“Complete nuleotide sequences of Potato Rough dwarf virus and Potato virus P: molecular comparison and phylogentic relationship with other carlaviruses”

MASSA GABRIELA ALEJANDRA; SEGRETIN MARIA EUGENIA; PORTANTIER MARINA; BRAVO-ALMONACID FERNANDO FELIX; FEINGOLD SERGIO ENRIQUE

Coylumbridge, Aviemore, Scotland, United Kingdom

Congreso; 13th European Association for Potato Research (EAPR) Virology Section Meeting; 2007

Immunological evidences (Butzonich, et al, 1996) as well as molecular and biological comparison (Massa et al, 2006 and Nisbet et al, 2006) indicated a high degree of similarity between two South American Carlaviruses potato rough dwarf virus (PRDV, Argentina) and potato virus P (PVP, Brazil). Hence, they were cataloged (Massa et al, 2006 and Nisbet et al, 2006) as strains of the same species. The objective of this work is to compare complete genomic sequences of PRDV and PVP in order to reveal the molecular comparison between both viruses and its phylogenetic relationship with others carlavirus. Viruses were purified from virus-infected leaves of Nicotiana occidentalis using the protocol of the International Potato Center, with modifications. Viral genomic RNAs were extracted from the purified viruses particles by SDS/proteinase K and phenol:clhoroform extraction method. cDNAs synthesis from purified RNAs were produced using the Universal RiboClone® cDNA Synthesis System (Promega, USA). Double stranded cDNA molecules were cloned into EcoRV-digested, pZErOTM (Invitrogen, USA) and subsequently transformed into E.coli DH10B electrocompetent cells. The independent clones were sequenced using SP6 and T7 vector specific primers. For sequencing of the 5`-ends of both viruses, 5´-rapid amplification of cDNA ends (RACE) was performed. The genome of PRDV was 8,404 nucleotides long (excluding the poly (A) tail) and showed the typical organization for a Carlavirus, with six open reading frames (ORFs) coding for replicase (219 kDa), triple gene block (TGB) 1, 2 and 3 (25kDa, 11 kDa and 7 kDa, respectively), coat protein (CP, 35 kDa) and nucleic acid binding protein (NABP, 11kDa). PRDV showed to be closely related to passiflora latent virus, blueberry scorch latent virus, potato virus S, and lily symptomless virus, with whole genome nucleotide similarities ranging from 44% to 55%. PVP genome was approximately the same size as PRDV, presenting six ORFs, in a general arrangement coincident with a typical Carlavirus, too. At time of writing, (~60% of PVP sequence information) the nt sequence analyzed showed very high identity with PRDV. Full detailed results of PVP sequence and comparison with PRDV and others carlaviruses will be presented.