Peucedanum praeruptorum Dunn, a traditional Chinese medicinal herb, is an important crop in Ningguo, China. Since 2010, leaf spot symptoms were observed yearly starting in June. Blighted leaf areas on individual plants ranged from 10 to 25% in many fields, and up to 200 ha were affected each year. Symptoms consisted of small, brown, necrotic spots uniformly distributed on the 1- to 2-week-old leaves. Small tissue pieces from the edges of lesions were disinfected in 2% NaClO for 3 min, rinsed twice in distilled water, plated on potato dextrose agar (PDA), and incubated at 25°C in darkness for 4 days. Single spore isolations were obtained for six strains. When inoculated on SNA media, the six strains produced typical septate mycelium, with the young hyphae hyaline and aged ones white greyish. Setae of the strains on SNA were brown, tip acute, 2- to 3-septate, and 32.5 to 85.6 μm long. Conidiogenous cells were hyaline, cylindrical, 2- to 3-septate, 6.2 to 16.5 μm in length, and 2.8 to 4.3 μm in width. The mature conidia were slightly curved, with round apex and truncate base, 1 to 5 oil globules, and were 13.3 to 23.8 μm in length and 3.0 to 3.9 μm in width, respectively. Appressoria were solitary or in loose groups, dark brown, irregular shapes, and were 6.8 to 9.2 μm in length and 4.3 to 7.1 μm in width. PCR amplification was carried out by utilizing the universal rDNA-ITS primer pair ITS4/ITS5 (1) and the actin gene primer pair ACT-512F and ACT-783R (2). The PCR products of ITS (GenBank Accession No. KC913201) and actin gene (KC913202) from six isolates were identical, respectively, and shared 100% identity to the ITS sequence of strain CBS 167.49 of Colletotrichum spaethianum (GU227807.1) and 99% similarity to the actin gene of strain CBS 167.49 of C. spaethianum (GU227905.1), which was isolated from Hosta sieboldiana in Germany (3). Based on the above, the isolates were identified as C. spaethianum. To confirm pathogenicity, conidial suspensions (105 conidia ml–1) of each of the six isolates were sprayed on four leaves per plant on five 6-month-old P. praeruptorum plants. Control plants were sprayed with water. Plants were maintained at 28°C in a greenhouse with constant humidity (RH 90%) and a 12-h photoperiod of fluorescent light. Symptoms similar to the original ones started to appear after 10 days, while the control plants remained healthy. The tests were repeated three times and the fungus was recovered and identified as C. spaethianum by both morphology and molecular characterization. To our knowledge, this is the first report of C. spaethianum causing leaf spot on P. praeruptorum in China. Since the C. spaethianum infections pose a serious threat to P. praeruptorum production, this disease needs to be considered for developing effective control strategies.
References: (1) I. Carbone and L. M. Kohn. Mycologia 91:553, 1999. (2) U. Damm et al. Fung. Divers. 39:45, 2009. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.