Taiwan cherry or Formosan cherry (Prunus campanulata Maxim.) is a beautiful ornamental tree that is native to Taiwan. In spring 2005, a severe disease was observed on 1- to 3-year-old seedlings of Taiwan cherry in a garden in Tungshih, Taichung, Taiwan. Infected plants showed symptoms of greenish water-soaked spots on leaves that became dark brown, 2 to 3 cm in diameter. Infected leaves withered and fell to the ground in 3 to 5 days and young shoots showed symptoms of withering and drooping. Infected roots showed symptoms of necrosis. Severely infected plants eventually died. A Phytophthora sp. was isolated consistently from diseased samples of Taiwan cherry and associated soil. Six isolates of Phytophthora, of the A1 mating type (1), were isolated from single zoospores. Two of these isolates, Tari 25141 (deposited as BCRC34932 in Bioresource Collection and Research Center, Shinchu, Taiwan) and Tari 25144 (BCRC34933), were used for pathogenicity tests on 1-year-old seedlings of Taiwan cherry to fulfill Koch's postulates. Inoculation was done by placing a cotton swab containing zoospore suspension on leaves or stem, or by soaking seedlings in the zoospore suspension. Inoculated seedlings were kept in a greenhouse at 20 to 25°C for 30 days and examined for appearance of symptoms. Results showed that both isolates were pathogenic on seedlings of Taiwan cherry, causing symptoms similar to those observed on naturally infected seedlings. The temperature range for growth of the six isolates of Phytophthora was 8 to 32°C with optimum temperature at 24°C. The linear growth rate was 72 mm per day on V8A culture (5% V8 vegetable juice, 0.02% CaCO3, and 2% Bacto agar) at 24°C. The colonies on potato dextrose agar produced sparse aerial mycelia with conspicuous radiate patterns. Sporangia were sparse on V8A agar blocks, but abundant when the agar blocks were placed in water under continuous white fluorescent light (average 2,000 lux) for 2 days. Sporangiophores branched sympodially. Sporangia were pear shaped, nonpapillate and nondeciduous, 50 to 75 (62) × 30 to 48 (40) μm, with a length/width ratio of 1.2 to 2.2 (1.6). New internal nested proliferate sporangia were formed inside the empty sac of old matured sporangia after releasing zoospores. No chlamydospores were formed on V8A. Hyphal swellings with distinctive irregular catenulation were produced on V8A and in water. The pathogen was stimulated to form its own oospores by the A2 tester using the method described by Ko (1). Oogonia were 28 to 50 (40) μm in diameter with smooth or irregularly protuberant walls. Oospores were mostly aplerotic and 18 to 42 (31) μm in diameter. Antheridia were amphigynous, mostly two-celled, and 10 to 42 (29) × 12 to 24 (19) μm. The sequence of the internal transcribed spacers (ITS) region of nuclear ribosomal DNA of isolate Tari 25141 (GenBank Accession No. GU111589) was 831 bp and had 99% sequence identity with a number of Phytophthora cambivora isolates such as GenBank Accession Nos. HM004220 (2), AY787030, and EF486692. Based on the morphological characteristics of sporangia and sexual structures and the molecular analysis of ITS sequences, the pathogen from Taiwan cherry was identified as P. cambivora (Petri) Buis. To our knowledge, this is the first report of P. cambivora on native Taiwan cherry in Taiwan and, so far, no other natural hosts have been reported. References: (1) W. H. Ko. J. Gen. Microbiol. 116:459, 1980. (2) P. W. Reeser et al. Mycologia 103:225, 2011.
Phylogenetics of Taxus Using the Internal Transcribed Spacers of Nuclear Ribosomal DNA and Plastid trnL-F Regions
