Comparison of three techniques for detection of grapevine leafroll-associated virus 1

Thirty seven plants of grapevine from the Research Station of Viticulture, Karlstejn was examined for the presence of leafroll viruses. Grapevine leafroll-associated virus 1 (GLRaV-1) was detected in the grapevines plants tested using double-antibody sandwich ELISA (DAS-ELISA), RT-PCR and molecular hybridization with non-radioactive RNA probes. Both molecular methods were based on a detection of the GLRaV-1 heat-shock protein 70 (HSP70) gene and showed a higher sensitivity in the detection of GLRaV-1 compared to DAS-ELISA. RNA probes are considered more suitable for the GLRaV-1 detection, as their application can overcome potential minor sequence variability, which may cause the detection by RT-PCR less reliable, especially when the variability occurs in the genome region targeted by RT-PCR primers. Based on additional DAS-ELISA, a mixed infection of GLRaV-1 and Grapevine leafroll-associated virus 3 (GLRaV-3) occurred frequently, while a mixed infection of GLRaV-1 and Grapevine virus A (GVA) or Grapevine fleck virus (GFkV) or a multiple infection of GLRaV-1, GLRaV-3 and GFkV occurred rarely in the tested plants. A mixed infection of all the four viruses mentioned above was not observed.