Real-time RT-PCR for detection of equine influenza and evaluation using samples from horses infected with A/equine/Sydney/2007 (H3N8).
Abstract: Equine influenza (EI) virus (H3N8) was identified in the Australian horse population for the first time in August 2007. The principal molecular diagnostic tool used for detection was a TaqMan real-time reverse transcription-polymerase chain reactions (RT-PCR) assay specific for the matrix (MA) gene of influenza virus type A (IVA). As this assay is not specific for EI, we developed a new EI H3-specific TaqMan assay targeting the haemagglutinin (HA) gene of all recent EI H3 strains. The IVA and the EI H3 TaqMan assays were assessed using in vitro transcribed RNA template, virus culture, diagnostic samples from the outbreak and samples from experimentally infected horses. The EI H3 TaqMan assay had a higher diagnostic sensitivity (DSe) when compared to the IVA TaqMan assay and also when using a conventional PCR for EI H3 as a standard of comparison. The performance of both TaqMan assays was compared with an antigen detection ELISA and virus isolation using nasal swabs collected daily from horses experimentally infected with the outbreak strain A/equine/Sydney/2888-8/2007. The EI H3 TaqMan assay was the most sensitive of the assays, able to detect EI from day 1 or 2 post-challenge, as early as virus isolation, and before clinical signs of disease were observed.
Publication Date: 2008-12-11 PubMed ID: 19153018DOI: 10.1016/j.vetmic.2008.12.006Google Scholar: Lookup
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- Journal Article
Summary
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The research focuses on the development of a new diagnostic tool for equine influenza (EI), particularly the H3N8 strain. The newly developed tool, an EI H3-specific TaqMan assay, was found to have a higher diagnostic sensitivity when compared to existing diagnostic methods.
Development of a new diagnostic tool for Equine Influenza
- The H3N8 strain of the Equine Influenza virus was detected in the Australian horse population for the first time in August 2007.
- The researchers aimed to develop a more specific diagnostic tool to enhance the detection of this particular strain. The existing matrix (MA) gene detection method, although useful, was not specific for the EI virus.
- A new EI H3-specific TaqMan assay was developed by the researchers targeting the haemagglutinin (HA) gene of recent EI H3 strains. This assay turned out to have increased diagnostic sensitivity compared to the existing tool.
Assessment and Comparison of Diagnostic Tools
- The researchers assessed the efficiency of the newly developed EI H3 TaqMan assay in comparison with the existing IVA TaqMan assay using different testing materials.
- These materials included in vitro transcribed RNA template, virus culture, and diagnostic samples from an outbreak and from experimentally infected horses.
- It was found that the EI H3 TaqMan assay demonstrated a higher diagnostic sensitivity compared to the IVA TaqMan assay and conventional PCR for EI H3.
Examining the Performance of the EI H3 TaqMan Assay
- Additional comparison was conducted between the newly developed assay and an antigen detection ELISA and virus isolation method.
- This comparison involved nasal swabs collected daily from horses experimentally infected with the outbreak strain A/equine/Sydney/2888-8/2007.
- The new EI H3 TaqMan assay was proven to be the most sensitive tool among the compared assays, allowing for early detection of EI within 1 or 2 days post-exposure.
- This proved to be as early as traditional virus isolation and even before clinical signs of the disease could be observed.
Cite This Article
APA
Foord AJ, Selleck P, Colling A, Klippel J, Middleton D, Heine HG.
(2008).
Real-time RT-PCR for detection of equine influenza and evaluation using samples from horses infected with A/equine/Sydney/2007 (H3N8).
Vet Microbiol, 137(1-2), 1-9.
https://doi.org/10.1016/j.vetmic.2008.12.006 Publication
Researcher Affiliations
- CSIRO Livestock Industries, Australian Animal Health Laboratory, 5 Portarlington Road (Private Bag 24), Geelong, VIC 3220, Australia.
MeSH Terms
- Animals
- Base Sequence
- Genes, Viral
- Horse Diseases / diagnosis
- Horse Diseases / virology
- Horses
- Influenza A Virus, H3N8 Subtype / genetics
- Influenza A Virus, H3N8 Subtype / isolation & purification
- Orthomyxoviridae Infections / diagnosis
- Orthomyxoviridae Infections / veterinary
- Orthomyxoviridae Infections / virology
- Reverse Transcriptase Polymerase Chain Reaction / methods
- Reverse Transcriptase Polymerase Chain Reaction / veterinary
- Sensitivity and Specificity
- Viral Regulatory and Accessory Proteins
Citations
This article has been cited 8 times.- Alaql FA, Alhafufi AN, Kasem S, Alhammad YMO, Albaqshi H, Alyousaf A, Alsubaie FM, Alghamdi AN, Abdel-Moneim AS, Alharbi SA. Full-Length Genome of the Equine Influenza A Virus Subtype H3N8 from 2019 Outbreak in Saudi Arabia. Animals (Basel) 2022 Oct 10;12(19).
- Ghoniem SM, El Deeb AH, Aggour MG, Hussein HA. Development and evaluation of a multiplex reverse-transcription real-time PCR assay for detection of equine respiratory disease viruses. J Vet Diagn Invest 2018 Nov;30(6):924-928.
- Singh RK, Dhama K, Karthik K, Khandia R, Munjal A, Khurana SK, Chakraborty S, Malik YS, Virmani N, Singh R, Tripathi BN, Munir M, van der Kolk JH. A Comprehensive Review on Equine Influenza Virus: Etiology, Epidemiology, Pathobiology, Advances in Developing Diagnostics, Vaccines, and Control Strategies. Front Microbiol 2018;9:1941.
- Gildea S, Garvey M, Lyons P, Lyons R, Gahan J, Walsh C, Cullinane A. Multifocal Equine Influenza Outbreak with Vaccination Breakdown in Thoroughbred Racehorses. Pathogens 2018 Apr 17;7(2).
- Skjold P, Sommerset I, Frost P, Villoing S. Vaccination against pancreas disease in Atlantic salmon, Salmo salar L., reduces shedding of salmonid alphavirus. Vet Res 2016 Aug 5;47(1):78.
- Firestone SM, Cogger N, Ward MP, Toribio JA, Moloney BJ, Dhand NK. The influence of meteorology on the spread of influenza: survival analysis of an equine influenza (A/H3N8) outbreak. PLoS One 2012;7(4):e35284.
- Firestone SM, Christley RM, Ward MP, Dhand NK. Adding the spatial dimension to the social network analysis of an epidemic: investigation of the 2007 outbreak of equine influenza in Australia. Prev Vet Med 2012 Sep 15;106(2):123-35.
- Lu Z, Chambers TM, Boliar S, Branscum AJ, Sturgill TL, Timoney PJ, Reedy SE, Tudor LR, Dubovi EJ, Vickers ML, Sells S, Balasuriya UB. Development and evaluation of one-step TaqMan real-time reverse transcription-PCR assays targeting nucleoprotein, matrix, and hemagglutinin genes of equine influenza virus. J Clin Microbiol 2009 Dec;47(12):3907-13.
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