High prevalence of viruses in table grape from Spain detected by real-time RT-PCR

Table grapes from one of the most important growing area in Spain (Vinalopó, Alicante) protected by the Designation of Origin “Vinalopó bagged table grape”, were surveyed and analysed to determine the prevalence of the five viruses included in the Spanish certification program: Arabis mosaic virus (ArMV), Grapevine fanleaf virus (GFLV), Grapevine fleck virus (GFkV), Grapevine leafroll associated virus-1 (GLRaV-1) and Grapevine leafroll associated virus-3 (GLRaV-3). Ninety five sampling points were selected and the position of grapevine plants georeferenced. Samples were collected in two different vegetative periods and analyses were performed by ELISA and real-time RT-PCR. Purified RNA and immobilized viral targets from plant extracts on nylon membranes were used in parallel assays as templates for PCR assays. In order to analyse these five viral species by real-time RT-PCR, new specific primers and TaqMan probes were designed for detection of ArMV and GFkV. Real time RT-PCR from purified RNA was more sensitive than spot version and ELISA tests. The most prevalent virus was GFLV (95.8%) followed by GLRaV-3 (94.7%), GLRaV-1 (66.3%) and GFkV (65.3%). ArMV was not detected in any sample. The high level of viral infections and the presence of mixed infections suggest that initial infected plant material and uncontrolled traffic of propagation material have played an important role in the spread of viruses.