PARTIAL MOLECULAR CHARACTERIZATION AND RT-PCR DETECTION OF A PUTATIVE CLOSTEROVIRUS ASSOCIATED WITH OLIVE LEAF YELLOWING

S. Sabanadzovic, N. Abou-Ghanem, P. La Notte, V. Savino, G.P. Martelli, G. Scarito

Abstract


Electrophoretic analysis of extracts from cortical tissues of olive trees of cvs ‘Biancolilla’ and ‘Nostrana’ from Sicily (southern Italy) showing bright chrome yellow discolourations of the leaves, consistently revealed a number of double-stranded RNA (dsRNA) bands, the largest of which had a size (ca 15 kbp) similar to that of the full genomic dsRNA of some species of the genus Closterovirus. A segment of 611 nucleotides, showing sequence homology with the HSP70 homologue gene of Closteroviridae was amplified by RT-PCR from symptomatic trees of both cultivars, using degenerated primers designed on the conserved phosphate 1 and 2 motifs of the HSP70 homologue gene sequence. Computer-assisted phylogenetic analysis showed that the 611 nt HSP70 homologue sequence from olive differed from that of other members of the family Closteroviridae, suggesting that it probably belonged to an undescribed closterovirus, for which the tentative name olive leaf yellowing- associated virus (OLYaV) is proposed. A set of OLYaV-specific primers was designed which amplified a 383 nt fragment of the HSP70 homologue. RT-PCR assays with these primers detected OLYaV sequences in olive trees with leaf yellowing from some Italian regions (Calabria and Latium) and Jerusalem, and in individuals of an unidentified pseudococcid mealybug species and of the psyllid Euphyllura olivina that had fed on symptomatic cv. ‘Biancolilla’ trees. No amplification was obtained with extracts from controls (symptomless olives and apparently healthy olive seedlings) or from olive trees from other Italian regions (Sardinia, Tuscany) that showed somewhat differing yellowing symptoms.

Keywords


olive; leaf yellowing; closterovirus; diagnosis; degenerate primers; RT-PCR; HSP70-like protein; sequencing

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DOI: http://dx.doi.org/10.4454/jpp.v81i1.1044

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