First Report of Grapevine Virus T Infecting Grapevine in China

Abstract

Grapevine virus T (GVT), a putative new member of the genus Foveavirus in the family Betaflexiviridae, was first described from transcriptomic data of the cultivar Teroldego (Jo et al. 2017). Since then, GVT has been reported from grapevines in Germany (Ruiz-Garcia et al. 2018), Croatia (Voncina et al. 2018), and other European countries (Glasa et al. 2018; Nourinejhad Zarghani et al. 2018). It is closely related to Grapevine rupestris stem pitting-associated virus and exhibited relatively high intraspecies variability. In June 2017, a field survey of grapevine disease in Yanqing district of Beijing was conducted, and grapevine plants with the symptoms of chlorosis and leaf rolling were observed with an incidence of 5 to 40%. Total RNA was extracted from leaf samples of a symptomatic Red Globe grapevine plant and used for constructing a transcriptome library followed by high-throughput sequencing on an Illumina HiSeq 4000 platform. De novo assembly of the 8,290,478 reads was performed by Trinity, and the BLASTn and BLASTX analysis of the assembled 14,834 contigs revealed the presence of one contig related to GVT (8,653 nt in length), with a nucleotide similarity ranging from 81 to 92%, together with the contigs related to Grapevine leafroll-associated virus 1, Grapevine Red Globe virus, and Grapevine Pinot gris virus. Furthermore, the complete genome of this GVT isolate (GVT-BJ) was amplified with conventional reverse transcription PCR (RT-PCR) using the primer pairs F1:5′-TGCTTTCACGTAATGGCTCTTCAC-3′/R1:5′- CCTCAGCAACTCAAAAATGTCAGAAT-3′ and F2:5′-GTGGTGGTCCTTGATGAAATTCAGCT-3′/R2:5′-AGTACGGTATGCCAGCGAACATATGC-3′ designed for the assembled contig and RACE (5′-GATTACGCCAAGCTTCGGGGGTATGAGATTTGCCAATGTTGGC-3′ for 5′ RACE and 5′-GATTACGCCAAGCTTGATGAAAGTGCTTTGGAGCCGGAAGG-3′ for 3′ RACE), and the PCR products were cloned and Sanger sequenced. The genome length of GVT-BJ was 8,700 nt (GenBank no. MH158659) with the typical foveavirus genomic organization with five open reading frames. A pairwise sequence comparison revealed GVT-BJ shared highest nucleotide sequence identity of 92.3% with the French isolate CF-3 (MH674180). Furthermore, 36 symptomatic grapevine samples from nine vineyards in Yanqing district were screened for the presence of GVT by RT-PCR with the primer pair targeting the CP region (CPf 5′-ATGGCGTCAAATGAGGAGC-3′/CPr 5′-TTATGAATCTCCCCCAAATGGATTG-3′); four were positive, and the presence of GVT was further confirmed by sequencing the PCR products. These four grapevine samples were also positive for Grapevine leafroll-associated virus 1 identified by RT-PCR with the reported primers pORF9F and pORF9R (Little and Rezaian 2006). This is the first report of GVT in China in mixed infection with other viruses. A larger-scale survey of commercial vineyards for the prevalence of GVT in China and its association with the observed symptoms is needed.