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Thirion Laurence, Charrel R. N., Boehmann Y., Corcostegui I., Raoul H., Lamballerie X. de. (2019). Development and evaluation of a duo Zaire ebolavirus Real-Time RT-PCR assay targeting two regions within the genome. Microorganisms, 7 (12), art. 652 [15 p.]

Fichier PDF disponiblehttp://horizon.documentation.ird.fr/exl-doc/pleins_textes/divers20-02/010077829.pdf[ PDF Link ]

Lien direct chez l'éditeur doi:10.3390/microorganisms7120652

En Libre Accès sur HAL https://www.hal.inserm.fr/inserm-02417164

Titre
Development and evaluation of a duo Zaire ebolavirus Real-Time RT-PCR assay targeting two regions within the genome
Année de publication2019
Type de documentArticle référencé dans le Web of Science WOS:000506646400070
AuteursThirion Laurence, Charrel R. N., Boehmann Y., Corcostegui I., Raoul H., Lamballerie X. de.
SourceMicroorganisms, 2019, 7 (12), art. 652 [15 p.]
RésuméPreparedness and response actions to mitigate Ebola virus disease (EVD) outbreaks rely on rapid diagnosis to be implemented locally to sort suspect patients attending health centers. Our aim was (i) to develop and evaluate an RT-qPCR assay combining primers and probes derived from two reference assays targeting different genomic regions; (ii) to study whether sensitivity and specificity of this dual-target assay were at least equal or better to the parental assays; (iii) to implement this dual-target assay onto the Cepheid GeneXpert open cartridge as a proof of principle for technological transfer aiming at bedsite testing locally. To do so, three home-made published RT-qPCR assays were selected to be compared with the RealStar((R)) Filovirus Screen RT-PCR kit 1.0 (Altona Diagnostics, Hamburg, Germany), a technique that was largely deployed during the 2014-2015 West African EVD outbreak. Primers and probes sequences of the custom-made assays were analyzed in silico against a multiple sequence alignment, including >250 complete sequences corresponding to strains that have caused EVD epidemics in the past. Genomic RNA purified from the Mekambo strain of Zaire ebolavirus (EBOV) was used to study the sensitivity of the five methods. Based on these results, two in-house methods were selected and adapted to design the dual-target assay, which performances were compared to those of the parental assays using a synthetic RNA control. The dual-target assay showed better sensitivity and limit of detection (LoD(95) at 0.4 copies/mu L) than the parental methods (1.7 and 2.2 copies/mu L). Ultimately, the dual-target assay was transferred onto the GeneXpert Flex-03 open cartridge, demonstrating a LoD(95) at 0.75 copies/mu L. Together these results indicate that EBOV dual-target assay has the potential to be used during EVD outbreak in the laboratory having performed molecular testing during the recent outbreaks.
Plan de classementEntomologie médicale / Parasitologie / Virologie [052] ; Santé : généralités [050]
Descr. géo.ZAIRE
LocalisationFonds IRD [F B010077829]
Identifiant IRDfdi:010077829
Lien permanenthttp://www.documentation.ird.fr/hor/fdi:010077829

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