Development of Methodologies for Rapid Detection of Grapevine Viruses

1994 Final Report – A low tittered antiserum to grapevine leafroll associated virus (GLRaV) type I (TI) has been produced. To produce this antiserum, a partially purified virus was used. The antibody produced from this antiserum worked well when used to coat the plate and trap the virus in ELISA assays. Antiserum to a new type of GLRaV has also been produced. The virus has been isolated from a vine with leafroll disease. The produced antiserum has a high titer and did not react with any of the known types of GLRaV (types I, II, III and IV) or GVA. We are in the process of characterizing and identifying this virus type. To overcome the shortage of polyclonal antibodies for GLRaV Til, III and IV, virus from each type has been isolated and purified. Rabbits have been immunized by Til and TIV and we are in the process of preparing and evaluating antisera to these viruses. A polymerase chain reaction (PCR) technique which is a very sensitive and reliable method for the detection of plant viruses has been developed for the detection of grapevine fanleaf virus (GFLV) in grapevine tissue. This technique was able to detect GFLV in a sample when one infected grapevine leaf was mixed with 200 healthy ones. We have modified and simplified the PCR methodology by combining immunology and PCR (immunocapture-PCR, IC-PCR) for the detection of GFLV and GLRaV Till. In this technique the lengthy process of nucleic acid extraction required for PCR has been eliminated. The results showed that this modification works quite well for the detection of these two viruses.