First Report of Root Rot in Phoebe bournei Caused by Fusarium oxysporum in Guizhou Province, China

Phoebe bournei, a national second-class protected plant in China, features superior wood (50,000–100,000 CNY/ton) and is widely cultivated in southern China for high economic value. In November 2024, wilt symptoms were observed in the P. bournei nursery in Longli County, Guizhou Province, China. The incidence of the disease was 40-45% (n=500). The initial symptoms manifested as leaf chlorosis spreading from the margin to the center, eventually causing the plant to wilt and die. Meanwhile, underground observations showed that the roots turned black, rotted, and exhibited epidermal detachment. Ten symptomatic roots were sampled as 1×1 cm tissue pieces and disinfected with 3% NaOCl for 30 s, followed by 75% ethanol for 1 min, and finally rinsed 5 times with sterile water before being incubated on potato dextrose agar (PDA) at 25 °C for 7 d. Single-spore isolation from 10 symptomatic roots yielded 15 isolates (1-2 per root). After 7 d of individual culture on PDA, 10 of these isolates showed consistent morphological characteristics and were obtained from different sampled plants. For isolates GF-1, GF-2, and GF-3, the colony was characterized by dense cottony aerial hyphae, with a white colony periphery and a purple central part. Microconidia were elliptical to reniform, measuring 7.9 to 9.3 × 2.5 to 3.0 μm (n = 50); macroconidia were slightly curved and falcate, measuring 13.7 to 19.3 × 3.1 to 4.5 μm (n = 50). These morphological characteristics are consistent with those of Fusarium oxysporum (Leslie et al., 2006; Lombard et al., 2019). To confirm the morphological diagnosis, genomic DNA was extracted from strains GF-1, GF-2, and GF-3 using the CTAB method (Sagar et al., 2014). The ITS, TEF1-α, and RPB2 genes of these strains were then amplified and sequenced for species identification (White et al., 1990; O'Donnell et al., 1998; Miller and Huhndorf, 2005). The ITS, TEF1-α, and RPB2 sequences of the three isolates were aligned to GenBank sequences; BLAST analysis showed identical sequences among GF-2, GF-3, and GF-1 across all three genes. For GF-1, sequence identity with F. oxysporum references was 100% (KU729045 ITS, 504/504), 100% (MZ359437 TEF1-α, 625/625) and 99.33% (LS479308 RPB2, 742/747). The sequences of isolate GF-1 were deposited in GenBank (ITS: PV864815; TEF1-α: PV933179; RPB2: PV999888). Isolate GF-1 was thus identified as F. oxysporum by morphological characteristics and sequence analysis. Fifteen two-year-old substrate-grown P. bournei seedlings were employed for pathogenicity assays; their roots were rinsed and soaked in a 1×10⁶ conidia suspension of GF-1 for 20 minutes, with sterile water as the control. Inoculated seedlings were transplanted into sterilized soil-filled pots (30 cm diameter × 50 cm depth) and cultured in a greenhouse (25 °C, 12-h photoperiod). After 30 d of inoculation, the leaves of the seedlings inoculated with GF-1 showed wilting symptoms, and the root systems showed typical root rot symptoms. The control group remained asymptomatic. The fungus re-isolated from the symptomatic tissues matched the original strains morphologically and genetically (based on ITS, TEF1-α and RPB2 sequences). Three separate trials confirmed the pathogenicity, fulfilling Koch's postulates. This is the first report of F. oxysporum causing root rot in P. bournei. The results of this study can lay the foundation for investigating the occurrence and prevalence of root rot in P. bournei, as well as its comprehensive prevention and control.