Sensitive and Specific TaqMan Real-Time PCR Assay for Beak and Feather Disease Virus in Psittacine Birds

Beak and feather disease virus (BFDV) is a small, icosahedral, non-enveloped virus with a circular single-stranded DNA genome, that belongs to the Circoviridae family. BFDV is globally distributed and poses a major threat to susceptible avian populations. This underscores the urgent need for rapid and reliable molecular detection methods to monitor and control the spread of the virus. Quantitative PCR assays offer several advantages, including high sensitivity, specificity, and the ability to quantify viral load. Here, we report the development and validation of a quantitative real-time PCR assay using a TaqMan probe targeting the replicase gene to detect BFDV in psittacine blood samples. The assay demonstrates high sensitivity, specificity, and suitability for routine diagnostic use with an LOD and an LOQ of 10 and 30 plasmid copies, respectively, determined using a recombinant plasmid containing a BFDV genomic fragment. The assay achieved 98.8% sensitivity and 100% specificity. The method also demonstrated strong repeatability and reproducibility, with intra- and inter-assay coefficients of variation of 4.14% and 4.44%, respectively. Both values are well below the acceptance threshold. Overall, this TaqMan-based real-time PCR assay is a reliable, sensitive, and efficient diagnostic tool for detecting BFDV in psittacine samples.