As an important industrial, pharmaceutical and evergreen shade tree (Singh and Jawaid 2012), the camphor tree (Cinnamomum camphora) has been coppiced in Jiangxi Province, China. From 2017 to 2020, we noticed many camphor trees with leaf spots, with an incidence estimated at 50 to 75%, which could severely inhibit leaf growth and reduce their biomass. A dark-green circle with a watery spot appeared on the infected leaves at the initial stage, and necrosis with forming shot-spots surrounded by yellow halos occurred (Figure 1 A). Five leaves with typical symptoms were sampled and washed with tap water for ca. 15 min. Isolation and morphological analysis were performed following the method of Bao et al. (2019). Among 61 fungal isolates, 49 showed the same culture characters. Colonies on PDA were villose and regular, the reverse was scarlet at the edge of the colony, which was ca. 8.75 cm after 7 days of inoculation (Figure 1 I). Chlamydospores were aseptate, dark brown, smooth, in chains or solitary, ellipsoidal to ovoid, 4.8–9.6 × 4.8–11.1 μm (Figure 1 J). The pycnidia were produced on PDA and varied from 47.4 to 85.8 µm (mean 60.2 µm) × 38.6 to 66.8 μm (mean 49.7 μm) (n = 17) (Figure 1 K). Conidia were hyaline, unicellular, elliptical to ovoid, 4.3-6.4 µm (mean 5.1 µm) × 2.3-3.3 µm (mean 2.8 µm) (n = 52) (Figure 1 L). Pathogenicity tests of isolate XW-9 was carried out in the field. Ten leaves were wounded with a sterilized insect needle and inoculated with mycelium plugs (7-mm diameter). Non-colonized PDA plugs served as the negative controlIn addition, conidial suspensions (105 conidia/mL) of isolate XW-9 were sprayed on surface-sterilized leaves with a further ten leaves being sprayed with sterile water as the control. Symptoms described in this study appeared in 100% of the mycelium-inoculated leaves and more than 80% of the conidium-inoculated leaves after 7 days post-inoculation (Figure 1 B-E). No symptoms were seen in the controls (Figure 1 C). Three days after inoculation, brown spots resembling those observed in the field developed on the inoculated leaves, and some lesions turned into shot holes on the infected leaves (Figure 1 G & H). However, no symptoms were observed on the controls (Figure 1 F). The fungus was re-isolated from the margins of the leaf spots and labelled P-XW-9A. The gene regions for ITS, LSU, tub2, RPB2 and ACT of isolates XW-9 and P-XW-9A were amplified and sequenced. The sequences of rDNA-ITS, LSU, tub2, RPB2 and ACT of XW-9 were GenBank MW142397, MW130844, MW165322, MW446945 and MW165324, respectively and those of P-XW-9A were GenBank MW142398, MW130845, MW165323, MW446946 and MW165325, respectively (Lumbsch, et al. 2000; Aveskamp, et al. 2009; Hou et al. 2020). Phylogenetic analysis using concatenated sequences of ITS, LSU, RPB2, and tub2 showed that isolates XW-9 and P-XW-9A formed a single clade with the reference strain of E. poaceicola CBS 987.95 (Figure 2). Thus, XW-9 was identified as E. poaceicola based on its morphological and molecular characteristics. Significantly, the recovered isolate P-XW-9A also aligned with E. poaceicola fulfilling the criteria for Koch's Postulates. E. poaceicola was only reported as a fungal pathogen of Phyllostachys viridis in China (Liu et al. 2020). To our knowledge, this is the first report of leaf spot disease on camphor trees caused by E. poaceicola in China and our findings will be useful for its management.
Essential oils of camphor tree (cinnamomum camphora nees & eberm) cultivated in Southern Brazil
