Manglietia decidua (Magnoliaceae) was a class I endangered plant in China. During 2018-2020, a severe root rot (about 10% - 90% disease incidence ) was observed on 2-year-old seedlings in the nursery in Yichun, Jiangxi province (N27°52’20”; E114°27’46”). Symptoms started on leaves showing dehydration and chlorosis, the root of diseased plant became black and rotted, and in severe cases, the plants withered and died. The symptomatic root tissues were cut and dipped in a 3% hydrogen peroxide solution for 5 mins, rinsed thrice with sterile water, and then placed on potato glucose agar medium containing ampicillin (50mg/L). The plates were kept in an incubator at 25-28°C for 2-3 days in the dark. The Calonectria-like fungus was consistently isolated from 100% of tissues and the colonies were feathery, moderate white aerial mycelium, surface pale brown, reverse with white outer margin, and brown inner region. The perithecia produced on carnation leaf agar were solitary, subglobose to ovoid, dark red-brown, and measured 273.8 - 427.2 × 362.6 - 628.9 µm (av. 360.9 × 429.9 µm) (n = 31). Clavate asci contained eight spores and tapered into a long thin stalk. Ascospores were hyaline, guttulate, straight to slightly curved with rounded ends, 36.8- 66.1×4.4-7.3 μm (av. 49.9 × 5.9 μm) (n = 52), 1-septate, constricted at the septum and aggregated in the upper third of the ascus. On PDA，conidia formed on penicillate conidiophores within 10 days were hyaline, 1(-3)-septate, cylindrical, rounded at both ends, straight, 36.5-61.7 × 5.0-7.2 μm (av. 50.7 × 6.2 µm) (n=48). Isolate HML 20 and 27 were used to further confirm species identity by five loci analysis：ITS (MZ389092 and MZ389093), ACT (MZ398252 and MZ398253), HIS3 (MZ398254 and MZ398255), TEF1-α (MZ398256 and MZ398257), and TUB2 (MZ398258 and MZ398259). NCBI BLASTN showed the high sequence identity with Calonectria ilicicola ex-type culture CBS 190.50 (CMW 30998) (Liu et al 2020): 100 % for ITS (MT359727), TUB2 (AY725631), and HIS3 (MT335506), 99.22% for ACT (MT335036), 99.80% for TEF1-α (MT412797). Maximum likelihood (ML) analysis and Bayesian inference (BI) based on the combined ITS, tub2, his3 and tef1 sequence using RAxML v.1.0.0 and MrBayes v. 3.2.1 software revealed that isolate HML 20 and 27 clustered together with C. ilicicola strains in C. kyotensis species complex. Thus, the fungus was identified as C. ilicicola (anamorph: Cylindrocladium parasiticum) based on morpho-molecular criteria (Lombard et al. 2010). Pathogenicity was determined under greenhouse conditions (25-30 ℃). The 2-year-old plants grown in 25-cm pots for 20 days were inoculated. Five 6-mm mycelial plugs from 7-day culture on PDA were buried 5 cm under the soil adjacent to the unwounded taproot of each plant and the plants were watered regularly to keep the soil moisture content at about 15%. After ten days, inoculated plants began to show chlorosis symptoms on leaves and collapsed within 15 to 20 days, while no symptoms were observed on control plants. The same colonial fungus was successfully reisolated. Calonectria ilicicola is an economically important plant pathogen worldwide, which causes diseases on Arachis hypogaea, Cinnamomum kanahirai, Glycine max, Medicago sativa, Sassafras randaiense, and Vaccinium spp. etc. in China (Gai et al 2017, Fei et al 2018, Zhang et al 2020 ). As far as we know, it is first report of C. ilicicola causing root rot on M. decidua. At present, this disease is an important threat to the conservation of M. decidua.
First Report of Root Rot Caused by Calonectria ilicicola on Blueberry in Yunnan Province, China