Detection and quantitation of the new world Squash leaf curl virus by TaqMan real-time PCR

Squash leaf curl diseases are caused by distinct virus species that are separated into two major phylogenetic groups, western and eastern hemisphere groups. The western group includes the new world Squash leaf curl virus (SLCV) which causes major losses to cucurbit production and induces severe stunting and leaf curl in squash plants. A TaqMan-based real time polymerase chain reaction (qPCR) assay has been developed for detection and quantitation of SLCV. Designed primers and probe targeted the AV1 (coat protein) gene and in silico analysis showed that they detect a large number of SLCV isolates. The developed assay could detect the virus in 18fg of total nucleic acid and 30 genomic units. The qPCR assay was about 1000 times more sensitive than PCR and amplified successfully SLCV from a wide range of cucurbit hosts and from viruliferous whiteflies. The developed qPCR assay should be suitable for detection and quantitation purposes for all reported SLCV isolates of the western hemisphere.