First Report of Brown Spot Caused by Alternaria alternata on Potato (Solanum tuberosom L.) in Korea

Abstract

In June 2020, brown spot symptoms were observed in a commercial potato field located in Yeocheon, Gyeonggi Province, Korea. The symptoms were similar to those associated with early blight. Brown lesions on leaves were circular and expanded rapidly under high humidity and warm temperatures ranging 12°C at night to 30°C during daytime. Over 60% of potato (Solanum tuberosum L. cv. Superior) leaves showed the symptoms. For fungal isolation, infected leaf tissues (5 × 5 mm) from 14 infected samples were immersed in 70% ethanol for 1 min, rinsed three times in sterilized water, dried, placed on water agar amended with 100 ppm of streptomycin, and then incubated in the dark at 25°C. Hyphae emerging from the tissues were subcultured on V8-Juice agar (8% of V8-Juice, 1.5% agar, pH 7), and the obtaining cultures were subjected to single-spore isolation, resulting in 14 isolates (SYP-934~947). Three representative isolates, SYP-934 to SYP-936, were deposited in the Korean Agriculture Culture Collection (Accession Nos. KACC 410058 to KACC 410060). Conidia (n = 100) produced on the colony were brown, ellipsoid to ovoid with walls ornamented, 1 to 6 transverse and 0-3 vertical septa, and length × width of 20-45 × 7 to 24 μm (n = 100). Their morphological characteristics were consistent with Alternaria alternata (Simmons, 2007; van der Waals et al., 2011; Woundenberg et al. 2015). Sequences of the following loci in the 14 isolates were determined as described in Woundenberg et al. (2013 and 2014: the internal transcribed spacer (primer pairs VG9/ITS4, GenBank accession nos. OP581413-25), glyceraldehyde-3-phosphate dehydrogenase (gpd1/gpd2, OP588286-99), RNA polymerase second largest subunit (RPB2-5F2/fRPB2-7cR, OP588314-27), translation elongation factor 1-alpha (EF1-728F/EF1-986R, OP588300-13), Alternaria major allergen gene (Alt-For/Alt-Rev, OP588328-41), endopolygalacturonase (PG3/PG2b, OP588342-55), and an unknown gene region (OPA10-2R/OPA10-2L, OP588356-68). A neighbor-joining phylogenetic analysis based on the concatenated gene sequences, which was performed using the MEGA X program (Kumar et al., 2018), placed the 14 isolates in the clade containing A. alternata isolates. To test pathogenicity, one-month-old potato (S. tuberosum cv. Superior) plants grown in a 25°C growth chamber were sprayed with conidial suspensions (1×106 conidia/mL) prepared from 14-day-old cultures of three isolates (KACC 410058 to KACC 410060). Sterile distilled water was used as the control treatment. The inoculated pots were placed in a plastic box to maintain high humidity and incubated in the dark at 25°C for 2 days. The plants were transferred to a growth chamber (16h light with over 70% humidity at 25°C). Symptoms were first observed after 3 days post inoculation (dpi) with all three isolates, and severe brown spot symptoms were observed after 7 dpi. No symptom was observed in the control treatment. The pathogenicity assay was repeated at triplicate. Reisolated cultures from lesions were confirmed to be A. alternata based on their sequence at the rpb2 locus, thus fulfilling Koch’s postulates. Alternaria alternata has been reported to cause brown spot and leaf blight on potato leaves in Israel (Dorby et al., 1984) and South Africa (van der Waals., et al. 2011). To our knowledge, this study is the first report of A. alternata causing brown spot disease in Korea.