FIRST REPORT OF GRAPEVINE LEAFROLL-ASSOCIATED VIRUS 9 IN SPAIN

C.V. Padilla, E. Cretazzo, M.J. Alcalá, I. Hita, N. López, V. Padilla, L. Velasco

Abstract


Grapevine leafroll is one of the most important viral diseases of grapevines (Vitis vinifera). To date, 11 grapevine leafroll-associated viruses (GLRaVs) from the family Closteroviridae have been described. A recent taxonomic revision, however, suggests the existence of only five valid species (GLRaV-1, -2, -3, -4, and -7), whereas GLRaV-4, -5, -6, -9, -Pr, -De and Grapevine leafroll-associated Carnelian virus (GLRaCV) form a uniform phylogenetic clade and represent strains of GLRaV-4 (Martelli et al., 2012). In Spain, IMIDA performs all sanitary tests for certification of candidate clones from all Spanish autonomous communities. The detection of GLRaV-4, -5, -6, -7 and -9 relies on the use of commercial ELISA kits (Bioreba, Switzerland), and the confirmation of positive results by real-time RT-PCR to assess the health status of the clones. In 2011, GLRaV-9 was detected using DAS-ELISA and TaqMan in a candidate clone of Mantúa (Man086), a traditional cultivar from Extremadura. To confirm the presence of this virus, conventional RT-PCR and sequencing were performed using total RNA from Man086 as template and specific primers designed in the HSP70h domain of GLRaV-9 (Alkowni et al., 2004). Pairwise comparisons of the corresponding sequence (351 bp, GenBank accession No. KC660077) with GLRaV-9 sequences available in GenBank showed the highest nucleotide (97.2%) and amino acid (95.7%) identities with an isolate from Washington state, USA (EU252530). Another set of primers was designed in the RNA-dependent RNA polymerase domain based on accession AY297819 from California, USA. Alignments between the sequence obtained (533 bp, GenBank accession No. KC660076) and AY297819 showed 96.1% and 97.2% nucleotide and amino acid sequence identities, respectively. This is the first report of GLRaV-9 in Spain.

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DOI: http://dx.doi.org/10.4454/JPP.V95I3.025

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