Microtiter plate test using liquid medium is an alternative method for monitoring metyltetraprole sensitivity in Cercospora beticola.

BACKGROUND Metyltetraprole is a new quinone outside inhibitor (QoI) fungicide that shows potent activity against QoI-resistant fungi that possess the G143A cytochrome b mutation, which confers resistance to existing QoIs such as trifloxystrobin. For its sustainable use, monitoring of metyltetraprole sensitivity is necessary and the establishment of appropriate methodology is important in each pathogen species. RESULTS In Cercospora beticola, the causal agent of sugar beet leaf spot, some isolates were less sensitive to metyltetraprole (EC50 > 1 mg L-1, higher than the saturated concentration) using the common agar plate method, even with 100 mg L-1 salicylhydroxamic acid, an alternative oxidase inhibitor. However, microtiter tests (EC50  80% control at 75 mg L-1 run-off spraying) confirmed that all tested isolates were highly sensitive to metyltetraprole. For trifloxystrobin, G143A mutants were clearly resistant upon microtiter plate tests (median EC50 > 2 mg L-1 ) and distinct from wild type isolates (median EC50 < 0.01 mg L-1 ). Notably, mycelium fragments were usable for the microtiter plate tests and the test was applicable for isolates that do not form sufficient conidia. Our monitoring study by microtiter plate tests did not indicate the presence of metyltetraprole-resistant C. beticola isolates in Hokkaido populations in Japan. CONCLUSION The microtiter tests were revealed to be useful for monitoring the sensitivity of C. beticola to metyltetraprole and trifloxystrobin. This article is protected by copyright. All rights reserved.