Dataset Information

Multiplex high-resolution melting analysis as a diagnostic tool for detection of plasmid-mediated AmpC ?-lactamase genes.

ABSTRACT: High-resolution melting (HRM) analysis can be a diagnostic tool to evaluate the presence of resistance genes with the added bonus of discriminating sequence modifications. A real-time, multiplex PCR assay using HRM was designed for the detection of plasmid-mediated ampC genes. The specificity and sensitivity of this assay were 96% and 100%, respectively.

SUBMITTER: Geyer CN

PROVIDER: S-EPMC3993522 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature

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Multiplex high-resolution melting analysis as a diagnostic tool for detection of plasmid-mediated AmpC β-lactamase genes.

Geyer Chelsie N CN Hanson Nancy D ND

Journal of clinical microbiology 20140129 4

High-resolution melting (HRM) analysis can be a diagnostic tool to evaluate the presence of resistance genes with the added bonus of discriminating sequence modifications. A real-time, multiplex PCR assay using HRM was designed for the detection of plasmid-mediated ampC genes. The specificity and sensitivity of this assay were 96% and 100%, respectively. ...[more]

PMID: 24478414

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Project description:Coronavirus disease 2019, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), poses a considerable threat to global public health. This study developed and evaluated a rapid, low-cost, expandable, and sequencing-free high-resolution melting (HRM) assay for the direct detection of SARS-CoV-2 variants. A panel of 64 common bacterial and viral pathogens that can cause respiratory tract infections was employed to evaluate our method's specificity. Serial dilutions of viral isolates determined the sensitivity of the method. Finally, the assay's clinical performance was assessed using 324 clinical samples with potential SARS-CoV-2 infection. Multiplex HRM analysis accurately identified SARS-CoV-2 (as confirmed with parallel reverse transcription-quantitative PCR [qRT-PCR] tests), differentiating between mutations at each marker site within approximately 2 h. For each target, the limit of detection (LOD) was lower than 10 copies/reaction (the LOD of N, G142D, R158G, Y505H, V213G, G446S, S413R, F486V, and S704L was 7.38, 9.72, 9.96, 9.96, 9.50, 7.80, 9.33, 8.25, and 8.25 copies/reaction, respectively). No cross-reactivity occurred with organisms of the specificity testing panel. In terms of variant detection, our results had a 97.9% (47/48) rate of agreement with standard Sanger sequencing. The multiplex HRM assay therefore offers a rapid and simple procedure for detecting SARS-CoV-2 variants. IMPORTANCE In the face of the current severe situation of increasing SARS-CoV-2 variants, we developed an upgraded multiplex HRM method for the predominant SARS-CoV-2 variants based on our original research. This method not only could identify the variants but also could be utilized in subsequent detection of novel variants since the assay has great performance in terms of flexibility. In summary, the upgraded multiplex HRM assay is a rapid, reliable, and economical detection method, which could better screen prevalent virus strains, monitor the epidemic situation, and help to develop measures for the prevention and control of SARS-CoV-2.

Dataset Information

Multiplex high-resolution melting analysis as a diagnostic tool for detection of plasmid-mediated AmpC ?-lactamase genes.

Publications

Multiplex high-resolution melting analysis as a diagnostic tool for detection of plasmid-mediated AmpC β-lactamase genes.

Similar Datasets

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