First Report of Basal Stem Rot and Foliar Blight Caused by Pythium sylvaticum on Miscanthus sinensis in Illinois.

Miscanthus sinensis Anderss., a perennial grass, is native to eastern Asia. It has been widely grown as an ornamental in temperate regions of the world, including the United States, and recently has become an important component of public and private sector bioenergy feedstock Miscanthus selection programs. In August 2008, stem rot and blight was observed on M. sinensis plants in two irregular patches, ~2 to 2.5 × 1 to 1.5 m each in a trial plot that was preceded by corn, at the University of Illinois Energy Farm near Urbana, IL. At the time of the observation, most plants were dead and the wilted tillers had black, soft rotted basal stems. A few plants were stunted and the crowns of the tillers had black-to-brown soft rot. Some tillers' leaves were dead and others had turned light brown. Sample tissue fragments were surface disinfested in 0.5% NaOCl and plated on 1% water agar (WA). After 3 days of incubation in the dark at 23°C, colonies were transferred to corn meal agar (CMA), potato dextrose agar (PDA), or 10% V8 juice agar and incubated at 23°C under continuous white light for up to 2 weeks. Morphological characteristics of the isolates correspond to those originally described for Pythium sylvaticum W.A. Campb. & J.W. Hendrix (1). The mycelia grew and covered the 10-cm-diameter plates within 5 days. On PDA, the culture was a creamy white mycelial mat of coenocytic hyphae. The isolates produced only globose, terminal or intercalary hyphal swellings ranging from 28 to 48 μm in diameter, but no oogonia were produced on any of the three growth media. No zoospores were produced when agar blocks bearing mycelium were flooded with distilled water or 1% soil water. Sequence analysis was performed with the internal transcribed spacer (ITS) region of the rDNA amplified with primer pair ITS1/ITS4 (3) and the mitochondrially encoded cytochrome c oxydase subunit II (cox II) gene using primers FM58/FM66 (2). The resulting 871-bp ITS nucleotide sequence (Accession No. HM991706) was identical among all three isolates analyzed and 99% identical (100% coverage) to ITS sequences of multiple isolates of P. sylvaticum in GenBank. Likewise, the 544-bp cox II sequence (Accession No. HQ454429) was 99% identical (97% coverage) to cox II sequences of multiple isolates of P. sylvaticum. Six pots of M. sinensis seedlings were inoculated by placing two CMA plugs of a 2-week-old culture of isolate F71 at the crown. The control pots were mock inoculated with sterile CMA plugs. The plants were incubated at ~90% relative humidity (RH) and 25°C day and 22°C night for 3 days, and thereafter left on the greenhouse bench at ~65% RH with alternating 9 h of darkness and 15 h of light. Three weeks after inoculation, two of the inoculated seedlings wilted, others were stunted with leaves wilting from the tip downwards and the stems rotting from the crown upward. A thick mat of mycelia was seen on the rotted basal stems. No symptoms were observed in the control. P. sylvaticum was reisolated from both the rotted basal stems and the wilted foliage. To our knowledge, this is the first report of P. sylvaticum on M. sinensis. Infestation of farm soils with P. sylvaticum could limit M. sinensis biomass production significantly by limiting seedling establishment. References: (1) W. A. Campbell and F. F. Hendrix. Mycologia 59:274, 1967. (2) F. M. Martin. Mycologia 92:711, 2000. (3) T. J. White et al. Page 38 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, 1990.