Decreased sensitivity of Leptosphaeria maculans to pyraclostrobin in Alberta, Canada

Leptosphaeria maculans, the causal agent of blackleg of canola (Brassica napus), can be managed with pyraclostrobin and other strobilurin fungicides. Their frequent application, however, poses a risk for the development of insensitivity in fungal populations. A collection of L. maculans single-spore isolates recovered from infected canola stubble in Alberta, Canada, in 2016 was evaluated for its pyraclostrobin sensitivity. In conventional growth plate assays, the concentration of pyraclostrobin required to inhibit fungal growth by 50% (EC50) was determined to be 0.28 mg/liter in a subset of 38 isolates. This EC50 was four times greater than the mean EC50 (0.07 mg/liter) of baseline isolates collected in 2011. Two hundred sixty-three isolates were screened further with two discriminatory doses of 0.28 and 3.5 mg/liter of pyraclostrobin, resulting in growth inhibition values ranging from 16 to 82% and 41 to 100%, respectively. In microtiter plate assays with the same isolates, the mean EC50 was determined to be 0.0049 mg/liter, almost 3.3 times greater than the mean EC50 (0.0015 mg/liter) of the baseline isolates. The sensitivity of the isolates was also evaluated in microtiter plate assays with discriminatory doses of 0.006 and 0.075 mg/liter of pyraclostrobin, resulting in inhibition values ranging from 20 to 88% and 49 to 100%, respectively. This is the first report of isolates of L. maculans with increased insensitivity to pyraclostrobin in Canada, suggesting the need for improved fungicide stewardship.