First Report of Pilidium concavum Causing Tan-Brown Leaf Spot on Strawberry in China.

Abstract

In April 2011, a survey of diseases was conducted on strawberry plants grown in greenhouses in the town of Xingshou, Changping District, Beijing, China. A tan-brown leaf spot with the presence of pink spore masses was observed on older leaves of strawberry plants. In general, the leaf spots began as small, round, water-soaked lesions in the middle or on the margin of leaves, which enlarged gradually up to 1 to 3 cm in diameter and were circular or irregular and brown to dark brown. Occasionally, the center of some spots cracked in the middle lesion under dry conditions. Eventually, black sporodochia were produced on the upper surface of spots and exuded pink conidial masses under humid conditions. Fungal structures were taken directly from the diseased leaves and examined microscopically for morphological characteristics. Sporodochia, 172 to 451 × 138 to 343 μm, were dark and suborbicular. Conidiophores, 8.4 to 48.3 × 0.8 to 2.1 μm, were hyaline, unicellular, and cylindrical. Conidia, 3.1 to 10.2 × 1.5 to 3.0 μm, were hyaline, aseptate, and canoe-shaped to allantoid, forming singly. For further study, the fungus was isolated in pure culture on potato dextrose agar (PDA) medium from symptomatic leaf tissue. In culture, the mycelium was white at first and then changed to brownish. The sporodochia were light colored at first and turned brownish or almost black in older cultures. To identify the fungus, the ITS1-5.8S-ITS2 rDNA region of the isolate was amplified by PCR with primers ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') and sequenced. The 462-nt sequence (GenBank Accession No. JQ995228) was identical to that of Pilidium concavum (1). To validate Koch's postulates, pathogenicity was tested by inoculating 20 leaves on 10 healthy strawberry plants with a mycelial plug from a 15-day-old colony (0.5 cm in diameter). Controls were treated with plugs of PDA medium. The inoculated and control plants were then maintained in growth chambers at 25°C (12 h of light per day, 80% humidity). After 12 days, 100% of the inoculated leaves showed symptoms identical to those observed on leaves in the field while the control leaves remained healthy. The original fungus was reisolated from inoculated leaves showing the symptoms. Thus, it was concluded that the fungus was the causal organism of the leaf spot. On the basis of morphological characteristics, molecular features, and pathogenicity tests, the pathogen of tan-brown leaf spot on strawberry was identified as P. concavum. To our knowledge, this is the first report of P. concavum causing tan-brown leaf spot on strawberry in China. This fungus was reported to cause leaf spot on Paeonia suffruticosa in China (2). It also caused tan-brown rot on strawberry fruit in our pathogenicity test.