First Report of <i>Phyllosticta capitalensis</i> Causing Leaf Blight on <i>Quercus acutissima</i> in China

Quercus acutissima is a major afforestation species in Longquan Mountain Reserve Forest. In May 2024, over 10% of 100 two-year-old seedlings showed leaf blight symptoms in a local nursery in Longquanyi District, Chengdu (30°39′57″N, 104°20′40″E; alt. 470 m). Symptoms began at leaf margins, spreading inward and causing wilting, necrosis, and crinkling of the affected tissue. Conidiomata form on necrotic tissue during disease progression, typically observable on the upper leaf surface. Five representative samples were collected from different diseased plants. A single-spore isolation was performed following the method of Chomnunti et al. (2014). Five fungal isolates, each from a different infected leaf, were deposited in the SICAU Culture Collection. The colonies presented consistent traits: a gray-green color with a black reverse side, thin aerial mycelia, irregular margins, and a diameter of 50 mm after 30 days at 25°C. Morphological characteristics on the host were examined using light microscopy. Conidiomata were black, scattered, semi-immersed or protruding, and 0.1 to 0.2 mm. Conidia were round to oval, 6-12.5 μm (x̄ = 8.7 μm, n = 50), transparent, thin-walled, smooth, with a single large central cavity, arounded with a thin gelatinous sheath. Each conidium bore a hyaline, linear apical appendage measuring 6.5-9.5 μm (x̄ = 7.8 μm, n = 50). The morphology matched the sexual stage of Phyllosticta capitalensis Henn. (Wikee et al. 2013a). The ITS, LSU, tef1-α, act, and gapdh regions were amplified using the primers ITS5/ITS4, LR0R/LR5, EF1-728F/EF-2, ACT-512F/ACT-783R, and Gpd1-LM/Gpd2-LM, respectively (Jiang et al. 2024). Sequences were deposited in NCBI with accession numbers, viz. SICAUCC 24-0181: PQ721658, PQ721643, PQ728300, PQ728301, and PQ728302; SICAUCC 25-0128: PV639329, PV639330, PV657991, PV657992, and PV657993. BLASTn analysis showed that sequences from SICAUCC 24-0181 showed 99%-100% similarity to P. capitalensis (CBS 128856, epitype): 570/571 bp (ITS), 763/763 bp (LSU), 209/210 bp (tef1-α), 224/224 bp (act), 646/650 bp (gapdh) (Glienke et al. 2011). Phylogenetic analysis based on a concatenated dataset of the five sequences revealed that the two isolates clustered within the same clade as the P. capitalensis species complex. Based on morphological and molecular characteristics, the isolates were identified as P. capitalensis. Pathogenicity tests were conducted on eighteen healthy, two-year-old Q. acutissima seedlings with 80% relative humidity at 25 to 30°C. Nine healthy plants (five leaves each) were inoculated with 500 μl of spore suspension on the adaxial surface (1 × 105 conidia/ml), and nine control plants were treated with sterile distilled water. The experiment was repeated three times. After fifteen days, similar symptoms appeared on the inoculated leaves as in the field, while no symptoms were observed on the control plants. The fungus reisolated from inoculated leaves was identified as P. capitalensis, and no pathogen was isolated from control plants, fulfilling Koch’s postulates. Phyllosticta capitalensis has previously been reported to cause leaf diseases on various host plants worldwide (Wikee et al. 2013b). However, this study is the first to report P. capitalensis as the causal agent of leaf blight on Q. acutissima in China. Further research is needed to understand the disease’s spread, impact, and effective control measures to prevent its dissemination.