scholarly journals Evaluation of a new automated Abbott RealTime MTB RIF/INH assay for qualitative detection of rifampicin/isoniazid resistance in pulmonary and extra-pulmonary clinical samples of Mycobacterium tuberculosis

2017 ◽  
Vol Volume 10 ◽  
pp. 463-467 ◽  
Author(s):  
Pilar Ruiz ◽  
Manuel Causse ◽  
Manuel Vaquero ◽  
Juan-Bautista Gutiérrez-Aroca ◽  
Manuel Casal
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Samples ◽  
Isoniazid Resistance ◽  
Qualitative Detection ◽  
Extra Pulmonary

scholarly journals Development of Au-Nanoprobes Combined with Loop-Mediated Isothermal Amplification for Detection of Isoniazid Resistance in Mycobacterium tuberculosis

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Jutturong Ckumdee ◽  
Thongchai Kaewphinit ◽  
Kosum Chansiri ◽  
Somchai Santiwatanakul
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multidrug Resistant ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
Isoniazid Resistance ◽  
Loop Mediated Isothermal Amplification ◽  
Specificity And Sensitivity ◽  
Mdr Tb ◽  
Induced Aggregation ◽  
Specific Sequences

Multidrug resistant tuberculosis (MDR-TB) is Mycobacterium tuberculosis that does not respond to isoniazid and rifampicin, so the condition worsens continuously and creates difficulties for treatment by public health control programmes, especially in developing countries. The real time polymerase chain reaction (PCR) combined with agarose gel electrophoresis or strip tests is useful molecular tools for diagnosis of MDR-TB. Novel loop-mediated isothermal amplification (LAMP) can also detect drug resistance, which is a one-point mutation, by designing inner primers of 5′ end specific with the mutant. Au-nanoprobes on hybridisation with LAMP products containing target-specific sequences remain red, whereas test samples without specific sequences in the probe turn purple due to salt-induced aggregation of the Au-nanoprobes. In this study, a strategy was designed based on the LAMP of a DNA sample coupled to specific Au-nanoprobes, which showed the potential to provide a rapid and sensitive method for detecting isoniazid resistance at katG gene position 315 (G→C). 46 clinical samples were tested and showed 100% specificity and sensitivity compared with Genotype® MDR-TB Plus. This method was advantageous because it is rapid, cheap, specific, and sensitive. Further, it does not require thermal cycles for MDR-TB detection.

scholarly journals Role of Polymerase Chain Reaction in diagnosis of Tuberculosis as compared to routine tests

2019 ◽  
Vol 6 (06) ◽  
pp. 4490-4494
Author(s):  
Dr Haris Memon ◽  
Dr Muhammad Haroon Mujtaba Memon ◽  
Dr Mahum Shahab ◽  
Dr Mahmood Iqbal ◽  
Dr Ghulam Murtaza Memon
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Synovial Fluid ◽  
Pleural Fluid ◽  
Tertiary Care ◽  
Clinical Samples ◽  
Pcr Analysis ◽  
Care Hospital ◽  
Specificity And Sensitivity ◽  
Pcr Method ◽  
Extra Pulmonary

Introduction: There is a need for rapid and sensitive detection of Mycobacterium tuberculosis in clinical samples. A study was conducted in which the target for the amplification being a segment of IS6110 in the M. tuberculosis chromosome was evaluated using real time PCR and its results were compared with routine tests, using pulmonary and extra-pulmonary specimen. Methods: In this descriptive cross-sectional retrospective study, specificity and sensitivity of PCR were analyzed. A total of 293 clinical samples were processed at a tertiary care hospital of Peshawar, during the time period of 2016-2018, from patients suspected of having pulmonary and extra-pulmonary tuberculosis and Follow up patients with DOTS treatment and MDR treatment that are referred by tertiary hospital were also included in this study after taking their informed consent. Patients not willing to participate in the study were excluded. For identification specimens were stained by Ziehl Neelsen staining (ZN), cultured on Lowenstein–Jensen (LJ) medium and then confirmed by PCR for the detection of Mycobacterium tuberculosis (MTB). Results: Of the 293 samples, 165(56.3%) were from males and 128(43.7%) females. Mean age was 44 years (2-85 years). Specimen types included: CSF (30.4%), pleural fluid (4.1%), sputum (15%), urine (2.4%), synovial fluid (2.4%), other fluids (33.1%) and biopsies (12.6%). Only 3.1% of specimens were ZN-smear positive for (MTB). LJ culture identified 7.2% whereas PCR method detected (MTB) in 15% of the total specimens. Using PCR as gold standard, ZN microscopy correctly identified 20.5% of total (MTB) positive specimens and LJ culture detected 47.7%.Specimen types showed significant association with PCR test: 42.9% of synovial fluid samples and 41.7% of pleural fluid samples; 28.6% of urine samples were positive for (MTB) by PCR method. This indicates that PCR analysis of these specimens’ exhibit greater positivity rates for (MTB) as opposed to CSF and other fluids and biopsies Conclusions: TB PCR is a rapid and reliable test in the diagnosis and management of tuberculosis.

scholarly journals An Altered Mycobacterium tuberculosis Metabolome Induced bykatGMutations Resulting in Isoniazid Resistance

2014 ◽  
Vol 58 (4) ◽  
pp. 2144-2149 ◽  
Author(s):  
Du Toit Loots
Keyword(s):  
Oxidative Stress ◽  
Mycobacterium Tuberculosis ◽  
Principal Component ◽  
Degradation Pathway ◽  
Clinical Samples ◽  
Research Approach ◽  
Isoniazid Resistance ◽  
Content Type ◽  
Genomics And Proteomics ◽  
Resistant Strains

ABSTRACTThe most common form of drug resistance found in tuberculosis (TB)-positive clinical samples is monoresistance to isoniazid. Various genomics and proteomics studies to date have investigated this phenomenon; however, the exact mechanisms relating to how this occurs, as well as the implications of this on the TB-causing organisms function and structure, are only partly understood. Considering this, we followed a metabolomics research approach to identify potential new metabolic pathways and metabolite markers, which when interpreted in context would give a holistic explanation for many of the phenotypic characteristics associated with akatGmutation and the resulting isoniazid resistance inMycobacterium tuberculosis. In order to achieve these objectives, gas chromatography-time of flight mass spectrometry (GCxGC-TOFMS)-generated metabolite profiles from two isoniazid-resistant strains were compared to a wild-type parent strain. Principal component analyses showed clear differentiation between the groups, and the metabolites best describing the separation between these groups were identified. It is clear from the data that due to a mutation in thekatGgene encoding catalase, the isoniazid-resistant strains experience increased susceptibility to oxidative stress and have consequently adapted to this by upregulating the synthesis of a number of compounds involved in (i) increased uptake and use of alkanes and fatty acids as a source of carbon and energy and (ii) the synthesis of a number of compounds directly involved in reducing oxidative stress, including an ascorbic acid degradation pathway, which to date hasn't been proposed to exist in these organisms.

DETECTING MYCOBACTERIUM TUBERCULOSIS RAPIDLY AND MYCOBACTERIUM TUBERCULOSIS STRAINS WITHOUT IS6110 SEQUENCE BY PCR ASSAY

2012 ◽  
pp. 15-19
Author(s):  
Thi Chau Anh Nguyen ◽  
Hoang Bach Nguyen ◽  
Hai Duong Huynh ◽  
Nu Xuan Thanh Le ◽  
Xuan Cuong Le ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
16S Rdna ◽  
False Negative ◽  
Clinical Samples ◽  
Tuberculosis Complex ◽  
Pcr Assay ◽  
Complex Objects ◽  
Method Performance ◽  
Gene Coding ◽  
16S Rdna Pcr

Background: The Nested IS6110 PCR is used for detecting tuberculosis, however IS6110 sequence is not present in the genome of all strains of M.tuberculosis, the result may be false negative. The gene coding 16S ribosome always contains a short sequence specific to M. tuberculosis complex. Objects: Performance of the 16S Real-time PCR to detect M. tuberculosis and combining to the nested IS6110 PCR to determine the rate of Mtb strains without IS6110 from clinical samples. Materials and method: Performance of 16S rDNA PCR by commercial kit of Viet A Inc. for all 480 samples, the samples which were positive with the 16S rDNA PCR were retested in IS6110 PCR assay by in-house kit. Results: The Realtime 16S rDNA PCR detected 258 cases (53.8%) of tuberculosis. There were 3 (1.2 %) M. tuberculosis strains which do not harbor IS6110 sequence in genome. Conclusion: The IS6110 nested PCR can be applied more widely than the 16S rDNA realtime PCR. In case of using IS6110 PCR assay, results may show a low proportion of false negative. Combining 16S rDNA PCR with the IS6110 based PCR allowed detection of deletion of IS6110 sequence in M. tuberculosis isolates.

scholarly journals Mutations in fbiD (Rv2983) as a Novel Determinant of Resistance to Pretomanid and Delamanid in Mycobacterium tuberculosis

2020 ◽  
Vol 65 (1) ◽  
pp. e01948-20
Author(s):  
Dalin Rifat ◽  
Si-Yang Li ◽  
Thomas Ioerger ◽  
Keshav Shah ◽  
Jean-Philippe Lanoix ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Clinical Evidence ◽  
Clinical Samples ◽  
Loss Of Function ◽  
Wild Type ◽  
Content Type ◽  
Solid Media ◽  
High Level ◽  
Level Resistance

ABSTRACTThe nitroimidazole prodrugs delamanid and pretomanid comprise one of only two new antimicrobial classes approved to treat tuberculosis (TB) in 50 years. Prior in vitro studies suggest a relatively low barrier to nitroimidazole resistance in Mycobacterium tuberculosis, but clinical evidence is limited to date. We selected pretomanid-resistant M. tuberculosis mutants in two mouse models of TB using a range of pretomanid doses. The frequency of spontaneous resistance was approximately 10−5 CFU. Whole-genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, of which 91% occurred in 1 of 5 genes previously associated with nitroimidazole activation and resistance, namely, fbiC (56%), fbiA (15%), ddn (12%), fgd (4%), and fbiB (4%). Nearly all mutations were unique to a single mouse and not previously identified. The remaining 9% of resistant mutants harbored mutations in Rv2983 (fbiD), a gene not previously associated with nitroimidazole resistance but recently shown to be a guanylyltransferase necessary for cofactor F420 synthesis. Most mutants exhibited high-level resistance to pretomanid and delamanid, although Rv2983 and fbiB mutants exhibited high-level pretomanid resistance but relatively small changes in delamanid susceptibility. Complementing an Rv2983 mutant with wild-type Rv2983 restored susceptibility to pretomanid and delamanid. By quantifying intracellular F420 and its precursor Fo in overexpressing and loss-of-function mutants, we provide further evidence that Rv2983 is necessary for F420 biosynthesis. Finally, Rv2983 mutants and other F420H2-deficient mutants displayed hypersusceptibility to some antibiotics and to concentrations of malachite green found in solid media used to isolate and propagate mycobacteria from clinical samples.

scholarly journals Detection of mutations associated with isoniazid resistance in multidrug-resistant Mycobacterium tuberculosis clinical isolates

2014 ◽  
Vol 69 (9) ◽  
pp. 2369-2375 ◽  
Author(s):  
Tomasz Jagielski ◽  
Zofia Bakuła ◽  
Katarzyna Roeske ◽  
Michał Kamiński ◽  
Agnieszka Napiórkowska ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Isoniazid Resistance ◽  
Detection Of Mutations
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