First Report of Epicoccum nigrum Causing Leaf Spot and Leaf Sheath Disease on Maize in Korea

Abstract

Maize (Zea mays L.) is one of the major food and feed crops in the Republic of Korea, and fungal foliar diseases can cause significant yield losses. In 2024 and 2025, maize plants showing spot symptoms on leaves and leaf sheaths were observed in commercial fields located in Yeoju, Yeongwol, Hwasun, and Yeosu, Korea. Spot symptoms included circular to irregular necrotic lesions on leaves and discolored lesions on leaf sheaths. Symptomatic tissues were cut into small pieces (1-3 cm), surface-sterilized in 1.5% NaOCl for 1 min, rinsed twice with sterile distilled water, and placed on potato dextrose agar (PDA) amended with chloramphenicol (50 μg/mL). Plates were incubated at 26°C in the dark. Fungal colonies produced abundant black sporodochia composed of aggregated conidiophores on PDA. Conidia were produced singly on densely compacted, non-specialized, determinate, slightly pigmented conidiophores. Conidia were globose to pyriform, 15-25 μm in diameter, with a funnel-shaped base and a broad attachment scar. Mature conidia were darkly pigmented, multicellular (dictyoconidia), and had a verrucose outer surface, consistent with descriptions of Epicoccum nigrum. For molecular identification, genomic DNA was extracted, and the internal transcribed spacer (ITS), large subunit (LSU), RNA polymerase II second largest subunit (RPB2), and beta-tubulin (TUB2) gene regions were amplified following Xu et al. (2022). The resulting sequences were deposited in GenBank under accession numbers (PX945540, PX945539, PX981668, and PX979520). BLAST analysis revealed approximately 99% of sequence identity to reference sequences of E. nigrum. Phylogenetic analysis based on the maximum likelihood method further confirmed that the isolates clustered with known E. nigrum strains previously associated with maize. A pathogenicity assay was conducted to fulfill Koch’s postulates. Leaves and stems of three-week-old healthy maize seedlings were inoculated with a conidial suspension (1 × 10⁵ conidia/mL). Control plants were treated with sterile distilled water. Inoculated plants were maintained in a dew chamber for two days and then transferred to a growth chamber at 26°C with 70% relative humidity. Necrotic lesions developed on inoculated leaves within 7 days, whereas control plants remained symptomless. The same fungus was re-isolated from symptomatic tissues and identified based on morphological characteristics and DNA sequence analysis, thereby fulfilling Koch’s postulates. Epicoccum species have been reported to cause leaf spot diseases on maize in other countries, including China and India (Chen et al. 2021; Das et al. 1985). To our knowledge, this is the first report of E. nigrum causing leaf spot and leaf sheath disease on maize in the Republic of Korea. This report provides important information for the diagnosis and management of maize diseases in Korea.