Development of a recombinase polymerase amplification combined with a lateral flow dipstick assay for rapid detection of the Mycoplasma bovis.

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  • Additional Information
    • Source:
      Publisher: BioMed Central Country of Publication: England NLM ID: 101249759 Publication Model: Electronic Cited Medium: Internet ISSN: 1746-6148 (Electronic) Linking ISSN: 17466148 NLM ISO Abbreviation: BMC Vet Res Subsets: MEDLINE
    • Publication Information:
      Original Publication: London : BioMed Central, 2005-
    • Subject Terms:
      Cattle Diseases/*diagnosis ; Molecular Diagnostic Techniques/*veterinary ; Mycoplasma Infections/*veterinary ; Nucleic Acid Amplification Techniques/*veterinary; Animals ; Cattle ; China ; DNA-Directed DNA Polymerase/metabolism ; Mycoplasma Infections/diagnosis ; Mycoplasma bovis/genetics ; Nucleic Acid Amplification Techniques/standards ; Recombinases/metabolism ; Sensitivity and Specificity
    • Abstract:
      Background: Mycoplasma bovis (M. bovis) is a major etiological agent of bovine mycoplasmosis around the world. Point-of-care testing in the field is lacking owing to the requirement for a simple, robust field applicable test that does not require professional laboratory equipment. The recombinase polymerase amplification (RPA) technique has become a promising isothermal DNA amplify assay for use in rapid and low-resource diagnostics.
      Results: Here, a method for specific detection of M. bovis DNA was established, which was RPA combined with lateral flow dipstick (LFD). First, the analytical specificity and sensitivity of the RPA primer and LF-probe sets were evaluated. The assay successfully detected M. bovis DNA in 30 min at 39 °C, with detection limit of 20 copies per reaction, which it was compared the real-time quantitative PCR (qPCR) assay. This method was specific because it did not detect a selection of other bacterial pathogens in cattle. Both qPCR and RPA-LFD assays were used to detect M. bovis 442 field samples from 42 different dairy farms in Shandong Province of China, also the established RPA-LFD assay obtained 99.00% sensitivity, 95.61% specificity, and 0.902 kappa coefficient compared with the qPCR.
      Conclusions: To the author's knowledge, this is the first report using an RPA-FLD assay to visualise and detect M. bovis. Comparative analysis with qPCR indicates the potential of this assay for rapid diagnosis of bovine mycoplasmosis in resource limited settings.
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    • Grant Information:
      31672556 the National Natural Science Fund of China
    • Contributed Indexing:
      Keywords: Isothermal nucleic acid amplification; Lateral flow dipstick; Mycoplasma bovis; Rapid and visual detection; Recombinase polymerase amplification
    • Accession Number:
      0 (Recombinases)
      EC 2.7.7.7 (DNA-Directed DNA Polymerase)
    • Publication Date:
      Date Created: 20181222 Date Completed: 20190111 Latest Revision: 20200225
    • Publication Date:
      20240105
    • Accession Number:
      PMC6302395
    • Accession Number:
      10.1186/s12917-018-1703-x
    • Accession Number:
      30572884