3.7.15
PEACH LATENT MOSAIC VIROID ISOLATED FROM STONE FRUITS IN JAPAN

H OSAKI 1, Y SATO2, Y TOMITA3, Y KAWAI4, Y MIYAMOTO5 and Y OHTSU1

1National Institute of Fruit Tree Science, Fujimoto, Tsukuba 305-8605, Japan; 2Akita Fruit Tree Experiment Station, Hiraka-cho, Hiraka-gun, Akita 013-01, Japan; 3Ibaraki Horticultural Research Institute, Minori-machi, Ibaraki 319-02, Japan; 4Nagano Fruit Tree Experiment Station, Suzaka, Nagano 382, Japan; 5

Background and objectives
Three graft-transmitted diseases of peach, yellow mosaic, oil blotch and star mosaic, are known in Japan. A small spherical viral particle was detected from yellow mosaic-infected peach trees. Namba et al. (1979) confirmed that the virus was the causal agent of peach yellow mosaic disease by back-inoculation with the purified preparations. However, agents of oil blotch and star mosaic diseases are unknown. On the other hand, peach latent mosaic (PLM) disease was initially identified in France. Peach latent mosaic viroid (PLMVd) is agent of PLM disease. Also peach mosaic disease is known in the USA. But PLMVd is not related to the agent of peach mosaic disease. We aimed to study the occurrence of PLMVd in stone fruit of Japan. Later we would like to study the relationship between PLMVd and the agents of three graft-transmitted diseases of peach, yellow mosaic, oil blotch and star mosaic.

Materials and methods
Stone fruit samples were collected at random, because PLM disease is generally latent, at several local orchards in Japan. Stone fruit trees maintained in the National Institute of fruit tree science were also used. Nucleic acid preparations were extracted from half gram of tissue by SDS-phenol. After ethanol precipitation, the pellet was suspended in 40l of distilled water. The preparation and dilution series of it were used for RT-PCR. A primer pair was designed according to the nucleotide sequence of PLMVd reported by Hernandez and Flores (1992). Products were cloned by TA Cloning Kit (Invitrogen). Each nucleotide was sequenced at least once on each strand. RT-PCR products were analyzed by electrophoresis in 5% polyacrylamide gel. Gels were stained with ethidium bromide.

Results and conclusions
Peach latent mosaic viroid (PLMVd) was detected from stone fruits in Japan, using RT-PCR method. Two PLMVd isolates from peach cultivars Nakatsu hakutou and Akatsuki were sequenced. Two isolates consisted of 337 nucleotides. Nakatsu hakutou isolate differed from the French isolate at 24 sites in the sequence, including 24 base exchanges, whereas Akatsuki isolate differed from at 28 sites, including 26 base exchanges, one insertion and one deletion. PLMVd was detected from 33 of 35 (94.3%) peach samples, from one of 19 (5.3%) Japanese plum samples, from 2 of 45 (4.4%) mume samples, from one of 27 (3.7%) sweet cherry samples and not detected from European plum, apricot samples in local orchards in Japan. PLMVd was also detected from many peach cultivars maintained in the National Institute of Fruit Tree Science.

References
1. Kishi K, Takanashi K, Abiko K, 1973. Bulletin of the Horticultural Research Station, Japan, Series A 12, 197-208 (in Japanese with English summary).
2. Hernandez C, Flores R, 1992. Proc. Natl. Acad. Sci. USA 89: 3711-3715.