rpoB mutation

We evaluated Tuberculosis-Spoligo-Rifampicin-Isoniazid Typing (TB-SPRINT), a microbead-based method for spoligotyping and detection of rifampicin and isoniazid resistance in Mycobacterium tuberculosis. For that, 67 M. tuberculosis complex strains were retrospectively selected. Membrane-based spoligotyping, restriction fragment length polymorphism, DNA sequencing/pyrosequencing of rpoB, katG, and inhA promoter, TB-SPRINT, and SNP typing were performed. Concordance between spoligotyping methods was 99.6% (2,785/2,795 spoligotype data points)...

This study aimed to evaluate the frequency of resistance to first- and second-line drugs using phenotypic and genotypic methods and its correlation with resistance-linked mutations in Mycobacterium tuberculosis (M. tb) isolated in Iran. Three different methods, including the indirect proportion method(PM), direct and indirect nitrate reductase assay(NRA), and direct sequencing were used to assess drug resistance. In this study, sensitivity, specificity, agreement, costs, and turnaround time of these methods were compared in 395 smear positive isolates...

INTRODUCTION: Resistance to second line tuberculosis drugs is common, but slow to diagnose with phenotypic drug sensitivity testing. Rapid molecular tests speed up diagnosis, but can only detect limited mutations. Whole genome sequencing (WGS) of culture isolates can generate a complete genetic drug resistance profile, but is delayed by the initial culture step. We previously successfully achieved WGS directly from sputum using targeted enrichment. CASE REPORT: A 29-year-old Nigerian lady was diagnosed with tuberculosis...

BACKGROUND: The GenoType MTBDRplus, a commercial Line Probe Assay (LPA) kit from Hain Lifescience, Germany, is endorsed by India's RNTCP Program for diagnosis of DRTB cases among smear-positive sputum samples. Although the LPA has been studied in several laboratories, there is a wide variation in existing M. tuberculosis strains across the globe, and false results can occur due to the presence of unique genetic mutations in different settings. AIM AND OBJECTIVE: An attempt was made to carry out band pattern analysis using LPA and also to observe uncommon mutations in MDR strains...

BACKGROUND: Multidrug/extensively drug-resistant tuberculosis (M/XDR-TB) is a major public health problem, and early detection is important for preventing its spread. This study aimed to demonstrate the distribution of genetic site mutation associated with drug resistance in M/XDR-TB in the northern Thai population. METHODS: Thirty-four clinical MTB isolates from M/XDR-TB patients in the upper northern region of Thailand, who had been identified for drug susceptibility using the indirect agar proportion method from 2005 to 2012, were examined for genetic site mutations of katG, inhA, and ahpC for isoniazid (INH) drug resistance and rpoB for rifampicin (RIF) drug resistance...

The various aspects of MDR-TB, type of pathogen, different drug sensitive methods and mutation (s) in specific genes were determined. The histone-like protein (hupB) gene of M. tuberculosis was targeted by using primer sets: N & S and M & S (produced 645 bp & 318 bp fragment respectively). The most significant risk factors were the poverty and male gender of ages 11-25 years. All samples were detected as M. tuberculosis. By Drug Proportion method, the highest percentage (37%) was found resistant to only Rifampin...

The relationship between bacterial drug resistance and growth fitness is a contentious topic, but some antibiotic resistance mutations clearly have a fitness cost in the laboratory. Whether these costs translate to deleterious effects in natural habitats is less certain however. Previously, fitness effects of resistance mutations were mostly characterized in nutrient-rich, fast-growth conditions, which bacteria rarely encounter in natural habitats. Carbon, phosphate, iron or oxygen limitations are conditions met by bacterial pathogens in various compartments of the human body...

BACKGROUND: The WHO recently has recommended the GenoType MTBDRplus version 1.0 and MTBDRsl version 1.0 assays for widespread use in countries endemic with drug-resistant tuberculosis. Despite this, these assays have rarely been evaluated in China, where the burden of drug-resistant tuberculosis is among the highest globally. METHODS: Mycobacterium tuberculosis clinical isolates were obtained between January 2008 and December 2008. Isolates were tested for drug resistance against rifampicin (RFP) and isoniazid (INH) using the GenoType MTBDRplus assay and drug resistance against ethambutol (EMB), ofloxacin (OFX), and kanamycin (KM) using the Genotype MTBDRsl assay...

Molecular tests to detect the presence of Mycobacterium tuberculosis and genetic polymorphisms in the rpoB gene conferring resistance to rifampicin (RMP) have become integral parts of tuberculosis diagnostics worldwide. These assays are often performed sequentially or in parallel to phenotypic drug susceptibility testing. Discordances between molecular and phenotypic tests invariably occur. Root causes range from pre-, post- and analytic errors to co-existence of non-tuberculous mycobacteria, silent mutations, mutations outside the 81 base-pair RMP resistance-determining region, non-canonical mutations conferring increased minimal inhibitory concentrations below the critical concentration in some phenotypic drug susceptibility tests, and heteroresistance...

Detection of drug resistance in Mycobacterium tuberculosis by conventional phenotypic drug susceptibility testing methods requires several weeks. Therefore, molecular diagnostic tests for rapid detection of multidrug resistance tuberculosis (MDR-TB) are urgently needed. Early diagnosis helps in initiating optimal treatment which would not only enable cure of an individual patient but also will curb the transmission of drug resistance in the community. Line probe assay (LPA) has shown great promises in the diagnosis of MDR-TB...

OBJECTIVES: Tuberculosis (TB) caused by Mycobacterium tuberculosis (MTB) poses a major public health problem in Nepal. Although it has been reported as one of the dominant genotypes of MTB in Nepal, little information on the Central Asian Strain (CAS) family is available, especially isolates related to multidrug resistance (MDR) cases. This study aimed to elucidate the genetic and epidemiological characteristics of MDR CAS isolates in Nepal. METHODS: A total of 145 MDR CAS isolates collected in Nepal from 2008 to 2013 were characterized by spoligotyping, mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) analysis, and drug resistance-associated gene sequencing...

BACKGROUND: The differentiation of Mycobacterium tuberculosis complex (MTBC) from non-tuberculous mycobacteria (NTM) is of primary importance for infection control and the selection of anti-tuberculosis drugs. Up to date data on rifampicin (RIF)-resistant tuberculosis (TB) is essential for the early management of multidrug-resistant TB. The aim of this study was to evaluate the usefulness of a newly developed multiplexed, bead-based bioassay (Quantamatrix Multiplexed Assay Platform, QMAP) for the rapid differentiation of 23 Mycobacterium species including MTBC and RIF-resistant strains...

Drug-resistant tuberculosis (DR-TB), especially multidrug-resistant tuberculosis (MDR-TB), is a serious medical and societal problem in China. The purpose of this study was to evaluate the mutation characteristics of drug-resistant Mycobacterium tuberculosis (M. tuberculosis) isolates in Yunnan, China. Drug susceptibility testing (DST) was performed in 523 clinical M. tuberculosis isolates. Six drug resistance genes (katG, inhA, rpoB, rpsL, embB, and pncA) were selected to screen for mutations. In total, 54 clinical M...

Recent studies in addition to studies based on whole genome sequencing (WGS) of clinical isolates of Mycobacterium tuberculosis (MTB) from diverse geographical regions have provided useful insights into the mechanisms of drug resistance. Of importance, are some of the findings pertaining to mechanisms of resistance to two of the first-line anti-tuberculosis (TB) drugs, namely, rifampicin (RIF) and pyrazinamide (PZA). For example, the implication of mutations in rpoA and rpoC genes that act as compensatory mutations for those in the rpoB gene with respect to RIF resistance is noteworthy...

Objectives: We previously reported the first case of vancomycin treatment failure due to development of vancomycin-intermediate resistance in a patient with an MRSA of ST72, a community genotype in Korea. We investigated two isogenic MRSA strains from this patient, who experienced treatment failure with vancomycin and rifampicin. Methods: We tracked the genetic alterations that confer reduced susceptibility to vancomycin on those two isogenic MRSA strains by WGS...

BACKGROUND: Drug-resistant tuberculosis, especially multidrug-resistant tuberculosis (MDR-TB), is a major public health problem. Effective management of MDR-TB relies on accurate and rapid diagnosis. In this study, we assessed the diagnostic accuracy of the Genotype MTBDRplus assay in diagnosing MDR-TB in Cameroon, and then discuss on its utility within the diagnostic algorithm for MDR-TB. METHODS: In this cross-sectional study, 225 isolates of Mycobacterium tuberculosis cultured from sputum samples collected from new and previously treated pulmonary tuberculosis patients in Cameroon were used to determine the accuracy of the Genotype MTBDRplus assay...

The objective of the present study was to evaluate the potential development of DNA microarray chips to detect rifampicin (RFP) and isoniazid (INH) resistance in Mycobacterium tuberculosis (MTB), using samples from clinical tuberculosis (TB) patients in Soochow City, China. The sputum samples of 42 patients with TB in the Affiliated Hospital of Infectious Diseases of Soochow University (Soochow, China) were collected. The conventional Lowenstein-Jensen culture medium (Gold Standard) was used to assess drug sensitivity using the absolute concentration method...

Mycobacterium tuberculosis (Mtb) encounters stresses during the pathogenesis and treatment of tuberculosis (TB) that can suppress replication of the bacteria and render them phenotypically tolerant to most available drugs. Where studied, the majority of Mtb in the sputum of most untreated subjects with active TB have been found to be nonreplicating by the criterion that they do not grow as colony-forming units (cfus) when plated on agar. However, these cells are viable because they grow when diluted in liquid media...

Amplicon-based Next Generation Sequencing (NGS) is an emerging method for Mycobacterium tuberculosis drug susceptibility testing (DST) but has not been well described. We examined 158 clinical multidrug-resistant M. tuberculosis isolates via NGS of 11 resistance-associated gene regions covering 3519 nucleotides. Across these gene regions, complete resistance or heteroresistance (defined as 1%-99% mutation) was present in at least one isolate in 6.3% of loci. The number of isolates with heteroresistance was highest for gyrA codon 94, rpoB codons 526 and 531, and embB codons 306, 372 and 406 (range 11-26% of isolates exhibited heteroresistance)...

Mutator strains were identified by screening random Tn5 insertion clones of Caulobacter crescentus. We identified clones with robust increases in mutation rates with Tn5 insertions in the mutY, mutS, mutL and uvrD genes, known to act in mutation-preventing pathways in Escherichia coli. Analysis of mutations in the rpoB gene revealed that in both the parental strain and mismatch repair-deficient mutants, A:T→G:C transitions predominate by a large margin over C:G→T:A. We have also investigated the role of the error-prone polymerase encoded by imuC (dnaE2) in spontaneous mutagenesis, and found that a imuC mutant strain shows mutation rates and sequences comparable to the parental strain...