Quantitative one-step real-time RT-PCR for the fast detection of the four genotypes of PPRV

Kwiatek Olivier, Keita Djénéba, Gil Patricia, Fernandez-Pinero Jovita, Jimenez Clavero Michel Angel, Albina Emmanuel, Libeau Geneviève. 2010. Quantitative one-step real-time RT-PCR for the fast detection of the four genotypes of PPRV. Journal of Virological Methods, 165 (2) : 168-177.

https://doi.org/10.1016/j.jviromet.2010.01.014

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Quartile : Q2, Sujet : BIOTECHNOLOGY & APPLIED MICROBIOLOGY / Quartile : Q3, Sujet : BIOCHEMICAL RESEARCH METHODS / Quartile : Q3, Sujet : VIROLOGY

Résumé : A one-step real-time Taqman® RT-PCR assay (RRT-PCR) for peste des petits ruminants virus (PPRV) was developed to detect the four lineages of PPRV by targeting the nucleoprotein (N) gene of the virus. This new assay was compared to a conventional RT-PCR on reference strains and field materials. Quantitation was performed against a standard based on a synthetic transcript of the NPPR gene for which a minimum of 32 copies per reaction were detected with a corresponding Ct value of 39. Depending on the lineage involved, the detection limit of RRT-PCR was decreased by one to three log copies relative to the conventional method. The lower stringency occurred with lineage III because of minor nucleotide mismatches within the probe region. The assay did not detect phylogenetically or symptomatically related viruses of ruminants (such as rinderpest, bluetongue, and bovine viral diarrhea viruses). However, it was capable of detecting 20% more positive field samples with low viral RNA loads compared to the conventional PCR method. When compared on a proficiency panel to the method developed by Bao et al. (2008), the sensitivity of the in-house assay was slightly improved on lineage II. It proved significantly faster to perform and hence better adapted for monitoring large numbers of at risk or diseased animals.

Mots-clés Agrovoc : peste des petits ruminants, génotype, PCR, virus peste petits ruminants, morbillivirus

Classification Agris : L73 - Maladies des animaux
U30 - Méthodes de recherche

Champ stratégique Cirad : Axe 4 (2005-2013) - Santé animale et maladies émergentes

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