First Report of Root and Hypocotyl Rot of Dry Bean Caused by Aphanomyces euteiches in Alberta, Canada.

Dry bean (Phaseolus vulgaris L.) is a high-value crop grown under irrigation on 45,000 ha in southern Alberta, Canada. In 2019, one field of red beans showed premature yellowing and stunting of shoots in mid-July near Bow Island, AB. Roots had brown lesions along the length of hypocotyl and tap root, and decay and sloughing of the root cortex of lateral roots (Fig. 1). Roots were washed for 10 min under running tap water, and portions of the lesions were excised for DNA extractions using the Plant DNeasy kit (Qiagen) and multiplex PCR assays for an array of pathogens, which included Aphanomyces euteiches (Chatterton et al. 2019). Lesion pieces were also surface sterilized and plated onto PDA or cornmeal agar amended with metalaxyl, benomyl and vancomycin (CMBV) and visualized microscopically for oospores. The roots were PCR positive for A. euteiches, Rhizoctonia solani, and Pythium ultimum. Isolations yielded R. solani, and Fusarium spp. on PDA, but cultures on CMBV were over-grown with Rhizopus. Oospores measuring 24.7 +/- 1.08 m, consistent with the expected size of A. euteiches oospores of 18 - 25 m (Papavizas and Ayers, 1974), were visible in some of the root pieces. To obtain an A. euteiches isolate, soil was collected from three areas of the red bean field and used in a bait assay. Tests were performed using pea (cv. CDC Meadow) and dry bean (cv. Redbond) seeds treated with metalaxyl. Five seeds per pot were planted into field soils in 10-cm pots with 4 replicate pots/field. Soils were watered as needed until the 2nd node stage and then kept at saturation for 14 days, at which time roots were washed. Roots from the dry bean, but not the pea, plants showed severe root rot, including honey-brown discolouration of the lateral roots and extending to the hypocotyl, degradation of the root cortex, and presence of oospores. Diseased root pieces were plated onto CMBV without surface sterilization. Cultures with fast growing, white, aerial mycelia characteristic of A. euteiches on CMBV were recovered from dry bean roots from the three soil samples, considered as three isolates, and were transferred to PDATo confirm identity, total DNA was extracted from 7-day old cultures of the three isolates growing on PDA using the Qiagen DNeasy Plant Kit. The ribosomal DNA internal transcribed spacer (ITS) region was amplified using the primer pair ITS1 and ITS4, and sequenced (White et al. 1990). The sequences, deposited in GenBank with accession numbers OM976770, OM976771, and OM976772, were 100% identical to the ITS rDNA sequence of several isolates of A. euteiches (e.g. KM486066.1; 669/669 bp; 670/670 bp; 670/670 bp) using a BLASTn query and 99.7% identical to the CBS15773 voucher specimen (HQ643116.1, 661/663 bp; 662/664 bp; 662/664 bp). To confirm pathogenicity of these three isolates, cultures were used to produce zoospores for inoculating 14-day old red bean and pea seedlings (Chatterton et al. 2015). Control plants received minimal salt media solution and did not develop symptoms. Dry bean, but not pea, seedlings displayed root rot symptoms (Fig. 2). A. euteiches was re-isolated from dry bean seedlings by plating onto CMBV, and identity confirmed as described above. Although a previous survey detected A. euteiches in a small number of dry bean fields in southern Alberta (Chatterton et al. 2019), tests to confirm Koch's postulates were not performed. Aphanomyces root rot can be a devastating pathogen of pulse crops. Precautionary measures should be taken to prevent spread of this pathogen in the small dry bean growing area of southern Alberta by encouraging producers to use rotation intervals, monitoring movement of the pathogen, and evaluating cultivars for resistance to this pathogen.