First Report of Cucumber leaf spot virus in Spain

Cucumber leaf spot virus (CLSV), reclassified as a species in the new genus Aureusvirus (family Tombusviridae) (1), has ≈30-nm isometric particles with a ≈4.4-kb positive-sense, single-stranded RNA. CLSV is transmitted by the chytrid fungus Olpidium bornovanus. The virus has been reported in Germany, Great Britain, Greece, Jordan, and Saudi Arabia. During the fall of 2000, abundant chlorotic spots with necrotic centers were observed on the leaves of cucumber plants grown in a commercial greenhouse in Granada (southeastern Spain). When sap from collected leaves was used to mechanically inoculate cucumber, symptoms were reproduced and were suggestive of CLSV. Based on the nucleic acid sequence of CSLV (2), the following specific primers were designed: CLSVU1440 (5'-AAGGTAGGGGAGATCTTG-3') and CLSVA2160 (5'-GCTTCGGCTGATTCTGA-3'). When used in reverse transcription-polymerase chain reactions (RT-PCR), leaves expressing symptoms yielded amplification products of the expected size (720 bp). These products were cloned into a pGEM-T vector and sequenced (GenBank Accession No. AY038365). The similarity of the nucleic acid and derived amino acid sequences with the one published for CLSV (2) was 94.5 and 99.1%, respectively. The amino acid sequence was 86% identical to that of Pothos latent virus (GenBank Accession No. AJ243370). Ten cucumber plants grown in vermiculite supplemented with rhizosphere soil (1/30, vol/wt) from infected plants developed symptoms on leaves after 1 month and were positive for CLSV when leaf and root tissues were analyzed by RT-PCR and Southern blot hybridization. Plants grown in vermiculite alone did not become infected with CLSV. Microscopic examination of root tissue revealed O. bornovanus only in infected plants. To our knowledge, this is the first record of CSLV in Spain. References: (1) G. P. Martelli et al. Arch. Virol. 143:1847, 1998. (2) J. S. Miller et al. Virus Res. 52:51, 1997.