In 2009, a disease survey was conducted in 97 commercial canola (Brassica napus L.) fields in Western Australia by the Department of Agriculture and Food, Western Australia (DAFWA). In about 20% of the fields from the northern agricultural region of Western Australia, small patches were observed where canola plants showed symptoms of stunting and wilting. These plants were collected and roots of affected plants were washed thoroughly and examined for the presence of root disease. Small galls and clublike structures were observed on the secondary roots and sometimes on the main root of the affected plants. Examination of thin free hand sections from the root galls revealed that several cortical cells were enlarged and full of resting spores. The diameter of resting spores ranged between 2.5 and 3.0 μm. Plasmodia and zoosporangia were also observed in the root hairs. The identity of Plasmodiophora brassicae Woronin was confirmed by PCR using a modified method of Cao et al. 2007 (1). DNA from spores and slices of the galls of 14 different samples were extracted using DNeasy plant mini kit (QIAGEN Australia) as per manufacturer's instructions. Samples were disrupted by placing them into MPBIO tube A and placed in the Fast Prep machine at speed of 6 ms–1 for 40 s. This was repeated twice. The species-specific primers TC1F (5′-GTGGTCGAACTTCATTAAATTTGGGCTCTT-3′)/TC1R (5′-TTCACCTACGGAACGTATATGTGCATGTGA-3′) and TC2F (5′-AAACAACGAGTCAGCTTGAATGCTAGTGTG-3′)/TC2R (5′-CTTTAGTTGTGTTTCGGCTAGGATGGTTCG-3′) were used (1). The primers TC1F and TC1R failed to produce a PCR product of 548-bp size but using the primers TC2F and TC2R the PCR reaction resulted in a 519- bp fragment. Seven out of 14 samples gave positive results for P. brassicae with primers TC2F and TC2R. This indicates that the P. brassicae pathotype from Western Australia may be different than the one found in Alberta, Canada. However, pathotypes of P. brassicae from brassica vegetables from Australia have been found similar to the populations of P. brassicae present in the United States (2). Pathogenicity of P. brassicae was tested by dipping roots of five 10-day-old canola plants var. Cobbler in a spore suspension (1 × 106 resting spores/ml). Roots of five control plants were dipped in sterile water. Five weeks after inoculation, small galls were observed on the roots of three inoculated plants and the control plants remained symptomless. Resting spores were recovered from the galls developed on the roots of affected plants. Presence of P. brassicae in the affected roots was further confirmed by PCR using the method described above. To our knowledge, this is the first report of club root of canola in Australia. Club root is reported from vegetable brassicas and white mustard (Sinapis alba L.) in Australia. Club root has become a serious disease of canola in Canada since its detection in Alberta in 2006 (3). The resting spores of the fungus can survive for several years in soil, and therefore, this disease could pose a significant threat to canola production in Western Australia. References: (1) Cao et al. Plant Dis. 91:80, 2007. (2) Donald et al. Ann. App. Biol. 148:239, 2006. (3) S. Streklov et al. Can. J. Plant Pathol. 28:467, 2006.
Experiments on obtaining precipitating sera by the Fajiwara method
