First report of Root rot in Cerasus subhirtella Caused by Fusarium solani and Fusarium oxysporum in China.

Abstract

Cerasus subhirtella (Miq.) Sok. is a widely used ornamental tree in urban areas around China and has a high ornamental value. From 2018 to 2020, a root rot disease was observed in C. subhirtella in Meitan County, Weng'an County, and Guiyang city of Guizhou, China (106.71 E, 26.57 N). Diseased C. subhirtella trees exhibited wilting with leaf chlorosis accompanied by brown to black root discoloration. In an area of 100 ha in total, with disease incidence ranging from 60 to 80%. Six symptomatic plants with root rot were randomly collected from three locations where disease symptoms were observed for pathogen isolation. Fifty fragments of diseased roots (5×5mm) were disinfected in 3% sodium hypochlorite for 30 s and 75% alcohol for 60 s, rinsed three times in sterile distilled water, plated on potato dextrose agar (PDA; BoWei, Shanghai), and incubated at 28 °C in the dark for 7 days. Eighteen isolates were purified by single spore culturing. Typical Fusarium spp. colonies were obtained from all root samples. On PDA, the colonies showed white and the hyphae were dense, while the colony of isolate YH15 showed pale yellow on the back, radial growth and produced chlamydospores. The macrospores (YH15) were straight to subarcuate, measured 15.3 to 25.1 × 2.5 to 6.2 μm (n=50). The microconidia (YH26) were ellipsoid to ovoid, measured 8.6 to 12.7 × 1.6 to 5.1 μm (n=50). These morphological characteristics were consistent with Fusarium spp., as described recently in Vitullo et al. (2014). To confirm the morphological diagnosis, genomic DNA from the isolates was extracted. The internal transcribed spacer (ITS) (White et al, 1990) region of rDNA and a β-tubulin (Varga et al, 2011) gene fragment were amplified with the primers ITS1/ITS4 and Bt2a/Bt2b, respectively, and were subsequently sequenced. Maximum likelihood analysis was carried out using MEGA 11.0. BLAST analysis revealed that the ITS and β-tubulin sequences of isolate YH15 were 100% homologous with F. oxysporum, and the isolate YH26 had a 99.69~100% homology with F. solani. Sequences of isolate YH15 and YH26 were deposited in GenBank (ITS: OQ363005 and OQ363049; β-tubulin: OQ398187 and OQ398180). The isolate YH15 was thus identified as F. oxysporum by the morphological characteristics and sequences analysis, and the isolate YH26 was identified as F. solani. A reconstructed phylogenetic tree also confirmed their phylogenetic position. The healthy 2-year-old C. subhirtella plants grown in autoclaved acid yellow soil were used for the pathogenicity tests. Then, 50 mL of conidial suspension (2.0×105 conidia/mL, in medium) of 7-day-old isolates YH15 and YH26 were gently applied to the soil in each of the 10 pots as the treatment. A sterilized fungal culture matrix (PDB; BoWei, Shanghai) was applied to each of 10 pots as a control. All pots (30 cm high, 25 cm upper diameter, 15 cm base diameter) were placed in a greenhouse (25 °C, 12 h photoperiod). After 30 days of inoculation, all plants inoculated with the isolates showed wilting symptoms, and the roots showed light-brown to dark-brown lesions. No symptoms were observed in the controls. The pathogen was reisolated from all symptomatic roots and identified as F. oxysporum and F. solani as described above. The pathogenicity test was repeated twice with similar results. Although this fungus was previously reported to cause root disease in many hosts (Li et al., 2020; Gibert et al., 2022), this is the first report of F. oxysporum and F. solani causing root rot in C. subhirtella in China.