<i>Erysiphe alphitoides</i> Causing Powdery Mildew on <i>Euonymus fortunei</i> in China

Euonymus fortunei, known as purple wintercreeper, is a widely grown evergreen vine with high horticultural and ornamental value in China (Zhao et al., 2003). Moreover, for hundreds of years, E. fortunei has been used as a traditional Chinese medicine. It has been widely used in various treatments of diverse maladies (Qu et al., 2002). In May 2021, powdery mildew symptoms were observed on 28 out of 50 inspected E. fortunei plants (56% incidence rate) of the cultivar Dart's Blanket at Henan Normal University, Xinxiang City, Henan Province, China (East Campus,113.36°E, 34.55°N). Subsequent annual surveys (2022-2025) confirmed disease recurrence with similar symptomology. White colonies were observed on adaxial and abaxial surfaces of leaves and covered up to 70 % of the leaf area, irregular chlorotic spots appear on the adaxial leaf surface. A total of 50 plants were systematically surveyed using a 5-point sampling method, with 3 symptomatic leaves per plant collected aseptically in sterile paper bags. With light microscope observations, foot cells of the powdery mildew (n = 50) were straight and 40 to 70 μm long. Cylindrical conidiophores (n = 50) were 60 to 100 × 6 to 14 μm with 3 to 4 conidia. The ellipsoid-ovoid-shaped conidia (n = 50) were 22 to 31 × 12 to 15 μm with a length/width ratio of 2.0 to 2.5. These morphological characteristics were similar to the previously reported powdery mildew fungus E. alphitoides. (Braun and Cook 2012). The powdery mildew pathogen was purified by directly transferring conidia and hyphae from infected leaves to healthy leaves of E. fortunei, with three successive subculturing cycles to obtain a pure isolate. Following previously described methods (White et al. 1990; Bradshaw et al. 2022; Zhu et al. 2022), the ITS (ribosomal transcribed spacer) and GAPDH (glyceraldehyde-3-phosphate dehydrogenase) gene regions of three isolates were amplified with specific primers ITS1/ITS4 (ITS1 5ʹ-TCCGTAGGTGAACCTGCGG-3ʹ; ITS4 5ʹ-TCCTCCGCTTATTGATATGC-3ʹ) and PMGAPDH1/PMGAPDH3R (PMGAPDH1 5ʹ-GGAATGGCTATGCGTGTACC-3ʹ; PMGAPDH3R 5ʹ-CCCCATTCGTTGTCGTACCATG-3ʹ), all three isolates shared identical ITS and GAPDH gene sequences, the sequences deposited in GenBank (ITS: OP102485, GAPDH: PV567403). BLASTn analysis showed that the ITS sequence shared 99.35% homology with E. alphitoides (OR940029) on Quercus sp., the GAPDH shared 95.67% homology with E. alphitoides (OR940020) on Quercus robur. The phylogenetic analysis clearly illustrated that the identified fungus clustered in the same clade with a previously reported E. alphitoides. To confirm the pathogenicity of identified powdery mildew, inoculation assays were conducted. The pathogenicity was tested according to the method previously described (Zhu et al. 2021). By blowing conidia on infected leaves with pressurized air, the fungus was inoculated onto the leaves of nine healthy 15-day-old Dart's Blanket E. fortune, while Nine uninoculated E. fortune served as controls. The infected plants and the control plants were placed in the culture room with the temperature 23 °C, humidity 50% and light/Dark 16/8 h, respectively, 7-11 days post inoculation, the leaves of the inoculated plants showed signs and symptoms of powdery mildew, while the control group was unaffected. After the disease occured, the symptoms and morphological characteristics of the pathogen were checked under the microscope and compared with the originally collected powdery mildew. The pathogenicity tests were repeated three times and the same results were obtained. Therefore, the pathogen was identified and confirmed as E. alphitoides (isolate ZM202201). the best of our knowledge, this is the first record of powdery mildew caused by E. alphitoides on E. fortunei in China. The outbreak of this economically important powdery mildew disease detrimentally affects plant health and thus may impact the ornamental and medical value of E. fortunei. This study provides the first confirmed report of E. alphitoides infecting E. fortunei in China, establishing a new host-pathogen relationship that informs disease management strategies for this economically important ornamental and medicinal plant species.