First Report of Ilyonectria robusta Causing Rusty Root of Asian Ginseng in China.

Abstract

Asian ginseng (Panax ginseng) is an economically important perennial herb, mainly cultivated in Jilin Province, China. In September 2013, Asian ginseng plants in Jilin showed rusty root symptoms. Typical symptoms included rusty superficial lesions of irregular shapes and margins. Ten symptomatic roots were collected from each of five fields for investigation. To isolate the pathogen, root epidermal tissues with typical lesions were excised, surface-sterilized, and placed on potato dextrose agar (PDA) amended with 50 μg/ml tetracycline. After incubation at 20 ± 1°C in the dark for a week, 18 single-spore isolates out of 50 samples were obtained and identified as Ilyonectria robusta (A.A. Hildebr.) A. Cabral & Crous based on morphological characters and DNA sequence analysis (1). After incubating 7 days on PDA in the dark at 20°C, colonies were cottony to felty in texture and orange white to brownish grey in color with average diameters of 60 ± 3 mm. Isolates were cultured on synthetic nutrient-poor agar for conidial measurements. Macroconidia formed on simple conidiophores predominately, with mostly one and occasionally up to three septa, and were cylindrical with both ends broadly rounded. Macroconidia varied in size depending on the number of cells as follows: one-septate, 7.0 ± 0.6 × 27.7 ± 2.7 μm; two-septate, 7.3 ± 0.7 × 33.3 ± 2.1 μm; three-septate, 7.4 ± 0.6 × 33.4 ± 2.2 μm. Microconidia that formed on complex conidiophores were ellipsoid to ovoid and ranged in size from aseptate 3.7 ± 0.5 × 8.7 ± 1.1 μm to one-septate 5.0 ± 0.6 × 13.1 ± 1.6 μm. Brown chlamydospores were abundantly produced on PDA, globose to subglobose in shape, and in size of 10.9 ± 1.3 × 11.8 ± 1.5 μm (n ≥ 30 observations per structure for each measurement). The isolates were further classified by amplifying and sequencing the ITS1-5.8S rRNA-ITS2 region and histone H3 gene with primer pairs ITS5 and ITS4 (4), and H3-1a and H3-1b (3), respectively. Sequences of the two loci (GenBank Accession Nos. KM015300 and KM015299) showed 100% identity among the three examined isolates and the published I. robusta isolates (JF735268 and JF735517). To confirm the pathogenicity, bare roots of 3-year-old Asian ginseng were inoculated with mycelial plugs of three isolates of I. robusta selected randomly. Four roots were inoculated as replicates for each isolate with pathogen-free agar plugs as a control. One week post-inoculation in the dark at 20 ± 1°C, all the inoculated ginseng roots showed light-brown to dark-brown lesions. I. robusta was recovered from symptomatic roots and confirmed by analyzing the DNA sequence of the histone H3 gene. The inoculation experiment was repeated, and both trials showed the same results. The ginseng tissue under the control agar plugs remained symptomless, and no fungi were isolated. To our knowledge, this is the first report of I. robusta causing rusty root of P. ginseng in China (1,2,5).