First Report on the Occurrence of Grapevine leafroll-associated virus 7 and 9 in Chilean Grapevines.

Grapevine is one of the oldest horticultural crops and represents a highly valuable agricultural commodity. So far, nine distinct Grapevine leafroll-associated viruses (GLRaVs) within the Closteroviridae family have been found to be associated with grapevine leafroll disease (3). Previous studies have demonstrated a high incidence of GLRaV-1, -2, and -3 in Chile (2). To determine if other GLRaVs were present, 21 dormant cane samples were screened with a comprehensive 70-mer oligonucleotide microarray designed to simultaneously detect all grapevine viruses with total or partial genomic sequence available. The array contained 570 unique probes designed against specific regions of more than 40 viral genomes (E. Engel et al., 15th ICVG [Abstr.], 2006). One sample (cv. Black Seedless) showing a microarray hybridization pattern compatible with a mixed infection of GLRaV-7 and GLRaV-1 was analyzed by ELISA using GLRaV-7 specific antibodies (Agritest, Valenzano, Italy) and reverse transcription (RT)-PCR using virus-specific primers LR7-F: 5'- TAT ATC CCA ACG GAG ATG GC -3' and LR7-R: 5'- ATG TTC CTC CAC CAA AAT CG -3' (based on GenBank Accession No. Y15987). The serological analysis confirmed the presence of GLRaV-7 with further confirmation by the RT-PCR product of 502 bp corresponding to a fragment of the HSP70h gene that was cloned and sequenced. The Chilean GLRaV-7 sequence (GenBank Accession No. EU334662) showed 94% nucleotide and 95% amino acid identity when compared with a corresponding region of another GLRaV-7 isolate from Albania (GenBank Accession No. Y15987). GLRaV-1 infection was confirmed by ELISA (Bioreba AG, Reinach, Switzerland) and RT-PCR. A second sample (cv. Tintorera) showing microarray hybridization pattern compatible with a mixed infection of GLRaV-9 and Grapevine virus A (GVA) was analyzed by RT-PCR using virus-specific primers LR9-F: 5'- CGG CAT AAG AAA AGA TGG CAC -3' and LR9-R: 5'- TCA TTC ACC ACT GCT TGA AC -3' (1). The RT-PCR product of 393 bp corresponding to a fragment of the HSP70h gene was cloned and sequenced (GenBank Accession No. EU334663), showing 94% nucleotide and 95% amino acid identity when compared with a corresponding region of another GLRaV-9 isolate from the United States (GenBank Accession No. AY297819). Since there are no commercial antibodies available for GLRaV-9 detection, a second pair of primers, LR9-F1: 5'- AAA GGT TTC TGC TGG TTA CC -3' and LR9-R1: 5'- CTT TCA GAA CAG TCC TCC TC -3' that amplified a fragment of ORF1a was also used. The 301-bp product was cloned and sequenced (GenBank Accession No. EU588989) showing 93.7% nucleotide and 98% amino acid identity when compared with a corresponding region of another GLRaV-9 isolate (GenBank Accession No. AY297819). GVA infection was confirmed by ELISA (Bioreba AG) and RT-PCR. To our knowledge, this is the first report of GLRaV-7 and GLRaV-9 in Chile. Further studies will help determine the effect and incidence of these viruses in Chilean grapevines. References: (1) R. Alkowni et al. J. Plant Pathol. 86:123, 2004. (2) N. Fiore et al. J. Plant Pathol. 90:125, 2008. (3) G. P. Martelli and E. Boudon-Padieu. Options Méditerr. B55, 2006.

First Report of Rupestris stem pitting associated virus in Argentina.

Rupestris stem pitting associated virus (RSPaV), a component of the rugose wood complex (RWC), is a worldwide graft transmissible disease of grapevines (Vitis vinifera L.). RSPaV has a single-stranded 8,726-nt RNA genome, belongs to the genus Foveavirus, and is often associated with Rupestris stem pitting (RSP) disease (2). In 1995, a grapevine sanitary selection program was implemented in Mendoza to investigate this and other grapevine viral diseases. RSP can be diagnosed when V. rupestris cv. St. George is used as a woody indicator for biological indexing. Chip-bud inoculated St. George plants developed a row of small pits and grooves on the wood cylinder below the graft or around and below the inoculated point (1,2). After three seasons in the field, 15 accessions with RSP wood markings were observed. Mature leaves and bark shavings were extracted, partially purified, and analyzed by a onestep reverse transcription polymerase chain reaction method. The expected 339-bp band was found in only six of the positively indexed samples using the specific 13/14 primer pair (2). Other viruses associated with RWC have been detected in Argentina, but to our knowledge, this is the first report of RSPaV. References: (1) A. C. Goheen. Page 53 in: Compendium of Grape Diseases, R. C. Pearson and A. C. Goheen, eds. American Phytopatological Society, St. Paul, MN, 1988. (2) B. Meng. Rupestris stem pitting: Insights on etiology and development of reverse transcription-polymerase chain reaction and immunoassays for diagnosis. Ph.D. Diss. Cornell University, Ithaca, NY, 1999.