Single-tube protocol for culture-independent spoligotyping of Mycobacterium tuberculosis based on MeltArray.

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  • Author(s): Xia Z;Xia Z; Su B; Su B; Tu C; Tu C; Sun S; Sun S; Tan Y; Tan Y; Xu Y; Xu Y; Li Q; Li Q
  • Source:
    Journal of clinical microbiology [J Clin Microbiol] 2024 Jan 17; Vol. 62 (1), pp. e0118323. Date of Electronic Publication: 2023 Dec 19.
  • Publication Type:
    Journal Article; Research Support, Non-U.S. Gov't
  • Language:
    English
  • Additional Information
    • Source:
      Publisher: American Society for Microbiology Country of Publication: United States NLM ID: 7505564 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1098-660X (Electronic) Linking ISSN: 00951137 NLM ISO Abbreviation: J Clin Microbiol Subsets: MEDLINE
    • Publication Information:
      Original Publication: Washington, American Society for Microbiology.
    • Subject Terms:
      Mycobacterium tuberculosis*/genetics ; Tuberculosis*/microbiology; Humans ; Molecular Epidemiology/methods ; Real-Time Polymerase Chain Reaction ; Multiplex Polymerase Chain Reaction ; Bacterial Typing Techniques/methods ; Genotype
    • Abstract:
      Importance: Spacer oligonucleotide typing (spoligotyping), the first-line genotyping assay for Mycobacterium tuberculosis (MTB), plays a fundamental role in the investigation of its epidemiology and evolution. In this study, we established a single-tube spoligotyping assay using MeltArray, a highly multiplex polymerase chain reaction (PCR) approach that runs on a real-time PCR thermocycler. The MeltArray protocol included an internal positive control, gyrB , to indicate the abundance of MTB via the quantification cycle and 43 spacers to identify the spoligotype via melting curve analysis. The entire protocol was completed in a single step within 2.5 hours. The lowest detectable copy number for the tested strains was 20 copies/reaction and thus sufficient for analyzing both culture and sputum samples. We conclude that MeltArray-based spoligotyping could be used immediately in low- and middle-income countries with a high tuberculosis burden, given its easy access, improved throughput, and potential applicability to clinical samples.
      Competing Interests: The authors declare no conflict of interest.
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    • Grant Information:
      2023G051 Fujian Provincial Department of Science and Technology; 3502Z20191007 Xiamen Municipal Bureau of Science and Technology; JCYJ20180306170526435 Shenzhen Municipal Science and Technology Innovation Council; 2022CXY0113 Xiamen Municipal Bureau of Science and Technology; 2021ZD0006 Department of Science and Technology of Inner Mongolia Autonomous Region; 2021Y4001 Fujian Provincial Department of Science and Technology
    • Contributed Indexing:
      Keywords: MeltArray; Mycobacterium tuberculosis; spoligotyping; sputum
    • Publication Date:
      Date Created: 20231219 Date Completed: 20240118 Latest Revision: 20240620
    • Publication Date:
      20240620
    • Accession Number:
      PMC10793361
    • Accession Number:
      10.1128/jcm.01183-23
    • Accession Number:
      38112521