African violet (Saintpaulia ionantha) is an ornamental species of the family Gesneriaceae and is characterized by fleshy leaves and colorful flowers. This popular, exotic ornamental, originally from Kenya and Tanzania, is vegetatively produced from cutting and tissue culture (1). In May 2013, virus-like foliar symptoms, including a mosaic with dark green islands and chlorosis surrounding the veins, were observed on an African violet plant in a greenhouse located in Icheon, Korea. Cucumber mosaic virus (CMV) was identified in the symptomatic plant by serological testing for the presence of CMV coat protein (CP) with a commercial immunostrip kit (Agdia, Elkhart, IN). The presence of CMV was confirmed by serological detection with a commercially available double-antibody sandwich (DAS)-ELISA kit (Agdia). Sap from the serologically positive sample was mechanically inoculated to test plants using 10 mM phosphate buffer (pH 7.0). The virus (named CMV-AV1) caused necrotic local lesions on Chenopodium amaranticolor at 5 days post-inoculation (dpi), while mild to severe mosaic was observed in Nicotiana glutinosa, N. tabacum ‘Samsun NN,’ Cucurbita pepo ‘Super-Top,’ Physalis angulate, and Solanum lycopersicum ‘Unicorn’ 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-AV1 is the cause of disease symptoms observed in African violet, virus-free African violet (10 plants) was mechanically inoculated by sap from local lesions on C. amaranticolor inoculated with CMV-AV1. At 8 weeks after inoculation, all plants produced systemic mosaic and chlorosis surrounding veins, resulting in strong DAS-ELISA reactions for CMV, whereas mock-inoculated African violet plants remained symptomless and virus-free. The presence of CMV-AV1 in all naturally infected and mechanically inoculated plants was further verified by reverse transcription (RT)-PCR. Total RNAs were extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany), according to the manufacturer's instructions. RT-PCR was carried out with the One-Step RT-PCR Kit (Invitrogen, Carlsbad, CA) using a pair of primers, CPTALL3 and CPTALL5 (2), amplifying the entire CP gene and part of an intergenic region and 3′-noncoding region of CMV RNA3. RT-PCR products (960 bp) were obtained from all naturally infected and mechanically inoculated plants as well as from positive control (viral RNAs from virions), but not from healthy tissues. The amplified RT-PCR products were purified with QIAquick PCR Purification Kit (Qiagen) and sequenced using BigDye Termination kit (Applied Biosystems, Foster City, CA). Multiple alignment of the CMV-AV1 CP sequence (Accession No. AB842275) 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-AV1 infection. CMV may pose a major threat for production of African violet since the farming of African violet plants is performed using the vegetative propagation of the African violet leaves in Korea. In particular, mosaic and chlorosis symptoms in African violet cause damage to ornamental quality of African violet. To our knowledge, this is the first report of CMV infection of African violet in the world. References: (1) S. T. Baatvik. Fragm. Flor. Geobot. Suppl. 2:97, 1993. (2) S. K. Choi et al. J. Virol. Methods 83:67, 1999.
Some Variables Affecting the Use of Cowpea as an Assay Host for Cucumber Mosaic Virus