Rapid detection of Israeli acute paralysis virus using multi-point ultra-rapid real-time PCR (UR-qPCR)

Abstract

Israeli acute paralysis virus (IAPV) is one of the main pathogens involved in the collapse of honey bee colonies. However, because the virus exhibits a high level of genetic variation and some IAPV strains exhibit high degrees of homology with related viruses, the detection of IAPV in infected honey bees is relatively challenging. To address this obstacle, the aim of the present study was to develop a new detection method that relies on multiple detection sites within the IAPV genome and on Ultra-rapid real-time polymerase chain reaction (UR-qPCR). The new system simultaneously targeted a RNA-dependent RNA polymerase gene (RdRp) and two capsid genes (VP3 and VP1). This multi-point PCR approach was highly efficient, with the ability to detect 100% of IAPV infections, and outperformed single-point PCR, which was only able to detect 86.96–95.6% of IAPV infections. Sequence analysis indicated that RdRp was more variable than the two capsid genes, and the specificity of the proposed method was demonstrated by the detection of IAPV from samples co-infected by IAPV and KBV. Importantly, both freezing-thawing RNA isolation and UR-qPCR could be performed in 27 min and 40 s. Therefore, the present provides an useful tool for the rapid identification of IAPV in apiaries.