First Record of Columnea latent viroid (CLVd) in Tomato in Italy

During spring-summer 2010 a serious disease was observed in protected tomato crops in Castrovillari, Calabria Region, southern Italy. Tomato plants (Solanum lycopersicum 'Santa') showing leaf malformation, bronzing in addition to stunting and necrosis, were submitted to laboratory analyses. Apparently, affected plants were estimated visually to be around 80% in the crop. This unusual symptomatology was noted since 2008 on tomato crop in the same greenhouse but on sporadic plants. Sap from diseased plants was inoculated on a set of indicator plants and used in ELISA tests to check for viruses infections including Tomato mosaic virus (ToMV), Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Potato virus Y (PVY), Potato virus X (PVX), Tomato spotted wilt virus (TSWV), Pepino mosaic virus (PepMV), Tomato yellow leaf curl virus (TYLCV), Tomato yellow leaf curl Sardinia virus (TYLCSV), Pelargonium zonate spot virus (PZSV). The plant samples were also tested by conventional RT-PCR, using a primer pair described for Potato spindle tuber viroid (PSTVd). Moreover, cuttings from the diseased plants were grafted on healthy tomato plants. Reaction of indicator plants suggested a ToMV infection which was confirmed by serological test. At same time, the RT-PCR gave an amplicon of about 370 bp. The amplicon was cloned and sequenced. Surprising, the sequence obtained was 98% identical to a sequence of Columnea latent viroid (CLVd). The amplification of CLVd with primers described for PSTVd, could be explained because of the good sequence homology between the two viroids in the same genomic region of the primers adopted. This is the first record of CLVd in tomato in Italy and this new finding confirms once again that this viroid posed a threat to tomato crops, since it was previously detected in tomato also in The Netherlands, France, Belgium and the United Kingdom. The possible modalities of diffusion of this viroid are proposed and discussed.