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Title: Deep sequencing of viral small-RNAs of citrus tristeza virus (CTV) reveals genomic differences between two Italian isolates of CTV

Author
item LICCIARDELLO, GRAZIA - Parco Scientifico E Tecnologico Della Sicilia
item RUSSO, MARCELLA - Parco Scientifico E Tecnologico Della Sicilia
item GIAMPETRUZZI, ANNALISA - National Research Council - Italy
item LOMBARDO, ALESSANDRO - Parco Scientifico E Tecnologico Della Sicilia
item FERRARO, ROSARIO - Parco Scientifico E Tecnologico Della Sicilia
item SCUDERI, GIUSEPPE - Parco Scientifico E Tecnologico Della Sicilia
item BAR-JOSEPH, MOSHE - Agricultural Research Organization, Volcani Center
item CATARA, ANTONINO - Parco Scientifico E Tecnologico Della Sicilia

Submitted to: Archives of Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/29/2015
Publication Date: 7/15/2015
Citation: Licciardello, G., Russo, M., Giampetruzzi, A., Lombardo, A., Ferraro, R., Scuderi, G., Bar-Joseph, M., Catara, A. 2015. Deep sequencing of viral small-RNAs of citrus tristeza virus (CTV) reveals genomic differences between two Italian isolates of CTV. Archives of Virology. Available: doi: 10.1007/s00705-015-2516-x.

Interpretive Summary: A recent epidemic of Citrus tristeza virus (CTV) in Sicily induced quick decline (QD), resulting in death of many sweet orange trees grafted on sour orange rootstock. Other trees in the same field on the same scion/rootstock combination remained asymptomatic and productive. Since recent reports showed that cross-protection of CTV involves exclusion of a severe CTV strain by a mild strain of the same genotype, a hypothesis was tested to determine if the genotype of the prevalent mild isolate (Bau282) matched that of the prevalent severe isolate, SG29, in the surviving trees. Deep sequencing of viral small RNAs from Hamlin sweet orange trees infected by each isolate was conducted to determine genome sequences. Full-length sequence comparisons showed that the two isolates were different genotypes. Results indicated that SG29 was a severe strain with strong sequence similarly to severe VT genotype strains from Spain and Japan; whereas Bau282 was a mild strain with strong sequence identity to mild T30 genotype isolates from Florida and Spain. Differences in degree of sequence variation between SG29 (with many point mutations) and Bau283 (with only a few mutations) suggested these two CTV genotypes resulted from separate introductions into Sicily. Furthermore, genetic differences between the isolates suggest that Bau282 would not be a good candidate to exclude infection by SG29 and that trees harboring Bau282 have escaped natural challenge from SG29-viruliferous aphid vectors. Therefore, search for mild variants of SG29 have been initiated to find candidate cross-protective isolates against the prevalent severe CTV strain in Sicily.

Technical Abstract: A recent Citrus tristeza virus (CTV) epidemic of quick decline (QD) killed many sweet orange trees grafted on sour orange rootstock in Sicily but left some asymptomatic trees in the same field. Recent reports indicated cross-protection involves exclusion of a severe CTV strain by a mild strain of the same genotype. Therefore, a genetic study of the CTV populations in Sicily was conducted to test the hypothesis that a mild strain was preventing superinfection of a severe strain resulting in surviving vigorous trees by cross-protection. The prevalent severe strain in Sicily was identified as SG29 and the mild strain was identified as Bau282. Deep sequencing of viral small (s)RNAs from Hamlin sweet orange trees infected by each isolate was conducted to determine genome sequence. The proportion of CTV-specific sRNAs reads of total sRNA reads was 53.2% for SG29; whereas that of Bau282 was 37.5%. The ratio of minus to plus strand reads of 21-22 nt CTV sRNAs was 1.8 for SG29 but was equal for Bau282. Overlapping reads were assembled, aligned with extant genome reference isolates, and phylogenetic analysis performed with 32 full-length CTV genomes. The sequence of the mild strain was found to be significantly different from the severe strain, suggesting that Bau282 was not a good candidate to confer cross-protection against SG29. Therefore, a search for mild variants of genotype SG29 has been initiated to find candidate isolates able to cross-protect against the prevalent severe CTV strain in Sicily.