Biological, Biochemical, Serological, Molecular and Tissue Cultural Studies on an Egyptian Isolate of Tomato Spotted Wilt Virus Infecting Chrysanthemum Plants | ||
| Egyptian Journal of Phytopathology | ||
| Article 6, Volume 37, Issue 2, December 2009, Pages 79-94 PDF (4.87 M) | ||
| Document Type: Original Article | ||
| DOI: 10.21608/ejp.2009.235050 | ||
| Authors | ||
| Manal A. El-Shazly; Rehab A. Dawood; Ahmed M. Soliman* | ||
| Virus and Phytoplasma Dept., Plant Pathology Research Institute, Agricultural Research Center, 12619, Giza, Egypt | ||
| Abstract | ||
| Tomato spotted wilt virus (TSWV) has been isolated and identified from naturally infected chrysanthemum plants, collected from the experimental farm of the Fac. of Agric., Cairo Univ. by indirect enzyme linked immunosorbent assay (Indirect-ELISA) and dot blotting immunobinding assay (DBIA) using an induced antiserum for TSWV. The symptoms consisted of dark colored leaf necrosis, necrotic line patterns, necrotic local lesion and tip necrosis on leaves, stunting, wilting and flower distortion. All the tested chrysanthemum cultivars were found to be susceptible when mechanically inoculated under greenhouse conditions. Wide variations of symptoms were found between Shakira, Feling, Grancl, Zambia and Maxx cultivars. Cv. Zambia was found to be more susceptible than any other cultivar tested in the present study, thus it was used for the production of virus-free plants using tissue culture technique. TSWV was purified from infected Gompherina globosa plants. The UV absorption spectrum had Amax at 260nm, Amin at 245nm and A260/280 ratio of 1.2. Coat protein subunit of TSWV had a value of 29 KDa. The IgG fraction of the prepared antiserum had Amax and Amin at 280 and 242 nm, respectively. Immunocapture-reverse transcription-polymerase chain reaction (IC- RT-PCR) was used to amplify 760 bp cDNA fragments from infected chrysanthemum leaves using the primers (SLS90-47) and (JLS90-46) specific to TSWV. The dilution end point of TSWV in infected tissue extracts was 1/1280, 1/2560 and l0-5for indirect ELISA, direct ELISA and IC- RT-PCR, respectively. IC- RTPCR proved to be more sensitive than indirect and direct ELISA. In this study, meristem-tips were excised from infected chrysanthemum plants. Thermotherapy and/or chemotherapy were used to eliminate TSWV. Results demonstrated that application of thermotherapy at 38°C for 28 days followed by 10 to 20 mg/l Virazole in vitro gave a survival rate of 53 and 77%, respectively. The presence of virus in the produced plants was evaluated by ELISA technique. In vitro thermotherapy combined with chemotherapy eliminated TSWV from infected chrysanthemum and reduced the risk of introducing this pathogen through vegetative propagation. | ||
| Keywords | ||
| Tomato; Solanum lycopersicum; Tomato spotted wilt virus; TSWV; Chemotherapy; Chrysanthemum; DBIA, indirect-ELISA; Meristem tip; Immunocapture RT-PCR; Thermotherapy; Tissue culture | ||
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