rpoB Mutations and Effects on Rifampin Resistance in Mycobacterium tuberculosis

Discordance between Xpert MTB/RIF Assay and Bactec MGIT 960 Culture System for Detection of Rifampin-Resistant Mycobacterium tuberculosis Isolates in a Country with a Low Tuberculosis (TB) Incidence

Journal of Clinical Microbiology ◽

10.1128/jcm.03412-14 ◽

2015 ◽

Vol 53 (4) ◽

pp. 1351-1354 ◽

Cited By ~ 17

Author(s):

Eiman Mokaddas ◽

Suhail Ahmad ◽

Hanaa S. Eldeen ◽

Noura Al-Mutairi

Keyword(s):

Mycobacterium Tuberculosis ◽

Culture System ◽

Content Type ◽

Rifampin Resistance ◽

Xpert Assay

Among 452 samples that were positive by the Xpert MTB/RIF (Xpert) assay and MGIT 960 system (MGIT), 440 and 10Mycobacterium tuberculosissamples were detected as rifampin susceptible and rifampin resistant, respectively. Two isolates that were rifampin susceptible by the MGIT system were rifampin resistant by the Xpert assay.rpoBsequencing identified a silent (CTG521TTG) mutation in one isolate and a missense (GAC516TAC) mutation in another. The detection of rifampin resistance is imperfect with both the Xpert assay and MGIT system. Any discordant rifampin resistance results should be confirmed by sequencing of therpoBgene.

Download Full-text

Amplification Refractory Mutation System – Polymerase Chain Reaction for Rapid Detection of rpoB Gene Mutations in Mycobacterium tuberculosis

International Journal of Applied Sciences and Biotechnology ◽

10.3126/ijasbt.v5i1.17020 ◽

2017 ◽

Vol 5 (1) ◽

pp. 81-85 ◽

Cited By ~ 1

Author(s):

Hemanta Kumari Chaudhary ◽

Mitesh Shrestha ◽

Prakash Chaudhary ◽

Bal Hari Poudel

Keyword(s):

Mycobacterium Tuberculosis ◽

Rpob Gene ◽

Amplification Refractory Mutation System ◽

Chain Reaction ◽

Arms Pcr ◽

Mutation System ◽

Mdr Tb ◽

Total Dna ◽

Polymerase Chain ◽

Rifampin Resistance

Multidrug-resistant tuberculosis (MDR-TB) has become a serious worldwide threat including in Nepal. MDR-TB refers to the pathological condition whereby Mycobacterium tuberculosis becomes resistant to the first line of drug treatment i.e. rifampin and isoniazid. Resistance to rifampin (RIF) is mainly caused by the mutations in the rpoB gene which codes for the β-subunit of RNA polymerase. In this study, Amplification Refractory Mutation System – Polymerase Chain Reaction (ARMS – PCR) technique has been used to detect mutations in the rpoB gene of Mycobacterium tuberculosis. Total DNA samples of 34 phenotypic MDR-TB were subjected to ARMS – PCR using three different codon specific primers (516, 526 and 531). These three codons occupy large portion of total mutation responsible for rifampin resistance. Out of the total DNA samples, all were bearing mutation in at least one of the three codons mentioned. Of those bearing mutation, the highest number had mutation in codon 531 (97.05 %) followed by codon 516 (17.64 %) and finally in codon 526 (11.76%) respectively. Hence, ARMS – PCR may be used as an alternative diagnostic technique for detection of rifampin resistance in Mycobacterium tuberculosis strains, especially for a developing country like Nepal.Int. J. Appl. Sci. Biotechnol. Vol 5(1): 81-85

Download Full-text

Evaluation of Direct Detection of Mycobacterium tuberculosis Rifampin Resistance by a Nitrate Reductase Assay Applied to Sputum Samples in Cotonou, Benin

Journal of Clinical Microbiology ◽

10.1128/jcm.00691-07 ◽

2007 ◽

Vol 45 (7) ◽

pp. 2123-2125 ◽

Cited By ~ 23

Author(s):

D. Affolabi ◽

M. Odoun ◽

A. Martin ◽

J. C. Palomino ◽

S. Anagonou ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Nitrate Reductase ◽

Direct Detection ◽

Nitrate Reductase Assay ◽

Rifampin Resistance

Download Full-text

Molecular Analysis of Codon 548 in therpoBGene Involved in Mycobacterium tuberculosis Resistance to Rifampin

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.04374-14 ◽

2014 ◽

Vol 59 (3) ◽

pp. 1542-1548 ◽

Cited By ~ 7

Author(s):

Yu-Tze Horng ◽

Wen-Yih Jeng ◽

Yih-Yuan Chen ◽

Che-Hung Liu ◽

Horng-Yunn Dou ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Bacterial Resistance ◽

Drug Susceptibility ◽

Content Type ◽

Stable Hydrogen Bond ◽

Resistance Patterns ◽

Resistant Strains ◽

Rifampin Resistance ◽

Susceptibility Tests ◽

Dna Complex

ABSTRACTMostMycobacterium tuberculosisrifampin-resistant strains have been associated with mutations in an 81-bp rifampin resistance-determining region (RRDR) in the generpoB. However, if this region alone were targeted, rifampin-resistant strains with mutations outside the RRDR would not be detected. In this study, among 51 rifampin-resistant clinical isolates analyzed by sequencing 1,681-bp-long DNA fragments containing the RRDR, 47 isolates contained mutations within the RRDR, three isolates contained mutations both within and outside the RRDR, and only one isolate had a single missense mutation (Arg548His) located outside the RRDR. A drug susceptibility test of recombinantMycobacterium smegmatisandM. tuberculosisisolates carrying mutatedrpoB(Arg548His) showed an increased MIC for rifampin compared to that of the control strains. Modeling of the Arg548His mutant RpoB-DNA complex revealed that the His548 side chain formed a more stable hydrogen bond structure than did Arg548, reducing the flexibility of the rifampin-resistant cluster II region of RpoB, suggesting that the RpoB Arg548His mutant does not effectively interact with rifampin and results in bacterial resistance to the drug. This is the first report on the relationship between the mutation in codon 548 of RpoB and rifampin resistance in tuberculosis. The novel mutational profile of therpoBgene described here will contribute to the comprehensive understanding of rifampin resistance patterns and to the development of a useful tool for simple and rapid drug susceptibility tests.

Download Full-text

Rifampin Resistance in Mycobacterium kansasii Is Associated with rpoB Mutations

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.11.3056-3058.2001 ◽

2001 ◽

Vol 45 (11) ◽

pp. 3056-3058 ◽

Cited By ~ 34

Author(s):

John L. Klein ◽

Timothy J. Brown ◽

Gary L. French

Keyword(s):

Mycobacterium Tuberculosis ◽

Molecular Basis ◽

Strong Association ◽

Missense Mutations ◽

Β Subunit ◽

Mycobacterium Kansasii ◽

Bacterial Rna ◽

Rifampin Resistance ◽

Potent Drug

ABSTRACT Rifampin is the most potent drug used in the treatment of disease due to Mycobacterium kansasii. A 69-bp fragment ofrpoB, the gene that encodes the β subunit of the bacterial RNA polymerase, was sequenced and found to be identical in five rifampin-susceptible clinical isolates of M. kansasii. This sequence showed 87% homology with the Mycobacterium tuberculosis gene, with an identical deduced amino acid sequence. In contrast, missense mutations were detected in the same fragment amplified from five rifampin-resistant isolates. A rifampin-resistant strain generated in vitro also harbored an rpoB gene missense mutation that was not present in the parent isolate. All mutations detected (in codons 513, 526, and 531) have previously been described in rifampin-resistant M. tuberculosis isolates. Rifampin MICs determined by E-test were <1 mg/liter for all rifampin-susceptible isolates and >256 mg/liter for all rifampin-resistant ones. In addition, four of the five rifampin-resistant isolates were also resistant to rifabutin. We have thus shown a strong association between rpoB gene missense mutations and rifampin resistance in M. kansasii. Although our results are derived from a small number of isolates and confirmation with larger numbers would be useful, they strongly suggest that mutations within rpoB form the molecular basis of rifampin resistance in this species.

Download Full-text