Defining the relative performance of isothermal assays that can be used for rapid and sensitive detection of foot-and-mouth disease virus

Howson, E. L.A., Kurosaki, Y., Yasuda, J., Takahashi, M., Goto, H., Gray, A. R., Mioulet, V., King, D. P. and Fowler, V. L. (2017) Defining the relative performance of isothermal assays that can be used for rapid and sensitive detection of foot-and-mouth disease virus. Journal of Virological Methods, 249, pp. 102-110. (doi: 10.1016/j.jviromet.2017.08.013) (PMID:28837842)

Abstract

This study describes the first multiway comparison of portable isothermal assays for the detection of foot-and-mouth disease virus (FMDV), benchmarked against real-time reverse transcription RT-PCR (rRT-PCR). The selected isothermal chemistries included reverse transcription loop-mediated isothermal amplification (RT-LAMP) and reverse transcription recombinase polymerase amplification (RT-RPA). The analytical sensitivity of RT-LAMP was comparable to rRT-PCR (10(1) RNA copies), while RT-RPA was one log10 less sensitive (10(2) RNA copies). Diagnostic performance was evaluated using a panel of 35 samples from FMDV-positive cattle and eight samples from cattle infected with other vesicular viruses. Assay concordance for RT-LAMP and RT-RPA was 86-98% and 67-77%, respectively, when compared to rRT-PCR, with discordant samples consistently having high rRT-PCR CT values (no false-positives were detected for any assay). In addition, a hierarchy of sample preparation methods, from robotic extraction to simple dilution of samples, for epithelial suspensions, serum and oesophageal-pharyngeal (OP) fluid were evaluated. Results obtained for RT-LAMP confirmed that FMDV RNA can be detected in the absence of RNA extraction. However, simple sample preparation methods were less encouraging for RT-RPA, with accurate results only obtained when using RNA extraction. Although the evaluation of assay performance is specific to the conditions tested in this study, the compatibility of RT-LAMP chemistry with multiple sample types, both in the presence and absence of nucleic acid extraction, provides advantages over alternative isothermal chemistries and alternative pen-side diagnostics such as antigen lateral-flow devices. These characteristics of RT-LAMP enable the assay to be performed over a large diagnostic detection window, providing a realistic means to rapidly confirm positive FMD cases close to the point of sampling.

• Item Type: Articles
• Keywords: Diagnostics, FMDV, foot-and-mouth disease, RT-LAMP, RT-RPA, rRT-PCR.
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Mioulet, Dr Valerie and King, Dr Donald and Fowler, Dr Veronica and Howson, Emma
• Authors: Howson, E. L.A., Kurosaki, Y., Yasuda, J., Takahashi, M., Goto, H., Gray, A. R., Mioulet, V., King, D. P., and Fowler, V. L.
• College/School: College of Medical Veterinary and Life Sciences; College of Medical Veterinary and Life Sciences > School of Biodiversity, One Health & Veterinary Medicine
• Journal Name: Journal of Virological Methods
• Publisher: Elsevier
• ISSN: 0166-0934
• ISSN (Online): 1879-0984
• Published Online: 31 August 2017
• Copyright Holders: Copyright © 2017 The Authors
• First Published: First published in Journal of Virological Methods 249: 102-110
• Publisher Policy: Reproduced under a Creative Commons license