Identification and Quantification of Pathogenic Binucleate Ceratobasidium sp. AG-K Responsible for Root Coarse Bark Disease (Makou) in Angelica sinensis in Dingxi, China

Makou disease is a grave disease in the production of Angelica sinensis, seriously affecting this plant’s yield and quality. Its typical symptoms include yellowing and necrotic spots on leaves of infected plants, and the belowground symptoms are longitudinal brown cracks (depth is about 1 to 2 mm) in the root epidermis. The symptoms of Makou disease are completely different from those of root rot disease, and the pathogen responsible has yet to be officially reported and is highly controversial surrounding the existing research. Symptomatic roots and infested soils were collected from 18 fields, from which 161 fungal isolates were isolated. Through morphological observation, 105 isolates were taxonomically identified by comparing the sequences of their internal transcribed spacer (ITS) region to those of known species in the NCBI database. Ceratobasidium sp. AG-K was the most abundant species (61.0%), followed by Fusarium spp. (31.4%) and Plectosphaerella cucumerina (5.7%). Phylogenetic analyses based on sequences of ITS of those isolates showed that those belonging to the same species were clearly separated in the derived dendrogram. Two representative isolates, R2 and P3, were selected for the pathogenicity test. Pathogenicity testing revealed Ceratobasidium sp. AG-K was the pathogenic fungus causing Makou disease in A. sinensis seedlings. Additionally, a strain of Bacillus sp. with inhibitory activity against Ceratobasidium sp. AG-K was identified in vitro. The RT-qPCR results showed that the abundance of Ceratobasidium sp. AG-K in the rhizosphere soil of seedlings inoculated with this pathogen reached 10 times that of the control group. Understanding the influence of Ceratobasidium sp. AG-K upon the occurrence and severity of Angelica Makou disease outbreaks will provide key information for devising prevention and control strategies. Furthermore, the detection method we developed can provide future guidance and a sound basis for studying the pathogen’s dynamics in the rhizosphere of A. sinensis.