A rapid RT-LAMP assay for the detection of all four lineages of Peste des Petits Ruminants Virus

• Published in: Journal of virological methods 2019-12, Vol.274, p.113730-113730, Article 113730
• Main Authors: Rajko-Nenow, Paulina, Flannery, John, Arnold, Hannah, Howson, Emma L.A., Darpel, Karin, Stedman, Anna, Corla, Amanda, Batten, Carrie
• Format: Article
• Language: English
• Subjects: Animals; Diagnostics; DNA Primers - genetics; Goat Diseases - diagnosis; Goats; Molecular Diagnostic Techniques - methods; Nucleic Acid Amplification Techniques - methods; Outbreak; Peste-des-Petits-Ruminants - diagnosis; Peste-des-petits-ruminants virus - genetics; Peste-des-petits-ruminants virus - isolation & purification; PPRV; RT-LAMP; Sensitivity and Specificity; Surveillance; Time Factors
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2019.113730

[Display omitted] •Designed N-gene RT-LAMP using all full-genomes available on Genbank.•The RT-LAMP showed comparable diagnostic sensitivity to the real-time RT-PCR assay.•The limit of detection was estimated between 0.3 to 0.8 TCID50 ml−1.•The RT-LAMP could detect PPRV as early as 4 dpi – before clinical signs were observed.•The RT-LAMP assay can support the global PPR eradication campaign. Peste des petits ruminants (PPR) is a viral disease of small ruminants that is caused by the PPR virus (PPRV) and is a significant burden on subsistence farmers across the developing world. Loop-mediated isothermal amplification (LAMP) provides cost-effective, rapid, specific and sensitive detection of nucleic acid and has been demonstrated to have field application for a range of viruses. We describe the development of a novel PPRV RT-LAMP assay utilising carefully-selected primers (targeting the N-gene) allowing for the detection of all known PPRV lineages in < 20 min. The assay was evaluated in comparison with a “gold standard” real-time RT-PCR assay using more than 200 samples, comprising samples from recent PPRV outbreaks, experimentally-infected goats, well-characterised cell culture isolates and samples collected from uninfected animals. The RT-LAMP assay demonstrated 100% diagnostic specificity and greater than 97% diagnostic sensitivity in comparison with the real-time RT-PCR assay. The limit of detection was between 0.3 and 0.8 log10 TCID50 ml−1 equating to a CT value of 31.52 to 33.48. In experimentally-infected animals, the RT-LAMP could detect PPRV as early as 4 days post infection (dpi) - before clinical signs were observed at 7 dpi. The RT-LAMP assay can support the global PPR eradication campaign.