Dataset Information

Evaluation of the invader assay with the BACTEC MGIT 960 system for prompt isolation and identification of mycobacterial species from clinical specimens.

ABSTRACT: Rapid and accurate identification of mycobacterial species is essential for patient management. We describe the use of the Invader assay in conjunction with the BACTEC MGIT 960 system that together provide an efficient procedure for clinical use. This assay discriminates single-base differences (e.g., genotyping single-nucleotide polymorphisms) under homogeneous and isothermal conditions and can measure directly on genomic DNA without prior target DNA amplification. To identify a wide variety of mycobacterial species, 20 Invader probes were designed to target the 16S rRNA gene and the 16S-23S rRNA gene internal transcribed spacer 1 (ITS-1) region. To validate the Invader probes, we used 78 ATCC strains, and 607 clinical mycobacterial strains, which were identified by DNA sequencing of the 16S rRNA gene and ITS-1. The Invader assay could accurately identify and differentiate these strains according to target sequences. Moreover, it could detect and identify 116 (95.1%) of 122 positive liquid cultures from the BACTEC MGIT 960 system and did not react to 83 contaminated MGIT cultures. Species identification takes 6.5 h by the Invader assay: 2.0 h for DNA extraction, 0.5 h for handling, and up to 4 h for the Invader reaction. The Invader assay has the speed, ease of use, and accuracy to be an effective procedure for the bacteriological diagnosis of mycobacterial infections.

SUBMITTER: Ichimura S

PROVIDER: S-EPMC2045373 | biostudies-literature | 2007 Oct

REPOSITORIES: biostudies-literature

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Evaluation of the invader assay with the BACTEC MGIT 960 system for prompt isolation and identification of mycobacterial species from clinical specimens.

Ichimura Sadahiro S Nagano Makoto M Ito Nobuko N Shimojima Masahiro M Egashira Toru T Miyamoto Chikara C Ohkusu Kiyofumi K Ezaki Takayuki T

Journal of clinical microbiology 20070808 10

Rapid and accurate identification of mycobacterial species is essential for patient management. We describe the use of the Invader assay in conjunction with the BACTEC MGIT 960 system that together provide an efficient procedure for clinical use. This assay discriminates single-base differences (e.g., genotyping single-nucleotide polymorphisms) under homogeneous and isothermal conditions and can measure directly on genomic DNA without prior target DNA amplification. To identify a wide variety of ...[more]

PMID: 17687020

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Project description:This study aimed to assess the possible causes of discordant results between Xpert MTB/RIF (Xpert) and Bactec MGIT 960 Culture System (MGIT960) regarding rifampicin (RIF) susceptibility in Mycobacterium tuberculosis. Patients with previous RIF-resistant tuberculosis who were admitted to Wenzhou Central Hospital from January 2020 to December 2022 were enrolled. The isolates obtained from these patients were subjected to RIF susceptibility tests using Xpert and MGIT960, and the minimum inhibitory concentration (MIC) of RIF was determined by the MYCOTB MIC plate test. Additionally, molecular docking and molecular dynamics (MD) simulations were performed to evaluate the binding efficacy of rpoB and RIF based on rpoB mutations detected in the isolates with discordant RIF susceptibility results. A total of 28 isolates with discordant RIF susceptibility test results were detected, 15 of them were RIF susceptible with MICs ≤ 0.5 µg/mL. Twelve out of 15 isolates contained borderline RIF resistance-associated mutations [L430P (n = 6), H445N (n = 6)], 1 isolate had D435Y and Q429H double mutation, and the remaining 2 isolates had a silent (Q432Q) mutation. Compared with the affinity of RIF toward the wild type (WT) (-45.83 kcal/mol) by MD, its affinity toward L452P (-55.52 kcal/mol), D435Y (-47.39 kcal/mol), L430P (approximately -69.72 kcal/mol), H445N (-49.53 kcal/mol), and Q429H (-55.67 kcal/mol) increased. Borderline RIF resistance-associated mutations were the main cause for the discordant RIF susceptibility results between Xpert and MGIT960, and the mechanisms of the resistance need further investigated.IMPORTANCEThis study is aimed at assessing discordant results between Xpert MTB/RIF (Xpert) assay and Bactec MGIT 960 Culture System (MGIT960) regarding the detection of rifampicin (RIF)-resistant Mycobacterium tuberculosis isolates in Wenzhou, China. The discordant results of RIF between these two assays were mainly caused by borderline RIF resistance-associated mutations, subsequently by silent mutations of rpoB. Borderline RIF resistance- associated mutations detected in our study were demonstrated to not be affected by the affinity of rpoB and RIF by molecular dynamics, and the mechanism of resistance was needed to be clarified. For the discordant results of RIF by Xpert and MGIT960 that occurred, rpoB DNA sequencing was recommended to investigate its association with resistance to RIF.

Dataset Information

Evaluation of the invader assay with the BACTEC MGIT 960 system for prompt isolation and identification of mycobacterial species from clinical specimens.

Publications

Evaluation of the invader assay with the BACTEC MGIT 960 system for prompt isolation and identification of mycobacterial species from clinical specimens.

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