A reverse transcriptase-loop mediated isothermal amplification assay (RT-LAMP) for rapid detection of bovine viral diarrhea virus 1 and 2

Document Type: Original Articles

Authors

1 Department of Biology, Faculty of Science, Shahid Chamran University of Ahvaz, Ahvaz, Iran

2 Department of Animal Science and Food, Khouzastan Ramin Agriculture and Natural Resources University, Ahvaz, Iran

3 Faculty of Veterinary Medicine, Shahid Chamran University of Ahvaz, Ahvaz, Iran

Abstract

Bovine viral diarrhea virus (BVDV) is a pathogen that infects cattle, and is globally important. It causes substantial financial losses to the livestock industry. In the current study, a one-step reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP) assay was set up for rapid and efficient detection of BVDV. For this purpose, four primers were designed to recognize six distinct regions on the target RNA based on a highly conserved sequence in the 5΄ UTR of the BVDV genome. Eighty blood specimens were collected from bovines suspected to suffer from BVDV infection, and were tested in parallel by RT-LAMP and RT-PCR. Twenty four of these samples were positive by RT-LAMP, while twenty were positive by RT-PCR. The RT-LAMP detection limit was estimated to be approximately 70PFU /mL of virus. Comparison of RT-PCR with RT-LAMP in this study revealed the recent developed RT-LAMP a highly sensitive and specific for BVD virus detection in the clinical samples.

Keywords

Main Subjects


Article Title [French]

Conception et évaluation de la méthode de transcriptase inverse combinée à une réaction d'amplification isothermique par loupe pour détecter le virus de la diarrhée virale bovine

Abstract [French]

Le virus de la diarrhée virale bovine (BVDV) est un agent pathogène chez la vache et a un impact économique important dans le monde. Cette maladie engendre des coûts énormes dans l'industrie du bétail. La méthode de transcriptase inverse en une étape suivie d’une amplification isothermique par loupe (RT-LAMP) a été conçue pour une identification rapide et efficace du virus BVD. Quatre amorces ont été conçues selon la zone non protégée et non traduite de l’extrémité (5'UTR)5' du génome du virus BVD afin de détecter six emplacements différents de l’ARN ciblée. Quatre-vingt échantillons de sang provenant d'animaux soupçonnés d'être porteur du BVD ont été simultanément recueillis et testés en utilisant les deux méthodes de RT-PCR et RT-LAMP. Parmi ces derniers, 24 et 20 échantillons positifs ont été respectivement détectés par les méthodes RT-LAMP et RT-PRC. La limite de détection a été estimée à environ 70PFU/mL pour la méthode RT-LAMP. La comparaison des deux méthodes RT-PRC et RT-LAMP a montré que le RT-LAMP représente une méthode sensible et spécifique pour détecter le virus BVD dans des échantillons cliniques.

Keywords [French]

  • la diarrhée virale bovine
  • transcriptase inverse et l'amplification isothermique par loupe
  • transcriptase inverse-réaction en chaîne de polymerase

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