Real-time reverse transcription recombinase polymerase amplification (RT-RPA) assay for detection of cassava brown streak viruses.

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  • Additional Information
    • Source:
      Publisher: Nature Publishing Group Country of Publication: England NLM ID: 101563288 Publication Model: Electronic Cited Medium: Internet ISSN: 2045-2322 (Electronic) Linking ISSN: 20452322 NLM ISO Abbreviation: Sci Rep Subsets: MEDLINE
    • Publication Information:
      Original Publication: London : Nature Publishing Group, copyright 2011-
    • Subject Terms:
      Manihot*/virology ; Plant Diseases*/virology ; Potyviridae*/genetics ; Potyviridae*/isolation & purification ; Recombinases*/metabolism; RNA, Viral/genetics ; RNA, Viral/isolation & purification ; Real-Time Polymerase Chain Reaction/methods ; Plant Leaves/virology ; Nucleic Acid Amplification Techniques/methods ; Reverse Transcription ; Sensitivity and Specificity ; Reverse Transcriptase Polymerase Chain Reaction/methods
    • Abstract:
      Cassava brown streak disease (CBSD) caused by Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) is the most economically important viral disease of cassava. As cassava is a vegetatively propagated crop, the development of rapid and sensitive diagnostics would aid in the identification of virus-free planting material and development of effective management strategies. In this study, a rapid, specific and sensitive real-time reverse transcription recombinase polymerase amplification (RT-RPA) assay was developed for real-time detection of CBSV and UCBSV. The RT-RPA was able to detect as little as 2 pg/µl of purified RNA obtained from infected cassava leaves, a sensitivity equivalent to that obtained by quantitative real-time reverse transcription PCR (qRT-PCR), within 20 min at 37 °C. Further, the RT-RPA detected each target virus directly from crude leaf and stem extracts, avoiding the tedious and costly isolation of high-quality RNA. The developed RT-RPA assay provides a valuable diagnostic tool that can be adopted by cassava seed certification and virus resistance breeding programs to ensure distribution of virus-free cassava planting materials to farmers. This is the first report on the development and validation of crude sap-based RT-RPA assay for the detection of cassava brown streak viruses (UCBSV and CBSV) infection in cassava plants.
      (© 2024. The Author(s).)
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    • Grant Information:
      BB/ROO5397/1 United Kingdom BB_ Biotechnology and Biological Sciences Research Council; Grant ID: RU/2018/CARP+/04) Regional Universities Forum for Capacity Building in Agriculture (RUFORUM); Grant Number OPP1149777 Bill and Melinda Gates Foundation
    • Contributed Indexing:
      Keywords: Cassava brown streak viruses; Early virus detection; Isothermal amplification; Rapid diagnosis; Reverse transcriptase recombinase polymerase amplification (RT-RPA)
    • Accession Number:
      0 (Recombinases)
      0 (RNA, Viral)
    • Subject Terms:
      Cassava brown streak virus
    • Publication Date:
      Date Created: 20240530 Date Completed: 20240530 Latest Revision: 20240603
    • Publication Date:
      20240603
    • Accession Number:
      PMC11139904
    • Accession Number:
      10.1038/s41598-024-62249-y
    • Accession Number:
      38816439