of 93% and susceptible-predictive value of 96%. This single comprehensive WGS assay has replaced seven molecular assays and has resulted in resistance profiles being reported to physicians an average of 9 days sooner than with culture-based DST for first-line drugs and 32 days sooner for second-line drugs.

scholarly journals Genome Sequencing of Polydrug-, Multidrug-, and Extensively Drug-Resistant Mycobacterium tuberculosis Strains from South India

2019 ◽  
Vol 8 (12) ◽  
Author(s):  
Sivakumar Shanmugam ◽  
Narender Kumar ◽  
Dina Nair ◽  
Mohan Natrajan ◽  
Srikanth Prasad Tripathy ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
South India ◽  
Sequence Data ◽  
Whole Genome ◽  
Drug Resistant ◽  
Resistance Mutations ◽  
Content Type ◽  
Extensively Drug Resistant

The genomes of 16 clinical Mycobacterium tuberculosis isolates were subjected to whole-genome sequencing to identify mutations related to resistance to one or more anti-Mycobacterium drugs. The sequence data will help in understanding the genomic characteristics of M. tuberculosis isolates and their resistance mutations prevalent in South India.

scholarly journals Whole-Genome Sequencing of Streptomycin-Resistant Mycobacterium tuberculosis Strain SBH145 from Sabah, Malaysia

2022 ◽  
Author(s):  
Zainal Arifin Mustapha ◽  
Jaeyres Jani ◽  
Cheronie Shely Stanis ◽  
Dg Syahidah Nadiah Abdull Majid ◽  
Chin Kai Ling ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Pulmonary Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
The Philippines ◽  
Whole Genome ◽  
Content Type ◽  
Tuberculosis Strain ◽  
Mycobacterium Tuberculosis Strain ◽  
Malaysian Borneo

This paper reports on the whole-genome sequencing of a streptomycin-resistant Mycobacterium tuberculosis strain that was isolated from a patient with pulmonary tuberculosis in Sabah state of Malaysian Borneo. The strain belongs to the EAI2-Manila family of lineage 1 and is clustered with M. tuberculosis strains from the Philippines, India, and Taiwan.

scholarly journals Direct DNA Extraction from Mycobacterium tuberculosis Frozen Stocks as a Reculture-Independent Approach to Whole-Genome Sequencing

2015 ◽  
Vol 53 (8) ◽  
pp. 2716-2719 ◽  
Author(s):  
K. Bjorn-Mortensen ◽  
J. Zallet ◽  
T. Lillebaek ◽  
A. B. Andersen ◽  
S. Niemann ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Dna Extraction ◽  
Genome Sequencing ◽  
Sequence Data ◽  
Whole Genome ◽  
Content Type ◽  
Direct Dna Extraction ◽  
Slow Growing

Culturing before DNA extraction represents a major time-consuming step in whole-genome sequencing of slow-growing bacteria, such asMycobacterium tuberculosis. We report a workflow to extract DNA from frozen isolates without reculturing. Prepared libraries and sequence data were comparable with results from recultured aliquots of the same stocks.

scholarly journals Activity of 2-(quinolin-4-yloxy)acetamides in Mycobacterium tuberculosis clinical isolates and identification of their molecular target by whole-genome sequencing

2018 ◽  
Vol 51 (3) ◽  
pp. 378-384 ◽  
Author(s):  
Fernanda Teixeira Subtil ◽  
Anne Drumond Villela ◽  
Bruno Lopes Abbadi ◽  
Valnês S. Rodrigues-Junior ◽  
Cristiano Valim Bizarro ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Molecular Target ◽  
Clinical Isolates ◽  
Whole Genome

scholarly journals Frequency and Mechanisms of Spontaneous Fosfomycin Nonsusceptibility Observed upon Disk Diffusion Testing of Escherichia coli

2017 ◽  
Vol 56 (1) ◽  
Author(s):  
Aaron E. Lucas ◽  
Ryota Ito ◽  
Mustapha M. Mustapha ◽  
Christi L. McElheny ◽  
Roberta T. Mettus ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Whole Genome ◽  
Zone Of Inhibition ◽  
Content Type ◽  
E Coli ◽  
Disk Diffusion Testing

ABSTRACTFosfomycin maintains activity against mostEscherichia coliclinical isolates, but the growth ofE. colicolonies within the zone of inhibition around the fosfomycin disk is occasionally observed upon susceptibility testing. We aimed to estimate the frequency of such nonsusceptible inner colony mutants and identify the underlying resistance mechanisms. Disk diffusion testing of fosfomycin was performed on 649 multidrug-resistantE. coliclinical isolates collected between 2011 and 2015. For those producing inner colonies inside the susceptible range, the parental strains and their representative inner colony mutants were subjected to MIC testing, whole-genome sequencing, reverse transcription-quantitative PCR (qRT-PCR), and carbohydrate utilization studies. Of the 649E. coliclinical isolates, 5 (0.8%) consistently produced nonsusceptible inner colonies. Whole-genome sequencing revealed the deletion ofuhpTencoding hexose-6-phosphate antiporter in 4 of theE. coliinner colony mutants, while the remaining mutant contained a nonsense mutation inuhpA. The expression ofuhpTwas absent in the mutant strains withuhpTdeletion and was not inducible in the strain with theuhpAmutation, unlike in its parental strain. All 5 inner colony mutants had reduced growth on minimal medium supplemented with glucose-6-phosphate. In conclusion, fosfomycin-nonsusceptible inner colony mutants can occur due to the loss of function or induction of UhpT but are rare among multidrug-resistantE. coliclinical strains. Considering that these mutants carry high biological costs, we suggest that fosfomycin susceptibility of strains that generate inner colony mutants can be interpreted on the basis of the zone of inhibition without accounting for the inner colonies.

scholarly journals Direct Whole-Genome Sequencing of Sputum Accurately Identifies Drug-Resistant Mycobacterium tuberculosis Faster than MGIT Culture Sequencing

2018 ◽  
Vol 56 (8) ◽  
Author(s):  
Ronan M. Doyle ◽  
Carrie Burgess ◽  
Rachel Williams ◽  
Rebecca Gorton ◽  
Helen Booth ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Antimicrobial Resistance ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Resistance Testing ◽  
Whole Genome ◽  
Accurate Identification ◽  
Content Type ◽  
Resistance Profile ◽  
Mgit Culture

ABSTRACT The current methods available to diagnose antimicrobial-resistant Mycobacterium tuberculosis infections require a positive culture or only test a limited number of resistance-associated mutations. A rapid accurate identification of antimicrobial resistance enables the prompt initiation of effective treatment. Here, we determine the utility of whole-genome sequencing (WGS) of M. tuberculosis directly from routinely obtained diagnostic sputum samples to provide a comprehensive resistance profile compared to that from mycobacterial growth indicator tube (MGIT) WGS. We sequenced M. tuberculosis from 43 sputum samples by targeted DNA enrichment using the Agilent SureSelectXT kit, and 43 MGIT positive samples from each participant. Thirty two (74%) sputum samples and 43 (100%) MGIT samples generated whole genomes. The times to antimicrobial resistance profiles and concordance were compared with Xpert MTB/RIF and phenotypic resistance testing from cultures of the same samples. Antibiotic susceptibility could be predicted from WGS of sputum within 5 days of sample receipt and up to 24 days earlier than WGS from MGIT culture and up to 31 days earlier than phenotypic testing. Direct sputum results could be reduced to 3 days with faster hybridization and if only regions encoding drug resistance are sequenced. We show that direct sputum sequencing has the potential to provide comprehensive resistance detection significantly faster than MGIT whole-genome sequencing or phenotypic testing of resistance from cultures in a clinical setting. This improved turnaround time enables prompt appropriate treatment with associated patient and health service benefits. Improvements in sample preparation are necessary to ensure comparable sensitivities and complete resistance profile predictions in all cases.

scholarly journals Rapid Whole-Genome Sequencing of Mycobacterium tuberculosis Isolates Directly from Clinical Samples

2015 ◽  
Vol 53 (7) ◽  
pp. 2230-2237 ◽  
Author(s):  
Amanda C. Brown ◽  
Josephine M. Bryant ◽  
Katja Einer-Jensen ◽  
Jolyon Holdstock ◽  
Darren T. Houniet ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Positive Culture ◽  
Clinical Samples ◽  
Whole Genome ◽  
Resistance Mutations ◽  
High Quality ◽  
Content Type ◽  
Negative Case

The rapid identification of antimicrobial resistance is essential for effective treatment of highly resistantMycobacterium tuberculosis. Whole-genome sequencing provides comprehensive data on resistance mutations and strain typing for monitoring transmission, but unlike for conventional molecular tests, this has previously been achievable only from cultures ofM. tuberculosis. Here we describe a method utilizing biotinylated RNA baits designed specifically forM. tuberculosisDNA to capture fullM. tuberculosisgenomes directly from infected sputum samples, allowing whole-genome sequencing without the requirement of culture. This was carried out on 24 smear-positive sputum samples, collected from the United Kingdom and Lithuania where a matched culture sample was available, and 2 samples that had failed to grow in culture.M. tuberculosissequencing data were obtained directly from all 24 smear-positive culture-positive sputa, of which 20 were of high quality (>20× depth and >90% of the genome covered). Results were compared with those of conventional molecular and culture-based methods, and high levels of concordance between phenotypical resistance and predicted resistance based on genotype were observed. High-quality sequence data were obtained from one smear-positive culture-negative case. This study demonstrated for the first time the successful and accurate sequencing ofM. tuberculosisgenomes directly from uncultured sputa. Identification of known resistance mutations within a week of sample receipt offers the prospect for personalized rather than empirical treatment of drug-resistant tuberculosis, including the use of antimicrobial-sparing regimens, leading to improved outcomes.

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