Omneity® Pellets
All-In-One Vitamin & Mineral Pellet
Development and evaluation of a reverse transcription loop-mediated isothermal amplification assay for H3N8 equine influenza virus.
Abstract: Reverse transcription loop-mediated isothermal amplification (RT-LAMP) was applied to the detection of equine influenza virus (EIV). Because equine influenza is caused currently by EIV of the H3H8 subtype, the RT-LAMP primer set was designed to target the hemagglutinin gene of this subtype. The detection limit of the RT-LAMP assay was a virus dilution of 10(-5); which was 10(3) times more sensitive than the Espline Influenza A&B-N test and 10 times more sensitive than a reverse transcription polymerase chain reaction (RT-PCR) assay. The specificity of the RT-LAMP assay was examined by using several equine pathogens and nasal swabs collected from horses with fever in 2010 after EIV was eradicated in Japan. No cross-reactions were observed. Using 100 nasal swabs collected from horses with fever during an EIV outbreak in 2007, the RT-LAMP assay detected EIV in 52 samples, whereas the Espline test and the RT-PCR assay detected EIV in only 17 and 41 samples, respectively. These results indicate that the RT-LAMP assay is specific for EIV and more sensitive than the Espline test and the RT-PCR assay. Because it provides high sensitivity and ease of manipulation without the need for a thermal cycler or gel electrophoresis, the RT-LAMP assay should be applicable for laboratory diagnosis of EIV.
Copyright © 2011 Elsevier B.V. All rights reserved.
Publication Date: 2011-08-30 PubMed ID: 21907240DOI: 10.1016/j.jviromet.2011.07.015Google Scholar: Lookup The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
- Evaluation Study
- Journal Article
Summary
This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.
This research investigates a novel technique known as “reverse transcription loop-mediated isothermal amplification” or RT-LAMP, finding it to be both more sensitive and specific than current diagnostic methods for the detection of H3N8 equine influenza virus, the primary cause of equine influenza.
Methodology
- The researchers focused on RT-LAMP, a method of DNA amplification that allows for rapid, simplified testing at a constant temperature, making it potentially more accessible than methods requiring lab-based temperature cycling.
- Specifically, they used this methodology to target the hemagglutinin gene present in the H3N8 subtype of the equine influenza virus (EIV).
- The detection limit of RT-LAMP in this study was one virus particle in a 10^5 dilution. This sensitivity level is significantly higher than standard tests, outperforming the Espline Influenza A&B-N test by a factor of a thousand and a reverse transcription polymerase chain reaction (RT-PCR) – another DNA testing method – by an order of magnitude.
Testing and Results
- The researchers also assessed the specificity of the RT-LAMP test, using it on various equine pathogens and nasal swabs collected from horses that had fever symptoms in 2010, after the EIV had been eradicated in Japan.
- In these tests, the RT-LAMP assay showed no cross-reactivity – it did not give a positive result for other pathogens, indicating a high level of specificity for EIV.
- Comparatively, in a set of 100 nasal swabs from horses during an actual EIV outbreak in 2007, the RT-LAMP test was able to accurately identify EIV in 52 samples, outperforming both the Espline (17 correct detections) and RT-PCR assays (41 correct detections).
Conclusion
- The RT-LAMP test showed high levels of sensitivity; it was able to detect the virus at lower levels than other tests and high specificity; it only provided positive results when the EIV was present.
- Due to the sensitivity, specificity, and relative ease-of-use (it does not require a thermal cycler or gel electrophoresis), the RT-LAMP assay could be particularly useful for lab-based diagnosis of EIV.
Cite This Article
APA
Nemoto M, Yamanaka T, Bannai H, Tsujimura K, Kondo T, Matsumura T.
(2011).
Development and evaluation of a reverse transcription loop-mediated isothermal amplification assay for H3N8 equine influenza virus.
J Virol Methods, 178(1-2), 239-242.
https://doi.org/10.1016/j.jviromet.2011.07.015 Publication
Researcher Affiliations
- Epizootic Research Center, Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. nemoto_manabu@epizoo.equinst.go.jp
MeSH Terms
- Animals
- DNA Primers / genetics
- Horse Diseases / diagnosis
- Horse Diseases / virology
- Horses
- Influenza A Virus, H3N8 Subtype / genetics
- Influenza A Virus, H3N8 Subtype / isolation & purification
- Japan
- Molecular Diagnostic Techniques / methods
- Nasal Mucosa / virology
- Nucleic Acid Amplification Techniques / methods
- Orthomyxoviridae Infections / diagnosis
- Orthomyxoviridae Infections / virology
- Sensitivity and Specificity
- Veterinary Medicine / methods
- Virology / methods
Citations
This article has been cited 11 times.- Knox A, Zerna G, Beddoe T. Current and Future Advances in the Detection and Surveillance of Biosecurity-Relevant Equine Bacterial Diseases Using Loop-Mediated Isothermal Amplification (LAMP). Animals (Basel) 2023 Aug 18;13(16).
- Knox A, Beddoe T. Isothermal Nucleic Acid Amplification Technologies for the Detection of Equine Viral Pathogens. Animals (Basel) 2021 Jul 20;11(7).
- Bakre AA, Jones LP, Bennett HK, Bobbitt DE, Tripp RA. Detection of swine influenza virus in nasal specimens by reverse transcription-loop-mediated isothermal amplification (RT-LAMP). J Virol Methods 2021 Feb;288:114015.
- Ahn SJ, Baek YH, Lloren KKS, Choi WS, Jeong JH, Antigua KJC, Kwon HI, Park SJ, Kim EH, Kim YI, Si YJ, Hong SB, Shin KS, Chun S, Choi YK, Song MS. Rapid and simple colorimetric detection of multiple influenza viruses infecting humans using a reverse transcriptional loop-mediated isothermal amplification (RT-LAMP) diagnostic platform. BMC Infect Dis 2019 Aug 1;19(1):676.
- 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.
- Zhuo Z, Wang J, Chen W, Su X, Chen M, Fang M, He S, Zhang S, Ge S, Zhang J, Xia N. A Rapid On-Site Assay for the Detection of Influenza A by Capillary Convective PCR. Mol Diagn Ther 2018 Apr;22(2):225-234.
- Ashraf W, Unger H, Haris S, Mobeen A, Farooq M, Asif M, Khan QM. Genetic detection of peste des petits ruminants virus under field conditions: a step forward towards disease eradication. BMC Vet Res 2017 Jan 25;13(1):34.
- Yamanaka T, Nemoto M, Bannai H, Tsujimura K, Kondo T, Matsumura T, Gildea S, Cullinane A. Evaluation of twenty-two rapid antigen detection tests in the diagnosis of Equine Influenza caused by viruses of H3N8 subtype. Influenza Other Respir Viruses 2016 Mar;10(2):127-33.
- Nemoto M, Bannai H, Tsujimura K, Kobayashi M, Kikuchi T, Yamanaka T, Kondo T. Getah Virus Infection among Racehorses, Japan, 2014. Emerg Infect Dis 2015 May;21(5):883-5.
- Malanoski AP, Lin B. Evolving gene targets and technology in influenza detection. Mol Diagn Ther 2013 Oct;17(5):273-86.
- Yamanaka T, Nemoto M, Bannai H, Tsujimura K, Kondo T, Matsumura T, Muranaka M, Ueno T, Kinoshita Y, Niwa H, Hidari KI, Suzuki T. No evidence of horizontal infection in horses kept in close contact with dogs experimentally infected with canine influenza A virus (H3N8). Acta Vet Scand 2012 Apr 16;54(1):25.
Use Nutrition Calculator
Check if your horse's diet meets their nutrition requirements with our easy-to-use tool Check your horse's diet with our easy-to-use tool
Talk to a Nutritionist
Discuss your horse's feeding plan with our experts over a free phone consultation Discuss your horse's diet over a phone consultation
Submit Diet Evaluation
Get a customized feeding plan for your horse formulated by our equine nutritionists Get a custom feeding plan formulated by our nutritionists
