Multicenter Laboratory Evaluation of the MB/BacT
            <i>Mycobacterium</i>
            Detection System and the BACTEC MGIT 960 System in Comparison with the BACTEC 460TB System for Susceptibility Testing of
            <i>Mycobacterium tuberculosis</i>

Garrigó, Montserrat; Aragón, Lina Marcela; Alcaide, Fernando; Borrell, Sònia; Cardeñosa, Eugenia; Galán, Juan José Unzilla; González-Martín, Juliàn; Martín-Casabona, N; Moreno, Carmen; Salvadó, Margarita; Coll, Pere

doi:10.1128/jcm.02162-06

Cited by 45 publications

(52 citation statements)

References 19 publications

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“…1 and Table 2). Of the 57 clinical strains carrying IGR mutations, 52 (91.22%) were found to be EMB resistant, as determined by the Bactec MGIT 960 method (5 mg/ml) (19). In 275 EMB-resistant strains, 149 strains carried only the embB mutation (e.g., site 306, 406, or 497), 31 strains carried both embC-embA IGR and embB mutations, and 22 strains carried only embC-embA IGR mutations.…”

Section: Resultsmentioning

confidence: 99%

“…To further analyze whether IGR mutations affect transcriptional activity, the IGR mutation region was cloned into the vector pMC210 upstream of the promoterless lacZ gene of vector pMC210 (19). The recombinant M. smegmatis strains transformed with embC-embA IGR mutation construct vectors (mutations of Ϫ8C to T [Ϫ8C-T], Ϫ11C-A, Ϫ11C-T, Ϫ12C-T, Ϫ16C-A, and Ϫ16C-T and the Ϫ27T deletion) had higher levels of ␤-galactosidase activity than the recombinant M. smegmatis with wild-type embC-embA IGR construct vectors (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…All isolates were initially classified as EMB resistant or susceptible in routine diagnostic laboratories by the Bactec MGIT 960 method (5 g/ml) (19). All strains were cultured in a mycobacterial growth indicator tube (MGIT) with the Bactec MGIT 960 growth supplement (Becton Dickinson Diagnostic Systems, MD).…”

Section: Strainsmentioning

confidence: 99%

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Mutations in the embC-embA Intergenic Region Contribute to Mycobacterium tuberculosis Resistance to Ethambutol

Cui

Song

et al. 2014

Antimicrob Agents Chemother

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The rapid increase in Mycobacterium tuberculosis resistance to ethambutol (EMB) threatens the diagnosis and treatment of tuberculosis (TB). We investigated the role of mutations in the embC-embA intergenic region (IGR) in EMB-resistant clinical strains from east China. A total of 767 M. tuberculosis clinical strains were collected and analyzed for their drug susceptibility to EMB using the MGIT 960 system and MIC assay, and the embC-embA IGRs of these strains were sequenced. The transcriptional activity of the embC-embA IGR mutations was examined by reporter gene assays in recombinant Mycobacterium smegmatis strains, and the effect of IGR mutations on its binding to EmbR, a transcription regulator of embAB, was analyzed by gel mobility shift assays. Correlation coefficient analysis showed that the embC-embA IGR mutation is associated with EMB resistance. The clinical strains carrying IGR mutations had a much higher level of embA and embB mRNA as well as higher MICs to EMB. IGR mutations had higher transcriptional activity when transformed into M. smegmatis strains. Mutated IGRs bound to EmbR with much higher affinity than wild-type fragments. The sensitivity of molecular drug susceptibility testing (DST) with IGR mutations as an additional marker increased from 65.5% to 73.5%. Mutations of the embC-embA IGR enhance the binding of EmbR to the promoter region of embAB and increase the expression of embAB, thus contributing to EMB resistance. Therefore, identification of IGR mutations as markers of EMB resistance could increase the sensitivity of molecular DST.T uberculosis (TB) remains a major global health problem. It affects millions of people each year and is the second leading cause of death from an infectious disease worldwide. In 2012, an estimated 8.6 million people developed TB, and 1.3 million died from the disease. The emergence of drug-resistant strains of Mycobacterium tuberculosis, especially those that are multidrug resistant (MDR) and extensively drug resistant (XDR), has posed a serious threat to global TB control programs (1). An estimated 3.6% of new patients and 20.2% of previously treated patients have MDR-TB, and there were 450,000 new cases of MDR-TB worldwide in 2012 (2). Given the alarming rise of drug-resistant TB, the identification of drug resistance genes is critical for the detection and treatment of TB. Much progress has been made to identify gene mutations in specific loci of the M. tuberculosis genome as the molecular basis for TB drug resistance and as drug targets for the development of anti-TB drugs. Although an association of mutations in these resistance genes with drug resistance has been observed, the exact role these genes play in the development of drug resistance is not fully understood. Furthermore, a significant number of anti-TB drug-resistant strains do not carry these mutations, suggesting that unknown gene mutations or variations are involved in the development of anti-TB drug resistance.Ethambutol (EMB) is an essential first-line anti-TB drug that inhibits the biosy...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Mutations in the embC-embA Intergenic Region Contribute to Mycobacterium tuberculosis Resistance to Ethambutol

Cui

Song

et al. 2014

Antimicrob Agents Chemother

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“…The MGIT 960 system is FDA-approved for susceptibility testing of Mtb complex against the first-line drugs including RIF (2 µg/ml), INH (0.4 µg/ml and 0.1 µg/ml), and EMB (7.5 µg/ml and 2.5 µg/ml), PYZ (100 µg/ml) and streptomycin (STR, 6.0 µg/ml and 2.0 µg/ml) (CLSI, 2008). Evaluation of the MGIT performance for the primary tuberculosis drugs showed results that were comparable to the BACTEC 460 radiometric methods as well as the agar proportion methods (>90% agreement) (Ardito et al, 2001;Adjers-Koskela & Katila, 2003;Scarparo et al, 2004), except for EMB and STR which had agreement varying from less than 80% to 98% (Adjers-Koskela & Katila, 2003;Scarparo et al, 2004;Hall et al, 2006;Garrigo et al, 2007). More recently, the MGIT 960 was evaluated for susceptibility testing of second-line drugs including levofloxacin, amikacin, capreomycin and ethionamide with overall agreement of 96% at critical concentrations when compared to the agar proportion method (Lin et al, 2009).…”

Section: Rapid Broth Methodsmentioning

confidence: 84%

Clinical Laboratory Diagnostics for Mycobacterium tuberculosis

Babady¹,

Wengenack²

2012

Understanding Tuberculosis - Global Experiences and Innovative Approaches to the Diagnosis

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“…All samples with a positive growth reading on the MGIT 960 system were tested for acid-fast bacilli by Ziehl-Neelsen stain and for the presence of M. tuberculosis complex DNA using the AccuProbe DNA probe assay (13,14). All MGIT 960 system cultures were subjected to DST against first-line drugs in the MGIT 960 system using the SIRE drug kit, which contains INH at 0.1 mg/liter, RMP at 1.0 mg/liter, streptomycin at 1.0 mg/liter, and ethambutol at 5.0 mg/liter (15). All isolates exhibiting RMP resistance by the Xpert assay and/or MGIT 960 system were tested by an in-house multiplex PCR assay specific for M. tuberculosis complex, the GenoType MDBDRplus assay, and direct DNA sequencing of three (N-terminal, RRDR, and cluster II) regions of the rpoB gene, performed as described previously (13,16).…”

mentioning

confidence: 99%

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

et al. 2015

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Among 452 samples that were positive by the Xpert MTB/RIF (Xpert) assay and MGIT 960 system (MGIT), 440 and 10 Mycobacterium tuberculosis samples 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. rpoB sequencing 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 the rpoB gene. Multidrug-resistant tuberculosis (MDR-TB) (defined as infection with a Mycobacterium tuberculosis strain resistant to at least the two most effective, rifampin and isoniazid, anti-TB drugs) is prevalent throughout the world, difficult to treat, and associated with higher rates of clinical failure and disease relapse (1, 2). The rapid and accurate laboratory diagnosis of MDR-TB is crucial for effective treatment, which will also limit the transmission of MDR-TB (2, 3). The resistance of M. tuberculosis to rifampin (RMP) in nearly 97% of isolates is due to mutations in an 81-bp rifampin resistance-determining region (RRDR) of the rpoB gene (4). Other RMP-resistant isolates contain mutations in either the N-terminal or cluster II region of the rpoB gene, or the resistance is due to other mechanisms (2, 4). Resistance to RMP is a key determinant in treatment failure and also correlates well with MDR-TB, since Ͼ85% of RMP-resistant M. tuberculosis isolates worldwide are also resistant to isoniazid (INH) (2-4). Molecular assays detect mutations in the RRDR of the rpoB gene for the rapid detection of RMP-resistant M. tuberculosis in clinical specimens and culture isolates (2, 3). The World Health Organization (WHO)-approved tests include two line probe assays, the INNO-LiPA Rif. TB (detecting resistance to RMP only) and the GenoType MTBDRplus (detecting resistance to RMP and INH), as well as the real-time PCR-based automated Xpert MTB/RIF (Xpert) assay (detecting resistance to RMP only) (3, 5). However, these tests are not specific, as silent mutations in the rpoB gene occasionally lead to the detection of false-positive RMP resistance (6-9). The current WHO recommendations are to use the Xpert assay as the initial diagnostic test and start treatment for MDR-TB if an RMP resistance result is expected, or, if unexpected, to repeat Xpert assay testing on another sputum sample, particularly in settings in which the prevalence of RMP-resistant TB is Ͻ15% (10). For those settings, treatment for MDR-TB should be initiated when the Xpert assay repeatedly detects RMP resistance. Treatment should be optimized by following susceptibility testing with other first-line and second-line drugs and confirmatory testing for RMP resistance by phenotypic or other genotypic methods; any discordant RMP susceptibility results can be resolved by sequencing of the rpoB gene (10).Phenotypic drug susceptibility testing ...

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Multicenter Laboratory Evaluation of the MB/BacT Mycobacterium Detection System and the BACTEC MGIT 960 System in Comparison with the BACTEC 460TB System for Susceptibility Testing of Mycobacterium tuberculosis

Cited by 45 publications

References 19 publications

Mutations in the embC-embA Intergenic Region Contribute to Mycobacterium tuberculosis Resistance to Ethambutol

Mutations in the embC-embA Intergenic Region Contribute to Mycobacterium tuberculosis Resistance to Ethambutol

Clinical Laboratory Diagnostics for 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

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