Abstract

Two dsRNA molecules with an estimated lenght of 1.5 Kbp were identified and characterized from leaves of a Japanese persimmon (Diospyros kaki) tree, showing veinlets necrosis on both sides of leaf blades.DOP-PCR recognized two genomic fragments of a bipartite cryptic virus, for which the name of Persimmon cryptic virus (PeCV) is proposed. RLM-RACE leaded to the sequencing of a 1,510 bp contig identified as dsRNA-1 and a 1,491 bp complete segment identified as dsRNA-2. The two genomic fragments resulted both monocistronic and harbored conserved domains related to RNA-dependent RNA polymerase (RdRP) and capsid protein (CP) of species associated to Alphacryptovirus genus. Phylogenetic analysis of RdRP sequence showed highest amino acid identity with Black raspberry cryptic virus (BrCV, 63 %), Pepper cryptic virus 2 (PCV-2, 52 %) and -1 (PCV-1, 46 %). Whereas the CP putatively encoded by dsRNA-2 shared highest identity with Mulberry cryptic virus 1 (MCV-1, 49 %), PCV-2 (39 %) and PCV-1 (32 %). N-J phylogenetic analysis confirmed those relationships and delivered PeCV in a cluster with phyto-cryptoviruses belonging to genera Alpha- and Betacryptovirus, quite far distinguished from myco-cryptoviruses, gathered in genus Partitivirus.Virus-specific primers for RT-PCR were successfully designed inside the CP region to detect PeCV in several symptomless trees found in different orchards of Apulia (Southern Italy), thus proving that infection may be fairly common and presumably latent.Policlonal antiserum (kindly provided by Dr. M. Turina, CNR-IVV, Torino, Italy) specific to the CP of family-related Beet cryptic virus 2 (BCV-2) was profitably used for western blot detection of a 45-50 KDa band, coherent with predicted size of dsRNA-2 product. Furthermore, antibodies were useful for ISEM observation and subsequent decoration of PeCV particles, proved to be isometric, around 30 nm in diameter, with rounded shape lacking in fine structural details, not easily permeable by negative stain.

Autore Pugliese

• Minafra Angelantonio , De Stradis Angelo , Boscia Donato , La Notte Pierfederico

Tutti gli autori

• Morelli M.; De Stradis A.; La Notte P.; Mercuri J.; Boscia D.; Minafra A.

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Anno di pubblicazione

2012

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