FIRST REPORT OF CUCUMBER MOSAIC VIRUS IN ANEMONE sp. IN GREECE

Abstract

Virus-like symptoms, such as severe stunting and mosaic, were observed in a greenhouse-grown Anemone sp. (cv. Jerusalem), in Pella (northern Greece) in January 2014. Foliar samples were collected from 20 symptomatic and five symptomless plants and subjected to three initial tests: (i) DAS-ELISA to check for the presence of Cucumber mosaic virus (CMV), (ii) RT-PCR using generic primers for the detection of potyviruses (Pappu et al., 1993) and (iii) PCR for phytoplasmas (Gundersen and Lee, 1996). CMV was detected in all symptomatic plants but i n none of the asymptomatic ones, whereas PCR-based tests for potyviruses or phytoplasmas resulted negative. In order to verify the results of serological tests, a nested RT-PCR was performed on total RNA extracts from all 25 samples. Degenerate primers CMVup624a (5′- ATGGACAAATCTGRATC-3′) and CMVdo1244a (5′- TGRTGCTCRAYGTCKACATGA-3′), used in the first round of PCR were followed by the set CMVup624b (5′- GGACAAATCTGRATCTCCCAATGC-3′) and CMVdo1244b (5′- TGCTCRAYGTCRACATGAAG-3′), designed to amplify a 622 nt- long fragment of the viral coat protein gene. A mplicons of t h e expected size were obtained from all the symptomatic samples but not from the asymptomatic ones. Sequencing of two randomly chosen PCR products (GenBank accession Nos LN810056, LN810057) showed 99% nucleotide identity with CMV isolates from various hosts (KC763473, EU191027). Phylogenetic analyses showed that studied CMV isolates from Anemone sp. belong to sub-Group II. Finally, CMV was mechanically transmitted from the original symptomatic plants onto Cucurbita pepo and Nicotiana benthamiana, and its presence was confirmed by DAS-ELISA. To our knowledge this is the first report of CMV in Anemone sp. in Greece.