First Report of Nigrospora oryzae Causing Leaf Spot of Cotton in China.

Cotton (Gossypium hirsutum L.) is widely cultivated for the important economic value of the fiber. In the summer of 2011, a leaf spot of cotton plants cv. Wanza40 was observed in 11 fields (total of about 4 ha) in Qianshan County in southwest Anhui Province, China. Approximately 30% of the plants in each field were symptomatic. Affected plants exhibited brown to reddish, irregular foliar lesions, each with a brown border near the vein of the leaves. A sign of fungal infection was a dark leaf mold observed on lesions on the abaxial surface of leaves. Sections of symptomatic leaf tissues were surface-sterilized (in 75% ethanol for 30 s, then 1% NaOCl for 1 min), rinsed three times in sterile distilled water, and plated onto potato dextrose agar (PDA). A fungus consistently recovered from symptomatic leaf samples produced colonies that were initially white and then became grayish brown with the onset of sporulation. Black, spherical to subspherical, single-celled conidia (10 to 12 × 14 to 16 μm) were borne on a hyaline vesicle at the tip of each conidiophore. Morphological characteristics of the fungus were similar to that of Nigrospora oryzae (2). The internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) from a representative strain of the fungus, AHC-1, was amplified using the primers ITS1/ITS4 (4) and sequenced (GenBank Accession No. JQ864579). The ITS sequence had 99% identity with >553 bp of the ITS sequence of an N. oryzae isolate (GenBank Accession No. EU918714.1). On the basis of morphological data and ITS rDNA sequence, the isolate was determined to be N. oryzae. A pathogenicity test was performed on detached, young leaves of 4-month-old healthy cotton plants of cv. Wanza40. Six leaves were inoculated by placing a colonized agar piece (5 mm in diameter) from 7-day-old cultures of the fungus on pushpin-wounded leaves. Another six leaves treated with sterile PDA plugs served as a negative control treatment. Leaves were incubated in petri dishes and maintained at 25°C in a growth chamber programmed for 12 hours of fluorescent white light/day. After 5 days, brown to black lesions were observed on all inoculated leaves, whereas no symptoms developed on control leaves. N. oryzae was consistently reisolated from symptomatic leaves but not from the control leaves. N. oryzae is a weak pathogen on a wide range of plants, and has been described as the causal agent of lint rot on cotton (1,3), but to our knowledge this is the first report of N. oryzae causing a leaf spot of cotton in China. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, ARS, USDA, Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , April 8, 2012. (2) H. J. Hudson. Trans. Br. Mycol. Soc. 46:355, 1963. (3) A. J. Palmatter et al. Plant Dis. 87:873, 2003. (4) T. J. White et al. In: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.