First Report of Didymosphaeria rubi-ulmifolii Brown Spot infection of Chinese quince fruit in South Korea.

Abstract

Chinese quince (Pseudocydonia sinensis (Thouin) CK Schneid.), a deciduous tree in the family Rosaceae, is native to China, Japan, and South Korea; the fruit is known as mogwa in South Korea. The ripened yellow fruit has been used as a traditional therapeutic for respiratory ailments and as an additive in health products such as syrups, tea, and candies (Sawai et al. 2008). From May to August 2020, Chinese quince trees showing symptoms of brown spots were observed on the Kyungpook National University premises, Daegu, South Korea, with an incidence of 30%-40%. The disease first appeared as small, round, yellow specks on the fruits, which necrotized over time and gradually enlarged to 0.7-2.7 cm in diameter. To isolate the pathogen, symptomatic tissues obtained from disease fruit were surface sterilized for 1 min with 70% ethanol, rinsed in sterile distilled water, and plated onto potato dextrose agar (PDA). The inoculated plates were incubated at 25°C for 7 days. Successively, pure cultures were obtained by transferring hyphal tips to new PDA plates. Twenty isolates were obtained from 25 fruit. Colonies on PDA reached a diameter of 30-40 mm. After incubation for 7 days at 25°C, spreading with an even, colorless-to-buff glabrous margin, a submarginal ring of conidiomata developed from day 5 to 12 and was visible as scattered dots on either side of the plate. Conidiogenous cells were discrete (3.5-6 × 3.5-5 μm); conidia were ellipsoid to short-cylindrical [3-5 × 2.1-3.5 μm (n = 60)] and olivaceous in color. These conidial dimensions corresponded to those of Didymosphaeria rubi-ulmifolii Ariyaw., Camporesi & K.D. Hyde (basionym: Paraconiothyrium brasiliense), which has been found on Rubus ulmifolius in Italy (Ariyawansa et al. 2014). Sequence data for the rDNA internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), and partial translation elongation factor 1-α (TEF) (White et al. 1990, Rehner and Buckley 2005) were obtained for one of the pure culture isolate (BT1) with GenBank accession numbers MW020087, MW020060 and MW027220, respectively. The sequences of BT1 isolate using a BLASTn analysis showed 100% identity with the ex-type MFLUCC 14-0023 of D. rubi-ulmifolii in ITS, and LSU portions (accession nos. MT310602, and MT214555, respectively) and 99% identity in TEF portion (accession no. MT394734). The procedure for Koch's postulates was followed to confirm fungal pathogenicity using 3-day-old mycelial discs. Fifteen healthy fruit were divided into three groups of five fruit each, with each group receiving a different treatment. One group of fruit was wounded by puncturing with a sterile pin and inoculated using 5-mm agar discs with mycelium on the wounds. Mycelium covered agar discs were also placed on the surfaces of five unwounded fruits. The remaining five fruit were maintained as a control and inoculated with sterile PDA plugs. The pathogenicity test was replicated thrice. The wounded fruits showed symptoms similar to those observed in the field. The control group remained asymptomatic and the morphology of the fungus re-isolated from the inoculated fruit was the same as that of D. rubi-ulmifolii. The phylogeny, together with the morphological identification and inoculation results, confirmed the identity of the fungus as D. rubi-ulmifolii (Ariyawansa et al. 2014). To the best of our knowledge, this is the first report of D. rubi-ulmifolii causing brown spot in Chinese quince.