Journal of Taiwan Agricultural Research

Author:Fen-Lang Chiang, An-Chin Chang, Ying-Huey Cheng, and Chin-Chih Chen*

Abstract:

    African lily (Agapanthus africanus L.), native in South-Africa, is currently a popular bulbous floral crop in Taiwan and many other countries. An African lily exhibiting foliar chlorotic strip and mottle symptom was collected from Mai-Fung nursery of Taiwan University in 2005. A virus isolate, designated as AL-1, was obtained from this plant by establishing a single lesion isolate in Chenopodium quinoa Willd. Back inoculation of AL-1 onto seedlings of African lilies confirmed that AL-1 was able to induce chlorotic mottle symptom which was similar to what was observed in the field. Virions of AL-1 were successfully purified from the inoculated tissue of C. quinoa by differential and equilibrium centrifugation in cesium sulfate. Elongated virus particles estimated at 508 nm in length were observed from the purified sample under an electron microscope. By SDS-PAGE, molecular weight of AL-1's coat protein was determined as 25 kDa. An antiserum against purified AL-1 virions was prepared in rabbit and shown to react strongly and specifically with its homologous antigens prepared from diseased samples of field-collected or inoculated ones in enzyme-linked immunosorbent assay (ELISA), SDS-immunodiffusion, and western blotting tests. Full-length genome of AL-1 was revealed by RT-PCR amplification with random primer and several primer pairs designed in this study according to those closely related virus sequences in the GenBank. Genome of AL-1 was found to compose 6577 nucleotides, excluding the poly (A) tails, in which five open reading frames (ORF) encoding presumably replicase (ORF 1), triple gene blocks (ORF 2–4) and coat protein (ORF 5) were located. This genome structure was equivalent to that of Potexvirus and most resembled to Nerine virus X (NVX) (Acc. No. AB219105) in the GenBank. The percent identities of nucleotide and amino acid sequences of the full-length genomes between AL-1 and NVX were 98 and 93%, respectively, indicating they were isolates of the same virus. A primer pair for specific detection of AL-1 was designed and shown to be useful in routine RT-PCR test. Our results revealed that AL-1 was an isolate of NVX based on the biological and serological characterizations and the molecular analysis of the full-length genomic sequences. To our knowledgement, it's the first report of NVX occurrence in Taiwan.

Key words:African lily, Agapanthus africanus, Virus disease, Nerine virus X, Polyclonal antiserum, Full-length genome, RT-PCR

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