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First Report of Leaf Spot of Lotus (Nelumbo nucifera) Caused by Nigrospora oryzae in China

Zhang, Q. H., Huang, L. L., Liu, Y. J., Ai, Y., Peng, D. H.
Plant disease 2018 v.102 no.5 pp. 1038
DNA primers, Khuskia oryzae, Lotus, Nelumbo nucifera, agar, aquatic plants, conidia, cultivars, culture media, ethanol, flowers, fungal diseases of plants, hyphae, industry, internal transcribed spacers, leaf spot, leaves, mycelium, pathogenicity, plant pathogenic fungi, ponds, rhizomes, ribosomal DNA, sodium hypochlorite, sporulation, streptomycin, tetracycline, vegetables, China, India
Lotus (Nelumbo nucifera) is an important aquatic plant in China. It is native to India and widely planted as an ornamental and a vegetable plant because of its beautiful flowers and edible rhizomes. In May 2016, distinct leaf spots were observed on lotus (cultivar Taikonglian No. 36) in Fuzhou City, Fujian province, casing approximately 40% of leaves to die in a 0.75 ha area. The initial foliar symptoms were small irregular grayish white spots (1 to 3 mm) that had black-brown banding patterns on the edges. The lesions later expanded (1 to 2 cm), turned brown, and were surrounded by a pale, yellow halo. Ten small pieces (5 mm²) of foliar tissue were excised from the junction of diseased and healthy tissue, surface-sterilized in 70% ethanol for 1 min and 3% sodium hypochlorite for 3 min, washed in three changes of sterile distilled water, and transferred to two potato dextrose agar (PDA) plates (5 pieces/plate) amended with streptomycin and tetracycline. Petri dishes were incubated at 25°C. Hyphal tips from the leading edge of 3-day-old colonies were transferred to fresh PDA plates to obtain pure cultures. Fungal colonies were initially white and then turned gray to black upon sporulation, with fast-growing aerial mycelium becoming darker on the underside as the entire colony darkened after 15 days at 25°C. Conidia were single-celled, brown or black, smooth, spherical or subspherical, and 10.5 to 16 μm (averaging 13.6 μm) in diameter. The fungus was identified as Nigrospora oryzae based on previous descriptions (Ellis 1971; Hudson 1963). Genomic DNA was extracted from a representative isolate, 7-1, and the internal transcribed spacer (ITS) region of the ribosomal DNA was amplified using the universal primers ITS1 and ITS4 (White et al. 1990). The ITS sequence (GenBank no. MG255311) showed 98% similarity in the overlapping 508-bp portion with N. oryzae (GenBank no. KC771498). Pathogenicity tests were performed in the laboratory with the isolate on newly matured leaves of a healthy lotus cultivar Taikonglian No. 36 by inoculating with fungal plugs. Plugs, 5 mm in diameter, from 5-day-old PDA cultures were directly placed onto leaves. Leaves with sterile agar plugs served as controls. Magenta boxes containing sterile distilled water and treated leaves were loosely lidded and incubated at 25°C. The inoculated leaves started showing disease symptoms after 3 days, but no symptoms were seen on mock-inoculated leaves. Koch’s postulates were fulfilled by reisolation of N. oryzae from diseased leaves. The pathogenicity tests were carried out twice with the same results. Review of the literature revealed that N. oryzae is known as a pathogen on many plants (Ellis 1971; Zheng et al. 2012) but has not been reported on lotus. To our knowledge, this is the first report of N. oryzae infecting lotus in China or worldwide. The leaf spot was found in four surveyed lotus ponds in Fuzhou and poses a threat to the lotus industry. The management of the disease should be further investigated.