ИСТИНА

Potato spindle tuber viroid (PSTVd) is a single-stranded, non protein-encoding, covalently-closed circular RNA molecule (359nt) that infects many horticultural and agricultural crops. PSTVd is mechanically transmitted, replicates in the nucleus, and moves cell-to-cell through plasmodesmata. Though it was well established that systemic spread of PSTVd occurs through the phloem sieve elements and was confirmed by modern genetic and microscopic analyses, there is a lack of the reliable data regarding PSTVd detection in xylem cells. In our study, the possible presence of PSTVd RNA in xylem sap was investigated by analysis of guttation fluid (GF) collected from viroid-infected tomato leaves. Our results revealed the absence of PSTVd in GF as confirmed by RT-PCR analysis. Thus, we can conclude that long distance trafficking of PSTVd through tomato plants does not involve xylem vessels. In addition, we applied a modified method based on an EDTA-mediated phloem exudation technique for detection of PSTVd in phloem exudates of tomato plants. Sequencing, RT-PCR and Southern blot analyses demonstrated the presence of viroid RNA in phloem exudates. To our knowledge, this is the first report of PSTVd RNA detection in phloem exudates obtained by the EDTA-mediated exudation technique, which proved to be a simple, inexpensive and reliable method for phloem exudate analysis. These results suggest the potential application of the EDTA-mediated exudation technique and GF analysis for nucleic acid (including signaling RNAs) isolation, and pathogen detection exclusively from phloem or xylem sap, respectively.