First Report of Shallot yellow stripe virus in Lilium lancifolium in China

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Lilium lancifolium (family Liliaceae), also called tiger lily, is widely cultivated in China for its edible bulbs and medicinal application (China Pharmacopoeia Committee 2005; Yu et al. 2015), with a production value of approximately six billion Yuan per year. L. lancifolium is a polyploid complex including both diploid (2n = 2x = 24) and triploid (2n = 3x = 36) cytotypes, and only the triploids are widely distributed in inland China (Kim et al. 2006; Noda 1986). Because the triploids are sterile, therefore L. lancifolium cannot extend its population through seeds but propagates only through an asexual mode. However, vegetative propagation is vulnerable to the accumulation of viruses over subsequent generations. Shallot yellow stripe virus (SYSV), belonging to the genus Potyvirus of the family Potyviridae, was isolated from shallot samples originating from Southeast Asia (Indonesia, China, and Thailand) (Van der Vlugt et al. 1999). A virulent strain of this virus caused striping in sap-inoculated garlic (Allium sativum) and Formosan lily (Lilium formosanum) (Van Dijk 1993). In June 2015, longitudinal yellow stripe symptoms on leaves were observed on approximately 10% of L. lancifolium plants in the greenhouse of the Chinese Academy of Agricultural Sciences (Beijing, China). Total RNA was extracted from 15 symptomatic leaves (lesion covering one-third of leaf area) randomly collected from 10 different plants, and a de novo transcriptome was constructed using the Illumina HiSeq 4000 platform (Novogene Bioinformatics Technology Co., Beijing, China). A total of 54,142,872 paired-end reads of 150 bp were obtained. De novo assembly of the reads generated one large contig (9,843 bp) sharing a maximum nucleotide identity of 85.0% and amino acid identity of 90.6% with SYSV (GenBank accession no. AJ865076) isolated from Welsh onion in China (Chen et al. 2005). To confirm the Illumina sequencing results, the full-length SYSV genome was amplified by reverse transcription PCR (RT-PCR) using seven pairs of specific primers designed from the contig obtained in this study and sequenced. Seven RT-PCR fragments were assembled to obtain an SYSV sequence with a full length of 9,843 bp, which contained a single 9,441-bp complete open reading frame. The sequence shared a nucleotide sequence identity of 98% with the sequence assembled from Illumina RNA-Seq data. The isolate of SYSV from L. lancifolium was named LL01 and was deposited in GenBank under accession MK127536. A pair of specific primers (5′-ATACCCTTTGGTTTCCGC-3′ and 5′-TGCTATCATCACCCTCGC-3′), which amplified a 269-bp fragment, was designed according to the genome sequence of the isolate LL01 to detect SYSV in Lilium species. In total, 30 out of 50 analyzed symptomatic leaf samples (60%) tested positive, whereas negative controls remained negative. SYSV was not detected in 40% of symptomatic samples, suggesting that divergent variants of the virus, or perhaps other viruses, could be associated with the observed symptoms, and further research is needed. To our knowledge, this is the first report on the occurrence and sequence of SYSV in L. lancifolium in China. This report suggests that L. lancifolium plays a possible role as a reservoir host of the virus for Allium species. In addition, growers should plant virus-free bulbs obtained through shoot tip culture combined with heat treatment or medicament treatment.