First Report of Brown Ring Patch Caused by Waitea circinata on Perennial Ryegrass ( Lolium perenne ) in Oklahoma

Brown ring patch of cool-season turfgrass, caused by Waitea circinata Warcup & Talbot, was first reported in Japan on creeping bentgrass (Agrostis palustris Huds.) in 2005 (Toda et al. 2005) and in the United States on annual bluegrass (Poa annua L.) in 2007 (de la Cerda et al. 2007). In April 2015, a rapid decline was observed on a stand of perennial ryegrass (Lolium perenne cv. Soprano, 1G Squared, and Apr2116) that had been overseeded on a bermudagrass stand at a baseball field in Stillwater, OK. Symptomatic leaf tissue was surface sterilized for 1 min in a 0.8% sodium hypochlorite solution, rinsed with sterile water, and plated onto potato dextrose agar (PDA). A Rhizoctonia-like fungus was consistently isolated and suspected to be the causal pathogen. After 3 days at 25[degrees]C white, cottony growth formed in culture, which turned orange to brown as the culture aged. Spherical sclerotia of irregular diameters were present in the 7-day-old culture that followed the same color development as mycelia. Colony and sclerotia morphology were consistent with previous descriptions of W. circinata. (de la Cerda et al. 2007; Toda et al. 2005). DNA was extracted and the internal transcribed spacer region (ITS) of the ribosomal RNA gene was amplified using primer pair ITS4/ITS5 and sequenced (White et al. 1990). Amplified fragments matched with 100% similarity to published ITS sequences (GenBank Accession Nos. HM807352, HQ166071, and FJ55889). A pathogenicity test was conducted on 3-week-old perennial ryegrass seedlings of the same cultivar blend on which the disease was initially observed. This blend was grown in 10-cm-diameter pots containing commercial planting mix (Sun Gro Horticulture, Bellevue, WA). One 4-mm-diameter plug was removed from the margin of a 4-day-old colony growing on PDA and placed on wet leaves. Five replicate pots of each inoculated and noninoculated ryegrass seedlings were kept in a moist chamber at 22[degrees]C. Within 4 to 5 days, inoculated leaves exhibited slight chlorosis, and water soaked lesions with white aerial mycelium were present. Koch's postulates were fulfilled as mycelia were recovered from diseased lesions, and confirmed by amplification and sequencing of the same ITS region. This demonstrates that W. circinata is present in Oklahoma and in the southern Midwest of the United States. To our knowledge this is the first report of W. circinata infecting perennial ryegrass in this region. Given that it has not been reported in t