Journal article
A quantitative, real-time polymerase chain reaction assay for beak and feather disease virus
Journal of Virological Methods, Vol.159(1), pp.98-104
2009
Abstract
PCR-based assays for the detection of BFDV DNA are in widespread use throughout the world. Quantitative real-time PCR assays are extremely sensitive and have the advantages over standard PCR assays that they do not require post-reaction processing to visualise PCR products and can quantify the amount of DNA present in a sample. This study describes a quantitative real-time PCR assay for the detection of BFDV DNA, using primers designed to amplify a conserved 81 bp fragment of ORFV1 and SYTO9, a fluorescent intercalating dye. A synthetic oligonucleotide was used to make standard curves for the quantitation of viral load in blood and feather preparations. The assay was very sensitive, with a detection limit of 50 copies/μL. The assay was developed using DNA extracts from the feathers of 10 different species of birds which had tested BFDV-positive previously and was validated with blood and feather samples from corellas vaccinated with an experimental BFDV vaccine, then challenged with live virus. Viral DNA was detected consistently in the blood of all control (non-vaccinated) birds and in some vaccinated birds. Contamination of the environment with feather dander from BFDV-infected birds meant that feather preparations used for the haemagglutination assay were unreliable for the detection and quantitation of viral excretion. Nonetheless, the assay should prove to be a useful and sensitive test for the detection of viral DNA in a range of samples.
Details
- Title
- A quantitative, real-time polymerase chain reaction assay for beak and feather disease virus
- Authors/Creators
- P.L. Shearer (Author/Creator) - Murdoch UniversityM. Sharp (Author/Creator) - Murdoch UniversityN. Bonne (Author/Creator) - Murdoch UniversityP. Clark (Author/Creator) - Murdoch UniversityS.R. Raidal (Author/Creator) - Charles Sturt University
- Publication Details
- Journal of Virological Methods, Vol.159(1), pp.98-104
- Publisher
- Elsevier BV
- Identifiers
- 991005544993807891
- Copyright
- © 2009 Elsevier B.V.
- Murdoch Affiliation
- State Agricultural Biotechnology Centre; School of Veterinary and Biomedical Sciences
- Language
- English
- Resource Type
- Journal article
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- Collaboration types
- Domestic collaboration
- Citation topics
- 1 Clinical & Life Sciences
- 1.104 Virology - General
- 1.104.1933 Porcine Viral Challenges
- Web Of Science research areas
- Biochemical Research Methods
- Biotechnology & Applied Microbiology
- Virology
- ESI research areas
- Microbiology