First Report of Magnaporthe poae, Cause of Summer Patch Disease on Annual Bluegrass, in Canada.

The ectotrophic, root-infecting fungus Magnaporthe poae Landschoot & Jackson, the causal agent of summer patch disease in the U.S. (2), is implicated in the damage and loss of annual bluegrass (Poa annua L.) on golf course greens. This pathogenic fungus, one of the important root pathogens of turfgrass, attacks and colonizes susceptible turfgrass roots suffering from environmental or cultural stresses. Over 100 turf samples that exhibited symptoms (chlorotic circular or irregular patches of ≥15 cm in diameter with necrotic crowns and discolored roots) reminiscent of summer patch were collected from 77 southwestern Ontario golf courses from July to August of 2009 and 2010. Roots and crowns were often covered with dark, ectotrophic runner hyphae, lobed hyphopodia, and growth cessation structures, characteristic of M. poae. Sections of root tissue were surface sterilized in 0.6% sodium hypochlorite (NaOCl) for 5 min. Sterilized root tissue was plated on potato dextrose agar (PDA) containing 50 mg L streptomycin sulfate and incubated at 28°C for 7 to 10 days. A fungus with morphological characteristics (hyaline mycelium that appears gray or olive-brown when mature) similar to those of M. poae (1) was consistently isolated (≥100 isolates were obtained) and used to identify M. poae through molecular techniques and Koch's postulates. DNA was extracted from the fungal mycelium of the collected isolates using the PowerPlant DNA isolation kit (MO BIO Laboratories, Inc., Carlsbad, CA). The rDNA internal transcribed spacer (ITS) regions of the isolates (≥100 isolates) were amplified by PCR using universal fungal rDNA primers ITS 4 (5'-TCCTCCGCTTATTGATATGC-3') and ITS 5 (5'- GGAAGTAAAAGTCGTAACAAGG-3') (3). The purified PCR products were sequenced (GenBank Accession No. JX134588 through JX134601) and a BLAST search exhibited seven isolates with 99% (MAG3, MAG6, MAG13, MAG16, and MAG17) and 100% (MAG1 and MAG14) similarity to M. poae in the GenBank database. Pathogenicity of four isolates (MAG1, MAG3, MAG6, and MAG14) was confirmed with Koch's postulates. Sixteen healthy P. annua core samples (four replicates of each treatment/isolate) collected from an Ontario golf course were inoculated with 25 mg M. poae-infested Kentucky bluegrass seed (Poa pratensis L.; 12.5 mg inoculum applied at the surface of the potting medium and 12.5 mg inoculum applied on the foliar surface) and were placed in a growth chamber with 12-h day/night cycles at 30/25°C and approximate relative humidity. After 2 to 3 weeks, inoculated plants exhibited chlorotic foliage and necrotic roots covered with dark ectotrophic runner hyphae and lobed hyphopodia. Infected root sections from each replication were surface sterilized and placed on PDA containing 50 mg L streptomycin sulfate. The fungal cultures exhibited morphological characteristics consistent with M. poae (1). To our knowledge, this is the first report of summer patch caused by M. poae in Canada. References: (1) B. B. Clarke and A. B. Gould, eds. Turfgrass Patch Diseases Caused by Ectotrophic Root-Infecting Fungi. The American Phytopathological Society, St. Paul, MN, 1993. (2) P. J. Landschoot and N. Jackson. Mycol. Res. 93:59, 1989. (3) T. J. White et al. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Pages 315-322 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al. eds. Academic Press, San Diego, CA, 1990.