First Report of <i>Pestalotiopsis kenyana</i> Causing Leaf Sheath Spot on <i>Coix lacryma-jobi</i> in China

Coix lacryma-jobi is a hardy grass valued for its edible, pearl-like grains and medicinal properties. In July 2025, a gray white leaf sheath spot disease on C. lacryma-jobi was found in a field in Fuyuan Yunnan province, China (25.66 °N, 104.25°E). About 35% of inspected plants (n = 200) showed symptoms, leading to considerable economic loss. Infected leaf sheaths displayed irregular gray-white lesions with water-soaked margins, later developing black conidiomata on the surface. To identify the pathogen, 15 symptomatic sheaths were collected from different plants. Black pycnidia were picked with a sterile needle, placed on water agar, and incubated at 25 °C for 24 h. Germinated spores were transferred to potato dextrose agar (PDA) for purification and morphological characterization. Three purified isolates (YR01–YR03) were obtained. On PDA, colonies reached 35–45 mm diameter after 10 days at 25 °C, with uneven margins, whitish aerial mycelium of medium density, black gregarious conidiomata, and yellowish reverse pigmentation. Conidia were fusoid, straight or slightly curved, 4-septate, 23–29 × 7–9 μm (n = 50). The basal cell was conical to obconic with a truncate base, hyaline, 3–5 μm long. The three median cells were doliiform, 16–18 μm long, pale brown, with septa darker than the cells; the second cell from the base measured 5.6–7 μm, the third 6–7.5 μm, and the fourth 5.6–7 μm. The apical cell was 3–4.5 μm long, filiform, hyaline, cylindrical to subcylindrical, bearing 3–4 tubular apical appendages 9.0–16.4 μm long. A solitary, tubular, unbranched basal appendage measured 4.3–7.6 μm. These traits matched published descriptions of Pestalotiopsis kenyana Maharachch., K.D. Hyde & Crous (Maharachchikumbura et al. 2014). For molecular identification, genomic DNA was extracted, and the internal transcribed spacer (ITS), β-tubulin (TUB), and translation elongation factor 1-alpha (TEF1) genes were amplified using primers ITS4/ITS5, T1/Bt2b, and EF1-728F/EF2 (Li et al. 2024), respectively. BLAST analysis of the sequences (GenBank accessions: PX411086–PX411088 [ITS], PX610109–PX610111 [TUB], PX610112–PX610114 [TEF1]) showed 99.64%, 99.58%, and 99.73% identity, respectively, with P. kenyana strain CBS 442.67. A maximum-likelihood phylogeny of concatenated sequences grouped the isolates with other P. kenyana references with 90% bootstrap support. To assess pathogenicity, the fifth leaf sheath from the middle of ten C. lacryma-jobi plants was punctured with a sterile needle and inoculated with 20 μL of a conidial suspension (1 × 10⁶ conidia/mL), while ten control plants received sterile water. All punctured sites were covered with sterile cotton balls and wrapped with plastic wrap to maintain moisture for 48 hours. Symptoms similar to field observations developed on inoculated leaves within 7 days, while controls remained asymptomatic. The pathogen was reisolated from symptomatic sheathes, and its identity was confirmed by both morphology and DNA sequence, fulfilling Koch’s postulates. The experiment was repeated two times with consistent results. P. kenyana was reported to cause leaf spots on multiple hosts like Rhododendron agastum (Li et al. 2024), Zanthoxylum schinifolium (Liu et al. 2023) and Camellia sinensis (Wang et al. 2019). To our knowledge, this is the first report of P. kenyana causing leaf sheath spot on C. lacryma-jobi. This finding provides important insights for diagnosing and managing this emerging disease. It helps anticipate potential economic losses in this valuable crop and supports the development of targeted control strategies.