First Report of Sanqi (Panax notoginseng) Root Rot Caused by Pythium vexans in China.

Abstract

Sanqi (Panax notoginseng (Burk.) F. H. Chen) is a precious traditional Chinese herbal medicine. During April of 2021, a root rot disease with approximate 15% incidence was observed on 2-year-old Sanqi plants in a field of Zhouning (27º12' N, 119°33' E), Fujian Province of China. The disease symptoms included severe stunting, leaf chlorosis, root rotting and necrosis, as the disease progressed, the whole plant gradually wilted and died. To recover the causal agent, symptomatic roots were excised, surface sterilized in 75% alcohol for 1.5 min, rinsed in sterilized water three times, dried, and placed on PARP selective medium (Jeffers and Martin 1986), and incubated at 20°C in dark. After 5 days, total of 26 Pythium-like isolates were obtained, and one representative isolate Py21-6 (available from the Institute of Plant Protection, Fujian Academy of Agricultural Sciences) was selected for further identification. Colonies of Py21-6 on PARP plate were white with dense, cottony, aerial, and transparent mycelia. Sporangia were terminal or intercalary, non-papillate, spherical, pyriform or ovoid, measuring 21.7 ± 2.8 × 19.3 ± 2.3 μm (n = 30). Zoospores were saucer-like, released out of sporangium after maturation, and dispersed quickly by swimming. Oogonia were spherical, terminal or occasionally intercalary. Oospores were globose, smooth and aplerotic. The dimensions of zoospores, oogonia, and oospores were 6.8 ± 0.7 μm, 21.6 ± 2.2 μm and 18.2 ± 2.7 μm (n = 30), respectively. Antheridia were bell-shaped or irregular, terminal, monoclinous, and usually one per oogonium. According to the morphological characteristics the isolate was initially identified as Pythium spp. (Van der Plaats-Niterink 1981, Yong et al. 2016). For further identification, DNA extracted from Py21-6, the cytochrome c oxidase subunit I (COI) gene and internal transcribed spacer (ITS) region were amplified and sequenced with primers FM55/FM52R (Long et al. 2012) and ITS1 /ITS4 (White et al. 1990), respectively. BLAST analysis of 680-bp COI (OM688194) and 728-bp ITS (OM663703) sequences revealed 99.86% and 99.99% similarity to Pythium vexans in GenBank (HQ708995 [COI], GU133572 [ITS]). Therefore, the pathogen was identified as P. vexans. In order to fulfill Koch's postulates, isolate Py21-6 was grown on Martin's liquid medium (Martin 1992) for 72 h to produce a spore suspensions of 106 oospores/ml, and the pathogenicity test was conducted by root-dip method. Three groups of 2-year-old Sanqi (15 plants per group) with root soaked for 20 min in oospore suspension were used for pathogenicity, and the other three groups (15 plants per group) with root dipped in sterilized water as control. All treated plants were replanted in (15-cm-diameter) pots (2 plants/pot) filled with mixture of sterilized soil: vermiculite: pearlite (2:1:1, v/v), maintained in greenhouse under 60% black shade cloth at 20 to 26°C with 80% relative humidity, and watered once every three days. After 21days, all inoculated plants showed the same symptoms observed on the original diseased plants in the field, whereas, the control plants remained symptomless. The same pathogen was successfully re-isolated from the inoculated plants, and identical to those of the originals based on morphological and sequence data. To our knowledge, this is the first report of P. vexans causing root rot on Sanqi in China (Farr and Rossman 2022). Root rot is one of the destructive diseases in Sanqi production, identification of the pathogen will be useful to develop effective field management strategies to control this disease.