Development of an on-site plum pox virus detection kit based on immunochromatography
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Sharka disease, caused by plum pox virus (PPV), is the most serious viral disease of stone fruit trees. Among the eight known strains of the virus, PPV-D is the most important due to its recent global spread. Although enzyme-linked immunosorbent assay (ELISA) is the most common approach for diagnosing sharka, it involves time-consuming steps and requires expensive equipment and trained technicians. In this study, an on-site PPV detection kit based on immunochromatography was developed using polyclonal antibodies against the coat protein (CP) of a PPV-D isolate. The immunochromatographic (IC) assay kit was as sensitive as a commercial ELISA system for detecting Japanese PPV-D isolates. Moreover, it was easy to use (a one-step procedure), and results could be obtained on-site within 15 min without special laboratory equipment. The IC assay kit detected the virus from every aerial part of symptomatic Japanese apricot trees. In a detailed study of viral localization in leaves, the most suitable plant parts for use in the IC assay were symptomatic mesophyll tissues and the region from the petiole to the main vein. A positive reaction was also observed using the CP of other major (PPV-M and PPV-Rec) and minor (PPV-EA, PPV-W, and PPV-T) strains.
KeywordsPlum pox virus (PPV) Diagnosis Antibody Immunochromatography Japanese apricot Stone fruits
We thank Dr. Ivanka Kamenova (AgroBioInstitute, Sofia, Bulgaria), Dr. Bojan Duduk (Institute of Pesticides and Environmental Protection, Belgrade, Serbia), and Dr. Mojca Virscek Marn (Agricultural Institute of Slovenia, Ljubljana, Slovenia) for providing European PPV-D, -M, and -Rec isolates. We also thank Mr. Hideo Hoshi (Tokyo Metropolitan Agriculture and Forestry Research Center) and the staff of The Ministry of Agriculture, Forestry and Fisheries of Japan (MAFF) for supporting sample collection in Western Tokyo. This study was supported through a grant from MAFF in Japan.
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