Munguti, Florence M., Kilalo, Dora C., Yegon, Hillary K., Macharia, Isaac, Seal, Susan ORCID: 0000-0002-3952-1562 , Mwango’mbe, Agnes W., Nyaboga, Evans N. and Silva, Goncalo ORCID: 0000-0001-5544-2947 (2024) Real-time reverse transcription recombinase polymerase amplification (RT-RPA) assay for detection of cassava brown streak viruses. Scientific Reports, 14:12438. pp. 1-11. ISSN 2045-2322 (Online) (doi:https://doi.org/10.1038/s41598-024-62249-y)

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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.

Item Type:	Article
Uncontrolled Keywords:	cassava brown streak viruses; early virus detection; isothermal amplification; rapid diagnosis; Reverse transcriptase recombinase polymerase amplification (RT-RPA)
Subjects:	Q Science > Q Science (General) Q Science > QR Microbiology Q Science > QR Microbiology > QR355 Virology
Faculty / School / Research Centre / Research Group:	Faculty of Engineering & Science Faculty of Engineering & Science > Natural Resources Institute Faculty of Engineering & Science > Natural Resources Institute > Agriculture, Health & Environment Department Faculty of Engineering & Science > Natural Resources Institute > Molecular Virology and Entomology Research Group
Last Modified:	04 Jun 2024 12:00
URI:	http://gala.gre.ac.uk/id/eprint/47331

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