W. X. Yang,
X. D. Ren, and
D. Q. Liu, Department of Plant Pathology, Biological Control Center of Plant Diseases and Plant Pests of Hebei Province, National Engineering Research Center for Agriculture in Northern Mountainous Areas, Agricultural University of Hebei, Baoding 071001, Hebei. This study was supported by the Technology Support Program of Hebei Province (10220309)
Since 2009, brown leaf spot and panicle blight of Zanthoxylum piperitum (L.) DC. (“liujin,” commonly known as Japanese pepper and Japanese pricklyash) has been observed on 40% of the plants in the test field of Foresty Academy of Science in Hebei Province of China. When symptoms formed on leaves, a thick yellow spot appeared, which then turned brown. When on the spikes, brown lesions were observed initially on the grain, which then spread down to fruit stem, and finally the whole spike wilted and dried up. Yield and quality losses were considerable. A fungus was isolated consistently from the diseased tissues using potato dextrose agar (PDA) (1). Three representative isolates were chosen for further characterization. All the isolates grew at 28°C on PDA and potato carrot agar (PCA) medium. Fungal colonies were initially white, then became olivaceous with some white mycelium on the top of the colony, and turned brown with age. When observed with the microscope, crineous septate hypha appeared, and conidiophore peduncles were upright or slightly curved, with a few branches, 33.0 to 75.0 μm long and 4.0 to 5.5 μm wide. Conidia were crineous short clubs or near oval in shape, 22.5 to 40.0 μm long and 8.0 to 13.5 μm wide, with a short conical beak, and had one to four longitudinal cross walls. On PCA, condia had three to seven transepta and one to five longisepta, and were produced in a branched, long chain with more than five conidia. The pathogen was identified based on morphological characteristics as Alternaria alternata (Fr.:Fr.) Keissl. (3). DNA was extracted from mycelium and PCR was performed on the internal transcribed spacer (ITS) region with primers ITS1 and ITS4. A 570-bp fragment was amplified and sequenced (GenBank Accession No. JQ973810). BLASTn analysis revealed there was 100% sequence identity with A. alternata strains (GU566303 and GQ121322). To further identify the fungus, A. alternata species-specific primers AAF2/AAR3 (2) were used to generate an amplicon which was then sequenced (JX308287). Sequence comparison showed there was 100% sequence identity with A. alternata (JQ927300 and JQ907485). Pathogenicity tests were performed by spraying with a cultured suspension (106 spores/ml) of approximately 100 μl onto healthy leaves in 15-cm-diameter glass dishes containing sterilized filter paper soaked with sterilized water at room temperature. Control plants were inoculated with sterile distilled water. Ten days after inoculation, symptoms were observed in all inoculated leaves and appeared to be identical to those observed in the field. No symptoms were noted on the control leaves. Identical results were also obtained when spikes were inoculated. The fungi reisolated from symptomatic plants were A. alternata. To our knowledge, this is the first report of A. alternata causing leaf spots and panicle blight of Z. piperitum in China.
References: (1) O. D. Dhingra and J. B. Sinclair. Basic Plant Pathology Methods. CRC Press, Boca Raton, FL, 1995. (2) P. Konstantinova. et al. Mycol. Res. 106:23, 2002. (3) T. Y. Zhang. China fungi records (Alternaria) (Volume 16) (in Chinese). Beijing: Science Press, 2003.