Development and validation of a one-step triplex real-time rt-pcr assay for the diagnosis of foot-and-mouth disease virus - Anses - Agence nationale de sécurité sanitaire de l’alimentation, de l’environnement et du travail Accéder directement au contenu
Communication Dans Un Congrès Année : 2021

Development and validation of a one-step triplex real-time rt-pcr assay for the diagnosis of foot-and-mouth disease virus

Résumé

Rapid and accurate laboratory detection of foot-and-mouth disease virus (FMDV) is an essential step in the control of the disease. In recent years, studies have focused on the development of molecular diagnostic methods that improved considerably the laboratory diagnosis of FMDV. Indeed, the detection of viral genomes by real-time RT-PCR is one of the most efficient, sensitive and rapid diagnostic method. In this study, a one-step triplex TaqMan® real-time RT-PCR assay for foot-and-mouth disease virus detection was developed and fully validated. This new tool allows the simultaneous detection of two of the most conserved FMDV coding regions: the RNA polymerase (3D) and the non-structural multifunctional protein (2B), and the GAPDH housekeeping gene as internal control of the RNA extraction step. This triplex was validated according to the OIE principles of validation of diagnostic assays and to the U47-600 French standard. Using field epithelium and nasal swab samples, the diagnostic sensitivity is 98% for the 2B target and 100% for the 3D target, so 100% for the triplex (95% confidence interval: 88,26 - 98,29% and 96,23 - 100% respectively). The diagnostic specificity is 100% for the tested species (i.e. cattle, goats, sheep and pigs). Inter-laboratory reproducibility was evaluated by providing a ready-to-use master mix and blind samples to ten European FMD national reference laboratories. The inter-laboratory coefficient of variation ranged from 0.16 - 8.39% and 0.01 - 9.37% for targets 2B and 3D respectively. This new tool has high specificity and sensitivity and should improve FMD diagnosis by allowing a rapid and accurate detection of FMDV in cases of clinical suspicion.
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Dates et versions

anses-04523929 , version 1 (27-03-2024)

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  • HAL Id : anses-04523929 , version 1

Citer

Cindy Bernelin-Cottet, Souheyla Benfrid, Anne-Laure Salomez, Aurore Romey, Anthony Relmy, et al.. Development and validation of a one-step triplex real-time rt-pcr assay for the diagnosis of foot-and-mouth disease virus. GFRA online meeting 2021, Global Foot-and-Mouth-Disease Research Alliance, Nov 2021, on-line meeting, Argentina. ⟨anses-04523929⟩
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