First Report of Leptosphaeria maculans and Leptosphaeria biglobosa, Causal Agents of Blackleg, on Canola in Washington State

Blackleg, caused by Leptosphaeria maculans (Desm.) Ces. & De Not., is a seed-borne pathogen endemic in the Canadian prairies, Midwestern and southern United States, Australia, and Europe (Rimmer et al. 2007). The canola production region of the Pacific Northwest United States was considered free of blackleg until 2011, when L. maculans was found in canola fields in northern Idaho (Agostini et al. 2013). An outbreak of blackleg occurred in the Willamette Valley, OR, in 2014-15 in diverse brassica crops and weeds (Ocamb et al. 2015). In spring 2015, blackleg was confirmed in >18 winter rapeseed and canola crops in Camas Prairie, ID (J. Davis and L. J. du Toit, personal observation). In eastern Washington, where 12,000 ha of dryland canola are grown, winter and spring canola fields (n = 25) were surveyed in 2015, including residues from 2014-15 crops. Lesions on winter canola leaves collected in April from a field in Okanogan Co. had pycnidia typical of the asexual state. Pycnidia were also found on canola residues. Single-conidia isolations onto water agar from cirrhi were transferred to PDA. The ITS1, 5.8s ribosomal RNA gene, and ITS2 sequences (Schroeder et al. 2006) of isolates Phl048 and Phl049 from Whitman Co. (GenBank accession nos. KX037027 and KX037028) had 99% homology with that of L. biglobosa Shoemaker & H. Brun ICMP:13281 (KT309866.1), and the sequence of isolate Phl050 from Okanogan Co. (KX037029) was 99% homologous to that of L. maculans ATCC 46318 (JX499035.1). Beta-tubulin gene sequence homology and phylogeny identified Phl048 and Phl049 as L. biglobosa subsp. australiensis (Vincenot et al. 2008). Isolates Phl048 to Phl051 were each tested for pathogenicity on six cabbage plants (14 days old, cv. Copenhagen Market) and six canola plants (cv. Westar). Each hypocotyl was punctured with a needle, and 10 µl of conidial suspension (106 CFU/ml) applied to the wound. Six plants of each species were inoculated with isolate Phl031 of L. maculans as a positive control treatment; and water was applied to six plants per species as a negative control treatment. Plants were grown at 20°C on a 12 h/12 h day/night cycle, and observed 10 and 14 days after inoculation. L. maculans isolates Phl050 and Phl051 caused a necrotic lesion, wilt, and seedling collapse, and formed pycnidia in the lesion. L. biglobosa isolates Phl048 and Phl049 caused a superficial lesion with limited or no pycnidia. The isolates caused similar symptoms on canola as on cabbage plants except for slower symptom expression and fewer pycnidia. Reisolates from plants inoculated with Phl048 and Phl049 were confirmed by morphology and/or sequencing as L. biglobosa; and reisolates from plants inoculated with Phl050, Phl051, and Phl031 were confirmed as L. maculans. Fungi were not recovered from negative control plants. Repeat tests gave the same results. The hot, dry summer of 2015 limited disease development. The results indicate L. maculans and L. biglobosa occur in inland Washington at a limited incidence. Washington quarantine regulations require testing brassica seed for the pathogens before planting to prevent introducing the fungi, especially in northwest counties and the Columbia Basin where a significant amount of brassica seed is produced for the United States and for export. © The American Phytopathological Society.