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First Report of Tomato ringspot virus in an Ohio Vineyard

Yao, X.-L., Han, J., Domier, L. L., Qu, F., Lewis Ivey, M. L.
Plant disease 2018 v.102 no.1 pp. 259
Tomato ringspot virus, Vitis vinifera, biotechnology, cultivars, decline, genome, high-throughput nucleotide sequencing, hybrids, lakes, leaves, nucleotides, raspberries, reverse transcriptase polymerase chain reaction, reverse transcription, ribosomal RNA, shoots, tissues, vines, vineyards, viruses, viticulture, wine grapes, Illinois, Ohio
Tomato ringspot virus (ToRSV) is known to be associated with raspberry and grapevine decline in Canada, Chile, and several U.S. states, but had not previously been found in Ohio grapevines (Rivera et al. 2016; Rott et al. 1991, 1995; Walker et al. 2015). This report documents the first discovery of ToRSV in an Ohio vineyard. ToRSV is a member of the genus Nepovirus, subfamily Comovirinae, family Secoviridae, with two single-stranded, positive sense RNA genome segments (RNA1 and RNA2) of approximately 8,200 and 7,200 nucleotides (nt), respectively (Rott et al. 1991, 1995). The ToRSV Ohio Grapevine (OG1) isolate was discovered through high throughput sequencing (RNA-Seq) of pooled RNA extracted from 140 grapevine leaf samples, collected from 40 Ohio vineyards during 2012 to 2014. Specifically, the 140 leaf samples were each subjected to total RNA extraction. One microgram aliquots were then withdrawn from the RNA samples, and those from the same of the seven Ohio grape-growing regions (northeast, lake shore, northwest, canal country, central, southwest, and Appalachian) were pooled to form seven regional pools. A final master pool was then made by combining equal aliquots of the regional pool RNAs (2 μg per regional pool), and sent for rRNA removal and RNA-Seq at the Roy J. Carver Biotechnology Center, University of Illinois at Urbana-Champaign, with a HiSeq2000 platform. Subsequent analysis of the sequencing reads revealed more than 20 different grapevine viruses, among them ToRSV. The ToRSV reads were assembled into two large contigs. The first contig was 5,987 nt long, sharing 94% sequence identities with RNA1 of a Canadian (Rasp1-2014) and a Chilean (Rasp-CL) ToRSV isolate (Rivera et al. 2016; Walker et al. 2015). A second, 6,074-nt contig shared 93% sequence identity with the RNA2 of the Rasp-CL isolate (Rivera et al. 2016). Both contigs also share varying levels of sequence identities (83 to 98%) with partial sequences of many other ToRSV isolates deposited in GenBank. The sequences of these two contigs have been deposited in GenBank with the accession numbers MF176958 and MF176959. To map the originating vineyard(s) of the ToRSV OG1 isolate, the 140 samples were tested by reverse transcription (RT)-PCR, with the following two primers we designed: ToRSVR1-5892F (5′-GATTTGCAAGCTATCTATTCTTCCCTGTA) and ToRSVR1-6426R (5′-GAATCGCGCATATGGCGTACGCGATGA). These experiments allowed us to trace the ToRSV OG1 isolate to two samples collected in August of 2012, from two neighboring grapevines (cultivar Vidal Blanc, a hybrid derived from Vitis vinifera and another hybrid) in the same Ohio vineyard located in the southwest region (50 km northeast of Cincinnati), showing symptoms of severe decline including stunted shoots, small fruit clusters with few, small fruits, and premature leaf senescence. Transmission of ToRSV was not carried out because additional virus-containing tissues could not be collected, due to removal of the diseased vines. The prevalence of ToRSV in Ohio vineyards and its impact on Ohio wine grape production will be assessed with additional investigations.