Diagnostic accuracy of a duplex real-time reverse transcription quantitative PCR assay for detection of African horse sickness virus.
Abstract: Blood samples collected from 503 suspect cases of African horse sickness (AHS) and another 503 from uninfected, unvaccinated South African horses, as well as 98 samples from horses from an AHS free country, were tested with an AHS virus (AHSV) specific duplex real-time reverse transcription quantitative PCR (RT-qPCR) assay and virus isolation (VI). The diagnostic sensitivity and specificity of this AHSV RT-qPCR assay and VI were estimated using a 2-test 2-population Bayesian latent class model which made no assumptions about the true infection status of the tested animals and allowed for the possibility of conditional dependence (correlation) in test results. Median diagnostic sensitivity and specificity of the AHSV RT-qPCR were 97.8% and 99.9%, respectively. Median diagnostic specificity of virus isolation was >99% whereas the estimated diagnostic sensitivity was 44.2%. The AHSV RT-qPCR assay provides for rapid, high-throughput analysis of samples, and is both analytically and diagnostically sensitive and specific. This assay is potentially highly useful for demonstrating freedom or infection of horses with AHSV, thus it is appropriate that its reproducibility be evaluated in other laboratories as a global standard for detection of AHSV.
Copyright © 2012 Elsevier B.V. All rights reserved.
Publication Date: 2013-01-03 PubMed ID: 23291102DOI: 10.1016/j.jviromet.2012.12.014Google Scholar: Lookup
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- Evaluation Study
- Journal Article
- Research Support
- Non-U.S. Gov't
- African Horse Sickness
- Bayesian Analysis
- Blood Analysis
- Clinical Study
- Diagnosis
- Diagnostic Imaging
- Diagnostic Technique
- Disease
- Disease Diagnosis
- Epidemiology
- Equine Health
- Horses
- Infection
- Infectious Disease
- Laboratory Methods
- Polymerase Chain Reaction
- Real-Time PCR
- Veterinary Medicine
- Veterinary Research
- Virology
- Virus
Summary
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The research paper analyzes the effectiveness of a diagnostic test, called a “duplex real-time reverse transcription quantitative PCR assay,” in detecting African horse sickness virus (AHSV). The study found that this assay demonstrated a high degree of accuracy in the detection of the virus.
Sample Collection and Testing
- Blood samples were collected from 503 suspected cases of African horse sickness, 503 uninfected and unvaccinated horses from South Africa, as well as 98 samples from horses in a country free from this disease.
- These samples were then tested using two methods: a duplex real-time reverse transcription quantitative PCR assay (AHSV RT-qPCR) and virus isolation (VI).
Estimation of Diagnostic Sensitivity and Specificity
- The study estimates the diagnostic sensitivity and specificity of the AHSV RT-qPCR assay and VI through a 2-test 2-population Bayesian latent class model.
- This method makes no assumptions about the true infection status of the animals tested. It also allows for the possibility of correlation in test results, a conditional dependence.
Findings of the Study
- The median diagnostic sensitivity of the AHSV RT-qPCR was 97.8%, while its specificity was 99.9%.
- The virus isolation method demonstrated a median diagnostic specificity of >99%, but its sensitivity was significantly lower at 44.2%.
- These results show that the AHSV RT-qPCR assay is both analytically and diagnostically sensitive and specific.
- Additionally, this method allows for rapid, high-throughput analysis of samples.
Conclusion and Potential Use
- The study concludes that the AHSV RT-qPCR assay is useful in demonstrating whether horses are free from or infected with the African horse sickness virus.
- It suggests further validation of this assay in other laboratories worldwide to establish it as a global standard for detecting AHSV.
Cite This Article
APA
Guthrie AJ, Maclachlan NJ, Joone C, Lourens CW, Weyer CT, Quan M, Monyai MS, Gardner IA.
(2013).
Diagnostic accuracy of a duplex real-time reverse transcription quantitative PCR assay for detection of African horse sickness virus.
J Virol Methods, 189(1), 30-35.
https://doi.org/10.1016/j.jviromet.2012.12.014 Publication
Researcher Affiliations
- Equine Research Centre, Faculty of Veterinary Science, University of Pretoria, Private Bag X04, Onderstepoort 0110, South Africa. alan.guthrie@up.ac.za
MeSH Terms
- Africa
- African Horse Sickness / blood
- African Horse Sickness / diagnosis
- African Horse Sickness / virology
- African Horse Sickness Virus / genetics
- African Horse Sickness Virus / isolation & purification
- Animals
- Horses
- Limit of Detection
- RNA, Viral / analysis
- RNA, Viral / genetics
- Reverse Transcriptase Polymerase Chain Reaction / methods
- Reverse Transcriptase Polymerase Chain Reaction / veterinary
- Sensitivity and Specificity
Citations
This article has been cited 17 times.- Penzhorn L, Crafford JE, Guthrie AJ. Enhancing African horse sickness virus detection: comparing and adapting PCR assays. J Vet Diagn Invest 2026 Feb 7;:10406387261417355.
- Morales J, Ruano MJ, Tena-Tomás C, van Schalkwyk A, Loundras EA, Valero-Lorenzo M, López-Herranz A, Romito M, Batten C, Villalba R, Agüero M. Modification and Validation of a Reference Real-Time RT-PCR Method for the Detection of a New African Horse Sickness Virus Variant. Microorganisms 2025 Nov 25;13(12).
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- Bustin SA. RT-qPCR Testing and Performance Metrics in the COVID-19 Era. Int J Mol Sci 2024 Aug 28;25(17).
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