First report of youcai mosaic virus in firecracker flower (Crossandra infundibuliformis) in Taiwan.

Abstract

Firecracker flower or crossandra (Crossandra infundibuliformis), an ornamental native to southern Asia, is commonly grown as bedding plants in the garden. In January 2021, crossandra plants showing mosaic, chlorotic ringspot, and leaf deformation (Suppl. Fig. 1) were observed at a recreational farm in Zhuolan Township (Miauli County, Taiwan) (E120°82'62'', N24°33'30'') . Transmission electron microscope (JEM-1400, JOEL, Japan) examination by negative staining of 10 affected plants indicated the presence of particles resembling a tobamovirus in all examined affected samples. However, no tobamovirus-like particles were observed in the crude sap prepared from healthy crossandra leaf tissues. Total RNA was extracted from affected leaves and used for RT-PCR amplification using the tobamovirus group-specific primer pair Tob Uni1 (5'-ATTTAAGTGGAGGGAAAACCACT-3') and Tob Uni2 (5'-GTYGTTGATGAGTTCGTGGA-3') (Letschert et al., 2002). A cDNA fragment of about 650-bp was amplified and Sanger sequenced (ABI PRISM 3730 DNA Sequencer, Biotechnology Center at National Chung Hsing University, Taichung, Taiwan ), revealing 98% sequence identity to that of a Brassica isolate of youcai mosaic virus (YoMV, AY318866). The virus was isolated through mechanical inoculation onto Chenopodium quinoa to yield two pure isolates, designated FC-1 and FC-2. Mechanical inoculation of FC-1 and FC-2 back to virus-free Crossandra infundibuliformis plants (5 for each isolate) resulted in systemic mosaic, chlorotic ringspot, and leaf deformation. All mock and healthy controls were symptomless and failed to obtain any RT-PCR products with YoMV-specific primers CPF1 (5'- ATGGTTTACAACATCACGAG-3') and CPR1 (5'-CTATGTAGCTGGCGCAGTAG-3'). Systemic symptoms of mild mosaic, ringspot, leafroll, and necrosis appeared on some of the tobaccos (Nicotiana benthamiana, N. tabacum, N. rustica), and cruciferous vegetables (Brassica rapa subsp. chinensis cv. Known-You No.2), B. rapa subsp. pekinensis cv. Autumn Sun), B. oleracea var. italica cv. Ching Hua), B. oleracea var. capitata cv. Green Peak), and Raphanus sativus cv. Snow Lady). However, inoculation of FC-1 and FC-2 on pepper (Capsicum annuum cv. Blue Star) resulted in severe necrosis on leaves and necrotic sunken spots on petioles and stems causing acute wilting and quick death. In stark contrast, FC-1 and FC-2 only induced local lesions on inoculated leaves of Chenopodium quinoa, Gomphrena globose, and Carica papaya. The infectivity of FC isolates to all plants used in host range tests were further confirmed by RT-PCR as mentioned. Oligonucleotide primers (Suppl. Table 1) specifically complementary to YoMV sequence were designed and used to amplify full-length genomic sequences of FC-1 and FC-2 isolates by RT-PCR and Sanger sequencing. Sequence analysis revealed that the genome of both FC-1 and FC-2 isolates consists of 6302 nucleotides. The viral genome has four open reading frames encoding a small replicase subunit, a RNA-dependent RNA polymerase , a movement protein, and a coat protein (CP), respectively. Both sequences, which shared 99.6% identity with each other, have been deposited in the NCBI database (Genbank Accession Numbers LC701592 and LC701593). FC-1 and FC-2 and the deduced amino acid sequences of CP shared 91.2% - 98.9% and 93.4 - 99.4% similarities, respectively, to those of published YoMV strains, confirming the identity of FC-1 and FC-2. RT-PCR analyses detected YoMV in all (100%) crossandra samples collected from the field (Suppl. Fig. 2). YoMV, formerly named Chinese rape mosaic virus (CRMV) or oilseed rape mosaic virus (ORMV) (Zhu et al., 2001), has been reported to infect cruciferous, solanaceous, and ornamental crops in Asia and Europe (Ju et al., 2019). The firecracker flower is a common and popular ornamental in Taiwan, even though its economic values are not so important. However, finding YoMV in firecracker flower may have epidemiological impacts as YoMV can infect economically important cruciferous and solanaceous crops. To our knowledge, this is the first report of YoMV infecting firecracker flowers in Taiwan.