Paris polyphylla var. yunnanensis is a perennial herb of the family Liliaceae. It is mainly distributed in southwest China, especially in Yunnan province [1]. Its rhizome, containing abundant active components including steroidal saponins and flavonoids, has anticancer and haemostatic pharmacological effects. Due to the important economic and medicinal value as well as scarity of the wild resources of P. polyphylla var. yunnanensis. P. polyphylla var. yunnanensis was domesticated and widely cultivated in Yunnan. Artifically planted P. polyphylla var. yunnanensis were frequently affected by fungal and viral diseases that lead to serious loss of yield and reduction in medicinal value. So far, there are five viruses reported from P. polyphylla var. yunnanensis including Paris mosaic necrosis virus (PMNV) [2], Paris polyphylla virus X (PpVX) [3], pepper mild mottle virus (PMMoV) [4], chilli veinal mottle virus (ChiVMV) [5], Paris virus 1 (ParV1) [6], which cause chlorosis, leaf deformation or yellowing of leaf veins symptoms on P. polyphylla var. yunnanensis.
In July 2021, P. polyphylla var. yunnanensis exhibiting yellow vein and mosaic symptoms were observed on paris plantation in Lijiang of Yunnan province, China (Fig. 1). Leaf samples were collected from the diseased plants and used for isolation of total RNA by TRIzol (Sangon, Shanghai, China). The presence of potyviruses were confirmed by RT-PCR using the degenerated primers for potyviruses [7, 8].
Symptomatic leaves were then high-throughtput sequenced by TsingKe (Kunming, China). 3121 contigs were obtained by transcriptom sequence. BLASTx analysis showed that a long contig consisting of 9563 nt shared 57.90% amino acid sequence identity with iris potyvirus A (IPA: accession QXU69567.1), suggesting it might be a distinct potyvirus, tentatively named as “ Paris yunnanensis mosaic chlorotic virus” (PyMCV).
The complete genomic RNA sequence of PyMCV was determined to be 9571 nt in length by RACE and RT-PCR, including the 5’-UTR (nt 1-171) and 3’-UTR (nt 9356-9571), excluding the poly (A) tail. It contains a large open reading frame (ORF) (nt 171-9356) encoding a polyprotein of 3,061 amino acids (aa) residues with a calculated molecular mass of 347 kDa. Nine highly conserved potyvirus proteolytic cleavage sites were found in the polyprotein, resulting in 10 putative mature proteins: P1 (299 aa), HC-Pro (457 aa), P3 (349 aa), 6K1 (53 aa), CI (635 aa), 6K2 (53 aa), VPg (195 aa), NIa (243 aa), NIb (513 aa) and CP (264 aa) (Supplementary Fig. S1). Each putative mature protein also has compared with other potyvirus proteins in NCBI (Supplementary Table S1). The highly conserved motif “ 2901GAAAAAAA2908 ” and a small ORF (PIPO) encoding 54 aa were found to be embedded in the P3 protein of PyMCV (Supplementary Fig. S2) [9]. Highly conserved potyviral motifs were identified in the PyMCV polyprotein by comparative analysis, including a 608PTK610 motif in HC-Pro; a 1243GAVGSGKST1251 , 1461VATNIIENGVTL1472 , 1505GERIQRLGRVGR1516 and 1332DECH1335 motifs in CI; a 2590GNNSGQPSTVVDNT2603 , 2631GDD2633 motifs in NIb.
To determine the phylogenetic status of PyMCV and volutionary relationship between PyMCV and other viruses of the genus Potyvirus, pairwise comparisons and construction of phylogenetic tree were performed using ClustalW Multiple alignment in MEGA-X [10], respectively. Pairwise comparison analysis showed that PyMCV polyprotein shared 54.20-59.60% nucleotide (Supplementary Table S2) and 51.80-57.90% amino acid sequence (Supplementary Table S3) identity with viruses of the genus Potyvirus. According to species demarcation criteria of the genus Potyvirus of ICTV, members of different species have complete ORF sequences that are generally <76% identical in nucleotide sequence and <82% identical in amino acid sequence [11]. Therefore, PyMCV belongs to be a new species of Potyvirus. The maximum likelihood phylogenetic tree based on the polyproteins indicated that lily yellow mottle virus (LYMV) is the closest known relative of PyMCV (Fig. 2).
In order to prevent Paris polyphylla var. yunnanensis from being infected by virus, further studies are needed to investigate the biological properties except genome organization of PyMCV.