First Report of <i>Nigrospora oryzae</i> Causing Leaf Spot on Peanut (<i>Arachis</i> <i>hypogaea</i>) in the Republic of Korea

Peanut (<i>Arachis hypogaea</i> L.) is one of the most profitable upland crops, yielding 10,711 tonnes in an area of 4,062 ha in the Republic of Korea (Ministry of Agriculture, Food and Rural Affairs 2023). In September 2023, dark gray spots surrounded by yellow halos were observed on the peanut leaves over an area of 880 m² at the National Institute of Crop Science (35°50'31.4"N 127°02'41.0"E), with a disease incidence up to 80%. Early symptoms appeared as small, brown, circular or irregular spots that enlarged and were surrounded by chlorotic halos. Leaf cuttings (5 mm x 5 mm) from five symptomatic plants were surface-sterilized with 70% EtOH for 1 min, followed by 1% NaClO for 1 min, and rinsed 3 times with sterile water. The pieces were placed on Potato Dextrose Agar (PDA) and incubated at 25 °C in the dark for 3 days. Three isolates obtained by single-spore isolation were designated as F23025, F23026, and F23027. Two isolates, F23025 and F23026 were deposited in the Korean Agricultural Culture Collection (https://genebank.rda.go.kr) under the accession numbers 410722 and 410723. Fungal colonies were initially white and turned sooty gray after 5 days. Conidia were unicellular, brown to black, and spherical or sub-spherical with 6.8 μm to 14.3 μm (mean = 11.1 μm ± 1.8, n = 50). The morphology of the three isolates was identical and showed the same characteristics as <i>Nigrospora oryzae</i> (Ellis 1971; Hudson 1963). For molecular identification, the Internal Transcribed Spacer (ITS) region (GenBank accession PP388306 and PP574448), beta tubulin (PP397027 and PP580108), and translation elongation factor 1- ɑ (PP397028 and PP580109) of isolates F23025 and F23026 were amplified and sequenced with primers of ITS5/ITS4, Bt2a/Bt2b, EF1-727F/EF2, respectively and showed high identity of 99.62% (530 bp/532 bp), 100% (384/384), and 99.79% (475/476) with N. oryzae strain LC2693 (GenBank accessions KX985994, KY019471, and KY019299, respectively). Multilocus sequence analysis showed isolates F23025 and F23026 were on the same clade with <i>N. oryzae</i> strain LC2693. To determine the pathogenicity to peanut, a conidial suspension (1 x 10⁶ conidia/mL) was sprayed onto leaves of five 3-week-old plants 'Sewon' grown in pots, while sterile distilled water was sprayed onto two plants used as negative control. Sprayed plants were placed in a dew chamber at 25℃ for two days and grown in a growth chamber at 25℃ and 80% of relative humidity with a 16L:8D cycle. Two weeks later, dark spots with chlorotic halos appeared only on leaves sprayed with conidia, and no symptoms on leaves sprayed with sterile distilled water. The pathogenicity test was repeated three times, and each time the pathogen was re-isolated and identified by ITS sequence, thus fulfilling Koch's postulates. <i>Nigrospora</i> species are cosmopolitan, and some species have a wide host range as plant pathogens. Recently, two species of the genus Nigrospora, <i>N. sphaerica</i> and <i>N. aurantiaca</i>, were reported to cause peanut leaf blight in China (Liu et al. 2020; He et al. 2023). To the best of our knowledge, this is the first report of <i>N. oryzae</i> causing leaf spot to <i>A. hypogaea</i> L. in the Republic of Korea. As identifying new pathogens and registering fungicides to control them are important for the continued cultivation of peanut, this report will help in that endeavor.