In May 2011, samples of Chinese cabbage (Brassica campestris L. subsp. chinensis Markino) seedlings at the two-to four-leaf stage with damping-off symptoms were collected from greenhouses in the Tibet Autonomous Region of China. Infected stems of the seedlings were constricted at or near the soil surface. On diseased stems, a light to dark brown coloration was demarcated from healthy tissue. Damping-off and death of seedlings occurred as the lesions enlarged, resulting in a significant reduction of seedlings. Diseased stems were cut into 3-mm-long segments, surface-sterilized with 3.5% sodium hypochlorite for 1 min, and rinsed in sterilized water three times before being placed on water agar. A fungus frequently isolated from diseased plants was hyphal tipped under a dissecting microscope and transferred onto potato dextrose agar (PDA) to obtain a pure culture. The isolate grew slowly on PDA at 25°C with a 12-h photoperiod. The colony was white at first and gradually turned gray or grayish-green. No conidia or chlamydospores developed. However, conidia were produced on potato-carrot agar. Conidia were yellow-brown, obpyriform or obpyriform with beaks, had two to eight transverse septa, one to three vertical or oblique septa, and were produced solitarily or often in chains of two or three and measured 43.5 to 85.2 × 21.5 to 28.0 μm. A few conidia without beaks were also present and were nearly round and slightly smaller than the conidia with beaks. Several adjacent chlamydospores with thickened walls were often intercalary. The isolate was tentatively identified as Alternaria japonica based on its morphological characteristics (1,2). For molecular analyses, the internal transcribed spacer (ITS) regions of ribosomal DNA from the isolate were amplified with universal primers ITS1 and ITS4. The resulting sequence (Accession No. JN654465) submitted to GenBank had a 99% identity to that of A. japonica (Accession No. AY154703.1) isolated from leaves of Raphanus sativus. To confirm the pathogenicity of A. japonica, nine healthy 10-day-old Chinese cabbage seedlings were inoculated at the stem base with one PDA plug from a 6-day-old culture, with nine noninoculated (PDA plus only) seedlings serving as controls. Two days after inoculation, symptoms similar to those on the naturally infected plants developed on the inoculated seedlings. No symptoms developed on the controls. The pathogen was reisolated from the stems of inoculated and diseased seedlings. To our knowledge, this is the first report of A. japonica leading to damping off on Chinese cabbage seedlings in China.
References: (1) M. P. Corlett and M. E. Corlett. Can. J. Plant Pathol. 21:298, 1999. (2) T. Y. Zhang. Flora Fungorum Sinicorum: Alternaria (in Chinese) Vol. 16. Science Press, Beijing, 2003.