Detection of rifampin resistance patterns in Mycobacterium tuberculosis strains isolated in Iran by polymerase chain reaction-single-strand conformation polymorphism and direct sequencing methods

Isfahani, Bahram Nasr; Tavakoli, A; Salehi, Mansoor; Mehdi, Tazhibi

doi:10.1590/s0074-02762006000600004

Cited by 22 publications

(17 citation statements)

References 28 publications

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“…Eight studies performed the PCR-SSCP assay on culture iso- lates, and two performed it on sputum samples. Eight studies used a standard phenol-chloroform method to extract chromosomal DNA for PCR amplification (3, 12-14, 16, 18, 31, 37), two studies used the cetyltrimethylammonium bromide (CTAB)-NaCl method (11,17) to extract DNA. Seven studies used the same PCR amplification primers (12-14, 16, 17, 18, 31), and three studies used other primers (3,11,37).…”

Section: Resultsmentioning

confidence: 99%

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PCR-Single-Strand Conformational Polymorphism Method for Rapid Detection of Rifampin-Resistant Mycobacterium tuberculosis : Systematic Review and Meta-Analysis

Jiang

Wang

et al. 2010

J Clin Microbiol

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The reference standard methods for drug susceptibility testing of Mycobacterium tuberculosis, such as culture on Lowenstein-Jensen or Middlebrook 7H10/11 medium, are very slow to give results; and due to the emergence of multidrug-resistant M. tuberculosis and extensively drug-resistant M. tuberculosis, there is an urgent demand for new, rapid, and accurate drug susceptibility testing methods. PCR-single-strand conformational polymorphism (PCR-SSCP) analysis has been proposed as a rapid method for the detection of resistance to rifampin, but its accuracy has not been systematically evaluated. We performed a systematic review and meta-analysis to evaluate the accuracy of PCR-SSCP analysis for the detection of rifampin-resistant tuberculosis. We searched the Medline, Embase, Web of Science, BIOSIS, and LILACS databases and contacted authors if additional information was required. Ten studies met our inclusion criteria for rifampin resistance detection. We applied the summary receiver operating characteristic (SROC) curve to perform the meta-analysis and to summarize diagnostic accuracy

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Section: Resultsmentioning

confidence: 99%

“…The PCR-SSCP assay is relatively simple and was initially promising. Recently, the PCR-SSCP assay has been used in some studies for the rapid detection of rifampin-resistant M. tuberculosis (3,11,12,17). However, individually, these trials found that the sensitivity and specificity were statistically inconsistent.…”

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confidence: 99%

PCR-Single-Strand Conformational Polymorphism Method for Rapid Detection of Rifampin-Resistant Mycobacterium tuberculosis : Systematic Review and Meta-Analysis

Jiang

Wang

et al. 2010

J Clin Microbiol

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“…Specific amino acid substitutions within the regions of the rpoB gene are strongly associated with resistance to other classes of drugs, most notably isoniazid (9,12,18,23,29). Codons 511, 516, 526, and 531 have been reported as the rpoB sites with the most frequent mutations worldwide (1,9,18,20,26,29), although variations in the relative frequencies of the mutations in these codons have been described for M. tuberculosis isolates from different geographic locations (8,18,27,34). These differences reflect the complex and crucial interactions between rifampin and drug targets at the molecular level, where the positions of the affected nucleotide changes seem variable.…”

Section: Discussionmentioning

confidence: 99%

High-Level Rifampin Resistance Correlates with Multiple Mutations in the rpoB Gene of Pulmonary Tuberculosis Isolates from the Afghanistan Border of Iran

et al. 2009

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The aim of this study was to investigate the significance of multiple mutations in the rpoB gene as well as predominant nucleotide changes and their correlation with high levels of resistance to rifampin (rifampicin) in Mycobacterium tuberculosis isolates that were randomly collected from the sputa of 46 patients with primary and secondary cases of active pulmonary tuberculosis from the southern region (Afghanistan border) of Iran where tuberculosis is endemic. Drug susceptibility testing was performed using the CDC standard conventional proportional method. DNA extraction, rpoB gene amplification, and DNA sequencing analysis were performed. Thirty-five (76.09%) isolates were found to have multiple mutations (two to four) in the rpoB (␤-subunit) gene. Furthermore, we demonstrate that the combination of mutations with more prevalent nucleotide changes were observed in codons 523, 526, and 531, indicating higher frequencies of mutations among patients with secondary infection. In this study, 76.08% (n ‫؍‬ 35) of all isolates found to have mutation combinations involving nucleotide changes in codons 523 (GGG3GCG), 531 (TCG3TTG or TTC), and 526 (CAC3CGC,

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“…Several scanning methods are available for detection of drug resistance mutations, including denaturing gradient gel electrophoresis, conformation-sensitive gel electrophoresis, temperature gradient capillary electrophoresis, denaturing high-performance liquid chromatography (dHPLC), and high-density oligonucleotide arrays (15,20,22,30,31,33). These methods vary in sensitivity and generally either are labor-intensive or require sophisticated instrument to perform analyses.…”

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confidence: 99%

Rapid Detection of Isoniazid, Rifampin, and Ofloxacin Resistance in Mycobacterium tuberculosis Clinical Isolates Using High-Resolution Melting Analysis

et al. 2011

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A high-resolution melting analysis (HRMA) assay was developed to detect isoniazid, rifampin, and ofloxacin resistance in Mycobacterium tuberculosis by targeting resistance-associated mutations in the katG, mabA-inhA promoter, rpoB, and gyrA genes. A set of 28 (17 drug-resistant and 11 fully susceptible) clinical M. tuberculosis isolates was selected for development and evaluation of HRMA. PCR amplicons from the katG, mabA-inhA promoter, rpoB, and gyrA genes of all 28 isolates were sequenced. HRMA results matched well with 18 mutations, identified by sequencing, in 17 drug-resistant isolates and the absence of mutations in 11 susceptible isolates. Among 87 additional isolates with known resistance phenotypes, HRMA identified katG and/or mabA-inhA promoter mutations in 66 of 69 (95.7%) isoniazid-resistant isolates, rpoB mutations in 51 of 54 (94.4%) rifampin-resistant isolates, and gyrA mutations in all of 41 (100%) ofloxacin-resistant isolates. All mutations within the HRMA primer target regions were detected as variant HRMA profiles. The corresponding specificities were 97.8%, 100%, and 98.6%, respectively. Most false-positive results were due to synonymous mutations, which did not affect susceptibility. HRMA is a rapid, sensitive method for detection of drug resistance in M. tuberculosis which could be used routinely for screening isolates in countries with a high prevalence of tuberculosis and drug resistance or in individual isolates when drug resistance is suspected.One-third of the world's population is reportedly infected with Mycobacterium tuberculosis, causing high mortality and morbidity. Eight million to nine million new tuberculosis (TB) cases and about 2 million deaths are reported each year (9, 18, 28). Multidrug-resistant TB (MDR-TB) is defined as resistance of the isolate to at least isoniazid (INH) and rifampin (RIF); extensively drug-resistant TB (XDR-TB) is MDR-TB in which the isolate has additional resistance to any of fluoroquinolones and at least one of three injectable second-line antituberculosis drugs used in TB treatment, namely, capreomycin, kanamycin, and amikacin (14, 21). MDR-TB and XDR-TB are major obstacles to the control of TB worldwide. More than 2.5 million patients were diagnosed with active TB in 116 countries and regions in 2006; of them, 4.8% had MDR-TB (3). About one in six (15%) patients with MDR-TB had previously received treatment for TB for at least 1 month (18), and at least 8.0% had XDR-TB, according to incomplete data collected from 37 countries between 2002 and 2007 (41).Drug susceptibility testing using molecular techniques can enhance the identification of drug-resistant M. tuberculosis. Several scanning methods are available for detection of drug resistance mutations, including denaturing gradient gel electrophoresis, conformation-sensitive gel electrophoresis, temperature gradient capillary electrophoresis, denaturing high-performance liquid chromatography (dHPLC), and high-density oligonucleotide arrays (15,20,22,30,31,33). These methods vary in sensitivity and...

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Detection of rifampin resistance patterns in Mycobacterium tuberculosis strains isolated in Iran by polymerase chain reaction-single-strand conformation polymorphism and direct sequencing methods

Abstract: Mutations in the rpoB locus confer conformational changes leading to defective binding of rifampin (RIF)

Cited by 22 publications

References 28 publications

PCR-Single-Strand Conformational Polymorphism Method for Rapid Detection of Rifampin-Resistant Mycobacterium tuberculosis : Systematic Review and Meta-Analysis

PCR-Single-Strand Conformational Polymorphism Method for Rapid Detection of Rifampin-Resistant Mycobacterium tuberculosis : Systematic Review and Meta-Analysis

High-Level Rifampin Resistance Correlates with Multiple Mutations in the rpoB Gene of Pulmonary Tuberculosis Isolates from the Afghanistan Border of Iran

Rapid Detection of Isoniazid, Rifampin, and Ofloxacin Resistance in Mycobacterium tuberculosis Clinical Isolates Using High-Resolution Melting Analysis

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