First Report of Leaf Spot caused by Alternaria alternata on Zanthoxylum dissitum in China

Zanthoxylum dissitum Hemsley is a rare medicinal plant (family Rutaceae) native to the southern China provinces of Guangdong, Guangxi, Hunan, and Sichuan, where it grows well in mountainous regions. Its roots, stems, and leaves are commonly used as a Chinese herbal medicine. In November 2012, symptoms of leaf spot were found on Z. dissitum leaves in Long'an County, Guangxi. The disease was found both in nurseries and in fields in Long' an, and also in fields in Guangxi Province. The incidence was less than 10% and only a few leaves per plant were affected. Lesions were initially small, brown, and oval or circular and developed into dark spots sometimes with yellow halos. A fungus was consistently recovered from symptomatic leaf samples (85% isolation rate from 40 leaf pieces), that had been surface sterilized with 0.1% mercuric chloride solution for up to 30 s. On potato dextrose agar (PDA), the cultures initially developed white colonies and the centers turned gray or brown after 5 days of incubation at 25[degrees]C. Conidiophores were single or fasciculate, straight or curved, brown with regular septa, and 35 to 65 x 4 to 5.5 [mu]m. Conidia were obclavate or ovate, brown, and 23 to 40 x 11 to 19 [mu]m with three to five transverse and one to three longitudinal or oblique septa. Club-shaped conidia were produced in branching chains. The fungus was identified morphologically as Alternaria alternata(Fr.) Keissl, based on descriptions in Simmons (2007) and (Zhang 2003). Small pieces of symptomatic tissues (4 mm super(2)) were plated on antibiotic-amended PDA, and grown at 25[degrees]C. Spores were obtained from cultures and streaked on fresh-water agar. Single-spore germinating cultures were transferred to fresh media. A single-spore isolate, Zhong1-1, was selected for molecular identification, and genomic 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. KT028771), and it showed 100% identity to Alternaria alternata(AY513941). Detached young, healthy leaves from 4-year-old, 1.5-m-high plants of Z. dissitum were inoculated with 5-mm-diameter mycelial plugs from PDA cultures that had been incubated for 7 days at 25[degrees]C. Control leaves were treated with agar plugs only. Leaves were placed in trays and covered with polythene film to maintain high humidity, and incubated at 25[degrees]C with a 12-h photoperio