First Report of Leaf Spot on <i>Cinnamomum burmanni</i> Caused by <i>Colletotrichum siamense</i> in China

Cinnamomum burmanni is an important essential oil economic and greening tree species, widely used in chemical industry, medicine and other fields (Reis-avila G et al. 2017). In May 2025, severe anthracnose was observed on C. burmanni in the campus greening area of Qujing Normal University, Qujing City, Yunnan Province, China (103.789642 E, 25.482519 N). The planting area (~0.2-0.3 ha) with approximately 100-120 mature trees had 60-80% affected leaves. The disease typically initiates at leaf margins, manifesting as pale-yellow spots that expand into oval to irregular brown lesions with wide yellow halos. Lesions enlarge to grayish brown patches (3.6 × 3.3-10.0 × 3.6 cm), often exceeding half the leaf area. In advanced stages, coalesced lesions cause leaf necrosis and premature defoliation, with occasional small black acervuli on lesions. To isolate the fungus, 10 symptomatic leaves were randomly collected from five trees and rinsed with sterile water. Small, infected tissue segments (~ 5 mm 2 ) were surface-sterilized with 75% ethanol for 30 s, then 3% hypochlorous acid for 3 min, rinsed thrice with sterile water, and inoculated on potato dextrose agar (PDA). Plates were incubated at 28°C with a 12h light cycle for 7 days. Ten isolates with similar morphology were obtained; three representatives (MPSTJ1-3) were purified. Purified colonies initially had sparse edges on PDA, becoming dense after 3 days, covering the dish by 7 days, with round, flat edges. Three to five brown-black ring marks with clear boundaries were observed, with black conidiophores appearing later. Microscopically, conidia were colorless, cylindrical, obtuse to slightly pointed, (8.25 - 19.67) × (5.92 - 8.47) μm, avg. (16.62 ± 2.77) × (7.20 ± 1.28) μm (n = 50). These morphological characteristics resembled Colletotrichum siamense (Qin et al. 2021). For confirmation, genomic DNA from the three isolates was extracted; sequences of ITS, GAPDH, ACT, CHS1, CAL, and TUB2 were amplified (Zhang et al. 2020), with GenBank accessions: ITS (PX844593-PX844595), GAPDH (PX996135-PX996136), ACT (PX996137- PX996139), CHS1 (PX996140-PX996142), CAL (PX996134, PX996143-PX996145), TUB2 (PX996146-PX996148). Phylogenetic analysis of six combined genes clustered the isolates within C. siamense clade, confirming identification. Pathogenicity of MPSTJ1-3 was tested on healthy 1-year-old leaves (needle-wounded, 5 mm mycelial plugs, 3 replicates). Inoculated leaves were incubated at 25-30°C, 85-90% RH, 12 h photoperiod. Five days post-inoculation, leaf spots developed on all inoculated leaves; controls (PDA plugs/sterile water) remained symptom-free. The fungus was re-isolated and identified as C. siamense, confirming Koch's postulates. C. siamense infects at least 60 plant species worldwide (Ji et al. 2019). To our knowledge, this is the first report of C. siamense causing leaf spot on C. burmanni in China via multilocus phylogenetic analysis. This finding aids targeted disease management to protect C. burmanni plantations, as C. siamense anthracnose reduces essential oil yield and quality, and supports the need for research on host-pathogen interactions and resistance breeding.