First report of sclerotinia stem rot of Ranunculus asiaticus caused by Sclerotinia sclerotiorum in Korea

Ranunculus asiaticus L. (Persian buttercup, Ranunculaceae) is an herbaceous perennial popularly cultivated in gardens and used as a florist crop worldwide. During winter of 2012-2013, typical symptoms of Sclerotinia stem rot were observed in commercial crops of R. asiaticus ‘Elegance’ grown in ground beds in plastic greenhouses in Hwaseong (37°12'16.8″ N; 127°00'31.8″ E) and Icheon (37°09'46.4″ N; 127°25'49.2″ E), Korea. About 20% of plants withered or died before harvest due to the disease, causing considerable economic losses. Initial symptoms were water-soaked lesions on stems at the soil line. Dark brown to black stem lesions enlarged and a cottony mycelium covered the affected area, followed by crown rot and wilt a few days afterward. Whitish aggregates of mycelia developed into sclerotia that were 2 to 8 mm in diameter outside and inside chlorotic stems. Voucher specimens were deposited in the Korea University Herbarium (KUS-F26436 and F27266). Two representative isolates were deposited in the Korean Agricultural Culture Collection (KACC46833 and 47217). Stem tissues were surface-disinfested with 0.5% sodium hypochlorite, and segments transferred to potato dextrose agar (PDA). Resultant colonies were white or faint gray and floccose, with black sclerotia (2 to 8 mm in diameter) forming on the colony surface near the margin. Based on the morphological and cultural characteristics, the fungus was identified as Sclerotinia sclerotiorum (Lib.) de Bary (Mordue and Holliday 1976). Fungal DNA was extracted with a DNeasy Plant Mini Kit (Qiagen Inc., Valencia, CA). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 (White et al. 1990) and sequenced. The resulting sequence of 501 bp was deposited in GenBank (Accession No. KP635275). A BLAST search revealed that sequences of the Korean isolates showed 100% identity with those of S. sclerotiorum (e.g., KJ696536). A pathogenicity test was achieved by placing PDA plugs (9 mm²) from a 7-day-old culture on the stems of three healthy plants near the soil line. Three plants inoculated with agar blocks served as controls. Plants were maintained in a greenhouse at 16 to 20°C and relative humidity >90%. After 3 days, all inoculated stems became discolored, soft, watery, and covered with white mycelia, whereas control plants remained symptomless. S. sclerotiorum was consistently reisolated from the symptomatic tissue, fulfilling Koch’s postulates. Sclerotinia stem rot of R. asiaticus caused by S. sclerotiorum has been recorded from Argentina, Italy, and New Zealand. Other Ranunculus species associated with this fungus were recorded in Norway and China; Ranunculus (species not identified) was listed as a host in the United States (Farr and Rossman 2015). To our knowledge, this is the first occurrence of Sclerotinia stem rot on R. asiaticus in Korea. Our observations suggest that low temperature, high humidity, poor ventilation, and continuous cultivation in nonheated plastic greenhouse cultivation systems can increase the incidence and severity of Sclerotinia stem rot on Ranunculus plants.