Detection protocols and in vitro eradication of the principal Ranunculus asiaticus L. viruses

The buttercup (Ranunculus asiaticus L.) belongs to the family of Ranuncolaceae and is a popular ornamental plant cultivated for the sale of cut flowers and for the production of potted or border plants. This species is native to the eastern Mediterranean basin and has become widespread in Western Europe, South Africa, California, Israel and Japan. New cultivars for the production of cut flowers are selected and multiplied by micropropagation combined with an additional year of rhizome development. The clonal lines have reached high quality and high economic value. Ranunculus hybrids are susceptible to infection by several viruses all associated with economically important diseases therefore the propagation material should be produced totally virus-free. In the present study, the main viruses infecting Ranunculus hybrids, grown in the greenhouse in the Liguria region (Imperia province), have been evaluated in leaf tissues by double step RT-PCR assays using specific primers. Moreover, it has been set up a protocol for in vitro viruses eradication by meristem tips excision combined with thermotherapy. Results show the presence of some viral species (Ranunculus mild mosaic virus (RMMV), Ranunculus leaf distortion virus (RLDV), Ranunculus white mottle virus (RWMV) and others in the samples analyzed, either in single or in mixed infection, and confirm that although the serological diagnosis (ELISA) is in many ways sufficient for the identification of viruses, the molecular evaluation, due to versatility of use and reliability of results, can be easily used as a tool for viruses detection on large scale propagation materials in order to prevent the spread of the principal viruses.