Development of Novel Diagnostic and Vaccine Options for Beak and Feather Disease Virus (BFDV)

PCR-based assays for the detection of BFDV DNA are in widespread use throughout the world. Quan-titative real-time PCR assays are extremely sensitive and have the advantages over standard PCRassays that they do not require post-reaction processing to visualise PCR products and can quan-tify the amount of DNA present in a sample. This study describes a quantitative real-time PCR assayfor the detection of BFDV DNA, using primers designed to amplify a conserved 81bp fragment ofORFV1 and SYTO9, a fluorescent intercalating dye. A synthetic oligonucleotide was used to makestandard curves for the quantitation of viral load in blood and feather preparations. The assay wasvery sensitive, with a detection limit of 50 copies/ L. The assay was developed using DNA extractsfrom the feathers of 10 different species of birds which had tested BFDV-positive previously andwas validated with blood and feather samples from corellas vaccinated with an experimental BFDVvaccine, then challenged with live virus. Viral DNA was detected consistently in the blood of all con-trol (non-vaccinated) birds and in some vaccinated birds. Contamination of the environment withfeather dander from BFDV-infected birds meant that feather preparations used for the haemaggluti-nation assay were unreliable for the detection and quantitation of viral excretion. Nonetheless, theassay should prove to be a useful and sensitive test for the detection of viral DNA in a range of sam-ples.© 2009 Elsevier B.V. All rights reserved.