First Report of Cucumber mosaic virus in Catharanthus roseus in Korea.

ABSTRACT

Catharanthus roseus, commonly known as Madagascar rosy periwinkle (also called vinca), is a tropical perennial herb of the family Apocyanaceae. Periwinkle is a bedding plant widely used in Korea because of its drought tolerance, low maintenance, and varied flower colors. In May 2013, virus-like foliar symptoms, including a mosaic with malformation of leaves, were observed on a periwinkle plant in a greenhouse located in Chonbuk Province, Korea. Cucumber mosaic virus (CMV) was identified in the symptomatic plant by serological testing for the presence of CMV coat protein (CP) with an immune-strip kit developed by our laboratory. The presence of CMV was confirmed by serological detection with a commercially available double-antibody sandwich (DAS)-ELISA kit (Agdia, Elkhart, IN). Sap from the serologically positive sample was mechanically inoculated to test plants using 10 mM phosphate buffer (pH 7.0). The virus (named CMV-Vin) caused necrotic local lesions on Chenopodium amaranticolor at 5 days-post-inoculation (dpi), while mild to severe mosaic was observed in Capsicum annuum, Cucumis sativus, Cucurbita pepo 'Cheonggobong,' Nicotiana glutinosa, N. tabacum'Samsun NN,' Physalis angulate, and Solanum lycopersicum 'Pink-Top' 10 to 14 dpi. Examination of the inoculated plant leaves by DAS-ELISA and electron microscopy (leaf dips) showed positive reactions to CMV and the presence of spherical virions ~28 nm in diameter, respectively. To verify whether CMV was the causal agent for the disease symptoms observed in naturally infected periwinkle, virus-free periwinkle (10 plants) was mechanically inoculated by sap from local lesions on C. amaranticolor inoculated with CMV-Vin. At 6 weeks after inoculation, all plants produced systemic mosaic and distortion of leaves, resulting in strong DAS-ELISA reactions for CMV, whereas mock-inoculated periwinkle plants remained symptomless and virus-free. The presence of CMV-Vin in all naturally infected and mechanically inoculated plants was further verified by reverse transcription (RT)-PCR. Total RNAs were extracted with a RNeasy Plant Mini Kit (Qiagen, Valencia, CA) and RT-PCR was carried out with the One-Step RT-PCR Kit (Invitrogen, Carlsbad, CA) using a pair of primers, CMVCPFor and CMVCPRev (1), which amplified the entire CP gene. RT-PCR products (657 bp) were obtained from all naturally infected and mechanically inoculated plants as well as from a positive control (viral RNAs from virions), but not from healthy tissues. The amplified RT-PCR products were directly sequenced using BigDye Termination kit (Applied Biosystems, Foster City, CA). Multiple alignment of the CMV-Vin CP sequence (Accession No. AB910598) with CP sequences of other CMV isolates using MEGA5 software revealed that 91.8 to 99.0% and 71.0 to 73.0% identities to those of CMV subgroup I and subgroup II, respectively. These results provide additional confirmation of CMV-Vin infection. Being perennial, periwinkle plants could serve as a reservoir for CMV to infect other ornamentals and cultivated crops (2). To our knowledge, this is the first report of CMV infection on periwinkle in Korea. References (1) S. K. Choi et al. Virus Res. 158271, 2011. (2) P. Palukaitis et al. Adv. Virus. Res. 41281, 1992.