Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus

Epizootic haemorragic disease (EHD) is an important disease of white-tailed deer and can cause a bluetongue-like illness in cattle. A definitive diagnosis of EHD relies on molecular assays such as real-time RT-qPCR or conventional PCR. Reverse transcription loop-mediated isothermal amplification (RT...

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Autores principales:	Paulina Rajko-Nenow, Emma L. A. Howson, Duncan Clark, Natasha Hilton, Aruna Ambagala, Nicholas Svitek, John Flannery, Carrie Batten
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	RT-LAMP EHDV rapid diagnostics multiplex LAMP AB 7500 fast instrument Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/28838e14fedb44e181c2f5fc6ff9aee6
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spelling	oai:doaj.org-article:28838e14fedb44e181c2f5fc6ff9aee62021-11-25T19:13:11ZDevelopment of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus10.3390/v131121871999-4915https://doaj.org/article/28838e14fedb44e181c2f5fc6ff9aee62021-10-01T00:00:00Zhttps://www.mdpi.com/1999-4915/13/11/2187https://doaj.org/toc/1999-4915Epizootic haemorragic disease (EHD) is an important disease of white-tailed deer and can cause a bluetongue-like illness in cattle. A definitive diagnosis of EHD relies on molecular assays such as real-time RT-qPCR or conventional PCR. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a cost-effective, specific, and sensitive technique that provides an alternative to RT-qPCR. We designed two sets of specific primers targeting segment-9 of the EHD virus genome to enable the detection of western and eastern topotypes, and evaluated their performance in singleplex and multiplex formats using cell culture isolates (n = 43), field specimens (n = 20), and a proficiency panel (n = 10). The limit of detection of the eastern and western RT-LAMP assays was estimated as ~24.36 C<sub>T</sub> and as ~29.37 C<sub>T</sub> in relation to real-time RT-qPCR, respectively, indicating a greater sensitivity of the western topotype singleplex RT-LAMP. The sensitivity of the western topotype RT-LAMP assay, relative to the RT-qPCR assay, was 72.2%, indicating that it could be theoretically used to detect viraemic cervines and bovines. For the first time, an RT-LAMP assay was developed for the rapid detection of the EHD virus that could be used as either a field test or high throughput screening tool in established laboratories to control the spread of EHD.Paulina Rajko-NenowEmma L. A. HowsonDuncan ClarkNatasha HiltonAruna AmbagalaNicholas SvitekJohn FlanneryCarrie BattenMDPI AGarticleRT-LAMPEHDVrapid diagnosticsmultiplex LAMPAB 7500 fast instrumentMicrobiologyQR1-502ENViruses, Vol 13, Iss 2187, p 2187 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	RT-LAMP EHDV rapid diagnostics multiplex LAMP AB 7500 fast instrument Microbiology QR1-502
spellingShingle	RT-LAMP EHDV rapid diagnostics multiplex LAMP AB 7500 fast instrument Microbiology QR1-502 Paulina Rajko-Nenow Emma L. A. Howson Duncan Clark Natasha Hilton Aruna Ambagala Nicholas Svitek John Flannery Carrie Batten Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
description	Epizootic haemorragic disease (EHD) is an important disease of white-tailed deer and can cause a bluetongue-like illness in cattle. A definitive diagnosis of EHD relies on molecular assays such as real-time RT-qPCR or conventional PCR. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a cost-effective, specific, and sensitive technique that provides an alternative to RT-qPCR. We designed two sets of specific primers targeting segment-9 of the EHD virus genome to enable the detection of western and eastern topotypes, and evaluated their performance in singleplex and multiplex formats using cell culture isolates (n = 43), field specimens (n = 20), and a proficiency panel (n = 10). The limit of detection of the eastern and western RT-LAMP assays was estimated as ~24.36 C<sub>T</sub> and as ~29.37 C<sub>T</sub> in relation to real-time RT-qPCR, respectively, indicating a greater sensitivity of the western topotype singleplex RT-LAMP. The sensitivity of the western topotype RT-LAMP assay, relative to the RT-qPCR assay, was 72.2%, indicating that it could be theoretically used to detect viraemic cervines and bovines. For the first time, an RT-LAMP assay was developed for the rapid detection of the EHD virus that could be used as either a field test or high throughput screening tool in established laboratories to control the spread of EHD.
format	article
author	Paulina Rajko-Nenow Emma L. A. Howson Duncan Clark Natasha Hilton Aruna Ambagala Nicholas Svitek John Flannery Carrie Batten
author_facet	Paulina Rajko-Nenow Emma L. A. Howson Duncan Clark Natasha Hilton Aruna Ambagala Nicholas Svitek John Flannery Carrie Batten
author_sort	Paulina Rajko-Nenow
title	Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
title_short	Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
title_full	Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
title_fullStr	Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
title_full_unstemmed	Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus
title_sort	development of a novel loop mediated isothermal amplification assay (lamp) for the rapid detection of epizootic haemorrhagic disease virus
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/28838e14fedb44e181c2f5fc6ff9aee6
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Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus

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