Dataset Information

Mutations Found in embCAB, embR, and ubiA Genes of Ethambutol-Sensitive and -Resistant Mycobacterium tuberculosis Clinical Isolates from China.

ABSTRACT: To better understand the molecular mechanisms of Ethambutol (EMB) resistance, the mutant hot spot region of five genes (embB, embA, embC, embR, and ubiA) was amplified and sequenced in 109 EMB-resistant and 153 EMB-susceptible clinical isolates from China. Twenty-seven EMB-susceptible isolates were found to have nonsynonym mutations, 23 of which were in embB. The mutations occurred most frequently in embB (85.3%, 93) and were seldom in embC (2.8%, 3), embA (3.7%, 4), embR (3.7%, 4), and ubiA (8.3%, 9) in EMB-resistant isolates. For the embB gene, 63 isolates showed mutations at embB306, 20 at embB406, nine at embB497, and five at embB354 in EMB-resistant isolates. In addition, the particular mutants at embB406 and embB497 indicated both high levels of EMB resistance (MICs > 5 ?g/mL) and broad anti-TB drug resistance spectrums. Our data supported the facts that embB306 could be used as a marker for EMB resistance with a sensitivity of 57.8% and a specificity of 78.8%.

SUBMITTER: Xu Y

PROVIDER: S-EPMC4568347 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Mutations Found in embCAB, embR, and ubiA Genes of Ethambutol-Sensitive and -Resistant Mycobacterium tuberculosis Clinical Isolates from China.

Xu Yuhui Y Jia Hongyan H Huang Hairong H Sun Zhaogang Z Zhang Zongde Z

BioMed research international 20150831

To better understand the molecular mechanisms of Ethambutol (EMB) resistance, the mutant hot spot region of five genes (embB, embA, embC, embR, and ubiA) was amplified and sequenced in 109 EMB-resistant and 153 EMB-susceptible clinical isolates from China. Twenty-seven EMB-susceptible isolates were found to have nonsynonym mutations, 23 of which were in embB. The mutations occurred most frequently in embB (85.3%, 93) and were seldom in embC (2.8%, 3), embA (3.7%, 4), embR (3.7%, 4), and ubiA (8. ...[more]

PMID: 26417605

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Project description:BackgroundEthambutol (EMB) is a first-line drug for the treatment of tuberculosis (TB). Resistance to EMB in Mycobacterium tuberculosis isolates is mediated by mutations in several genes involved in arabinan synthesis notably three emb (arabinosyl transferase) and iniA (isoniazid-inducible) genes. Most epidemiologically unrelated EMB-resistant M. tuberculosis strains contain mutations at embB codons 306, 406 and 497, embC-embA intergenic region (IGR) and iniA codon 501 (iniA501).ObjectiveTo develop a more comprehensive molecular screen for EMB-resistance detection among epidemiologically unrelated EMB-resistant M. tuberculosis strains previously analyzed for embB codon 306, 406 and 497 mutations by including analysis of mutations at iniA501 and in embC-embA IGR.MethodsFifty consecutive and phenotypically documented EMB-resistant and 25 pansusceptible M. tuberculosis strains isolated from 75 different TB patients over a four-year period in Kuwait were analyzed. Mutations at iniA501 were detected by PCR amplification followed by restriction fragment length polymorphism (RFLP) patterns generated with Hpy 99 I. Direct DNA sequencing was used to confirm RFLP results and for detecting mutations in embC-embA IGR.ResultsNearly same number of EMB-resistant M. tuberculosis strains were resistant to EMB alone and EMB together with additional resistance to rifampicin and isoniazid (9 of 50, 18% and 11 of 50, 22%, respectively). All the 25 pansusceptible strains contained wild-type sequences at iniA501 and in embC-embA IGR. The analysis of 50 EMB-resistant M. tuberculosis isolates showed that only one strain contained a mutated iniA501 while no mutation was detected in embC-embA IGR in any of the isolate.ConclusionAnalysis of iniA501 and embC-embA IGR in epidemiologically unrelated EMB-resistant M. tuberculosis isolates in Kuwait indicate that mutations at these locations occur very infrequently and their inclusion for the development of a comprehensive molecular screen will make only minor contribution towards rapid EMB resistance detection.

Project description:BackgroundStreptomycin (SM) is a broad spectrum antibiotic and is an important component of any anti-tuberculosis therapy regimen. Several mechanisms have been proposed to explain the emergence of resistance but still our knowledge is inadequate. Proteins form a very complex network and drugs are countered by their modification/efflux or over expression/modification of targets. As proteins manifest most of the biological processes, these are attractive targets for developing drugs, immunodiagnostics or therapeutics. The aim of present study was to analyze and compare the protein profile of whole cell extracts from Mycobacterium tuberculosis clinical isolates susceptible and resistant to SM.ResultsTwo-dimensional gel electrophoresis (2DE) and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry was employed for analyzing the protein profiles. Homology and in silico characterization for identified proteins was assessed using BLAST, InterProScan and KEGG database searches. Computational studies on the possible interactions between SM and identified proteins were carried out by a battery of online servers and softwares, namely, CLUSTALW (KEGG), I-TASSER, VMD, PatchDock and FireDock. On comparing 2DE patterns, nine proteins were found consistently overexpressed in SM resistant isolates and were identified as Rv0350, Rv0440, Rv1240, Rv3075c, Rv2971, Rv3028c, Rv2145c, Rv2031c and Rv0569. In silico docking analysis showed significant interactions of SM with essential (Rv0350, Rv0440 and Rv2971) and non essential (Rv1240, Rv3075c and Rv2031c) genes.ConclusionsThe computational results suggest high protein binding affinity of SM and suggested many possible interactions between identified proteins and the drug. Bioinformatic analysis proves attributive for analysis of diversity of proteins identified by whole proteome analysis. In-depth study of the these proteins will give an insight into probable sites of drug action other than established primary sites and hence may help in search of novel chemotherapeutic agents at these new sites as inhibitors.

Dataset Information

Mutations Found in embCAB, embR, and ubiA Genes of Ethambutol-Sensitive and -Resistant Mycobacterium tuberculosis Clinical Isolates from China.

Publications

Mutations Found in embCAB, embR, and ubiA Genes of Ethambutol-Sensitive and -Resistant Mycobacterium tuberculosis Clinical Isolates from China.

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