Establishment of a Real-time RT-PCR Assay for Avian Encephalomyelitis Virus

Abstract

Avian encephalomyelitis is an epidemic disease caused by avian encephalomyelitis virus (AEV), which exerts egg drop, slow growth, decreased egg hatchability, rapid tremors and ataxia of the head and neck in chicks and laying hens. The availability of a robust diagnostic assay to confirm the infection of AEV is important for its prevention and control. Thus, this study aimed to develop and validate a rapid Real-time RT-PCR assay for the detection and quantification of AEV. The authors developed and validated a TaqMan-based fluorescent real-time RT-PCR with sensitivity, specificity and reproducibility for rapid detection of EAV. This assay was then applied on 52 clinical samples from 37 suspected chickens. The minimal detection limit of this method was 20 copies. No cross-reactivity with Newcastle disease virus (NDV), avian influenza virus (AIV), Marek’s disease virus (MDV) or avian leukosis virus (ALV) was observed. The intra- and inter-assay coefficients of variation were both below 2%. Of the 52 clinical samples analyzed using this assay, 33 samples were positive for AEV, yielding an agreement rate of 91.2% with RT-PCR and 88.2% with the agar diffusion test. In conclusion, we have successfully developed and validated a sensitive Real-time RT-PCR for rapid detection of AEV, which could be used in the early diagnosis, prevention and control, and epidemiological investigation of AEV.