Immunocapture-RT-PCR assay for detection and molecular epidemiology studies of Rabbit Haemorrhagic Disease and European Brown Hare Syndrome viruses.
Rabbit Haemorrhagic Disease Virus and European Brown Hare Syndrome Virus are two members of the genus Lagovirus in the family Caliciviridae. They are the causative agents of highly contagious and fatal diseases of rabbits and hares respectively. We adjusted one assay for the detection and the genomic characterisation of each virus, based on viral purification by immunocapture and genomic amplification by reverse transcription-polymerase chain reaction (IC-RT-PCR). It is carried out directly with the liver exudate obtained after thawing and suppresses the viral nucleic acid preparation step. This assay combines the advantages of an ELISA test (rapidity) because immunocapture and the RT reaction were carried out in the same microtitre plate, and the advantages of PCR (sensitivity). The procedure described allows the processing of large numbers of samples and is suitable for phylogenetic studies of lagomorphs caliciviruses. In addition, it was compared with sandwich-ELISA used for Rabbit Haemorrhagic Disease or European Brown Hare Syndrome diagnosis. A good correlation was found between ELISA and IC-RT-PCR results for Rabbit Haemorrhagic Disease diagnosis, whereas for European Brown Hare Syndrome diagnosis, the results confirmed the higher sensitivity of the molecular method.
Le Gall-Reculé
G
G
Agence Française de Sécurité Sanitaire des Aliments, Laboratoire d'études et de recherches avicoles et porcines, B.P. 53, 22440, Ploufragan, France. g.legall@ploufragan.afssa.fr
Zwingelstein
F
F
Portejoie
Y
Y
Le Gall
G
G
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis / Q000276:immunology; D017250:Caliciviridae Infections / Q000175:diagnosis* / Q000276:immunology / Q000662:veterinary / Q000821:virology*; D002645:Chickens; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006168:Guinea Pigs; D017929:Hemorrhagic Disease Virus, Rabbit / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D010802:Phylogeny; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral