First Report of Leaf Spot Caused by Alternaria alternata on Ginseng (Panax ginseng) in China

Abstract

Ginseng (Panax ginseng) is a perennial herbaceous plant and a precious traditional Chinese medicine. P. ginseng is widely cultivated in Asia, particularly the Changbai Mountain area in China (Wang et al. 2019). Alternaria species can grow under a wide range of temperatures that occur worldwide (Konstantinova et al. 2002). Alternaria panax was reported as the main cause of Alternaria leaf spot of ginseng (Deng et al. 2013); however, a small-spored Alternaria species was isolated from leaf spot symptoms during recent surveys. In our study, leaf spot disease of P. ginseng was observed during 2017 in the Fusong district, Jilin, China. The disease was widespread, with 20 to 30% of the fields affected. Leaves of the affected plants showed brown to black necrotic spots. The fungus was isolated from symptomatic leaves on potato dextrose agar and characterized morphologically under the microscope. Colonies of pure cultures were initially dark brown and then turned black after 7 days. Strains (TCS 15013001 and TCS 15013002) were grown on water agar embedded with surface-sterilized ginseng spot tissues under a 12/12-h light/dark cycle at room temperature. Conidia observed on the lesions were dark brown or olive-brown, ovoid or ellipsoid, short beaked, with three to seven transverse and zero to five longitudinal septa, and 14.9 to 36.5 μm (average 25.8 μm) × 5.8 to 17.9 μm (average 12.5 μm). Based on its morphological characteristics, the pathogen was identified as Alternaria sp. (Simmons 2007). Single-spore cultures were obtained, and their genomic DNA was extracted. The rDNA internal transcribed spacer (ITS, GenBank accession nos. MN394879 and MN394880), endopolygalacturonase (endoPG, MN410909 and MN410910), Alternaria major allergen (Alta1, MN410911 and MN410912), and the portion of the histone 3 (H3, MN410913 and MN410914), elongation factor 1-alpha (EF1-α, MN410915 and MN410916), the RNA polymerase II 2nd largest subunit (RPB2, MN410917 and MN410918), and glyceraldehyde 3-phosphate dehydrogenase (gpd, MN410919 and MN410920) gene were amplified from the two isolates, respectively (Woudenberg et al. 2015). Sequence analysis of the two isolates showed 99 to 100% identity to that of A. alternata (MG182428 for ITS, MG598798 for endoPG, MG598796 for Alt a1, MG182429 for H3, KU738723 for EF1-α, KU738707 for RPB2, and KP851761 for gpd). Pathogenicity tests were conducted in a greenhouse. Sixty healthy 2-year-old P. ginseng seedlings grown in potting medium were tested in a greenhouse at 20/26°C and about 90% relative humidity with natural light. Of the plants, 30 were sprayed with spore suspension (1 × 10⁵ spores/ml) until most of the leaves were wet. Control treatments (30 plants) were treated with distilled water. The two isolates were tested, and all test plants were covered with clear plastic bags for 3 days to increase humidity. Each treatment had three replicate plants. Six days after inoculation, leaf spots similar to the original ones developed at inoculation sites for all isolates, and A. alternata was consistently reisolated from the induced symptoms. The pathogenicity test was performed three times. A. alternata causing leaf spot of American ginseng was reported in China (Hill and Hausbeck 2008). Also, leaf spot of Panax notoginseng caused by A. alternata was reported in China (Yang et al. 2018). To our knowledge, this is the first report of A. alternata on P. ginseng in China.