Articles
Comparing methods for detection of QoI fungicide resistance in Colletotrichum spp. associated with postharvest disease of avocado in Australia
Article number
1451_12
Pages
89 – 94
Language
English
Abstract
This study compared azoxystrobin-amended agar plate (Petri dish) and microplate assays for the detection of strobilurin fungicide resistance in Colletotrichum species.
Seven Colletotrichum isolates which were classified as resistant (RES) or sensitive (SEN) to strobilurin fungicides based on a molecular detection of the G143A mutation in the cytochrome b gene region, were used for this study.
The isolates were grown in media amended with azoxystrobin fungicide (10, 1, 0.1, 0.01 and 0 mg a.i. mL‑1 for the Petri dish assay and 0.1, 0.01, 0.001, 0.0001 and 0 mg a.i. mL‑1 for the microplate assay), and the concentration inhibiting growth by 50% (EC50) was calculated.
A significant positive correlation was observed between fungal growth measured via the Petri dish and microplate assays (R2=0.4223, p≤0.01). The Petri dish assay generally produced higher EC50 values, except for one isolate.
For both assays, all RES isolates had EC50≥9.7 mg a.i. mL‑1. In the Petri dish assay, the EC50s for SEN isolates were lower than the RES isolates by more than 10-fold for most isolates.
The EC50s for SEN isolates in the microplate assay, were lower by 100- to 1000-fold than those of the RES isolates.
Despite the variations in EC50 values, results from both the Petri dish and the microplate assays were consistent with those obtained from the molecular assay.
The microplate assay was more sensitive in detecting growth inhibition and multiple isolates could be screened for resistance simultaneously demonstrating how the 96-well microplate method could be utilised for monitoring fungicide resistance in Colletotrichum spp.
Seven Colletotrichum isolates which were classified as resistant (RES) or sensitive (SEN) to strobilurin fungicides based on a molecular detection of the G143A mutation in the cytochrome b gene region, were used for this study.
The isolates were grown in media amended with azoxystrobin fungicide (10, 1, 0.1, 0.01 and 0 mg a.i. mL‑1 for the Petri dish assay and 0.1, 0.01, 0.001, 0.0001 and 0 mg a.i. mL‑1 for the microplate assay), and the concentration inhibiting growth by 50% (EC50) was calculated.
A significant positive correlation was observed between fungal growth measured via the Petri dish and microplate assays (R2=0.4223, p≤0.01). The Petri dish assay generally produced higher EC50 values, except for one isolate.
For both assays, all RES isolates had EC50≥9.7 mg a.i. mL‑1. In the Petri dish assay, the EC50s for SEN isolates were lower than the RES isolates by more than 10-fold for most isolates.
The EC50s for SEN isolates in the microplate assay, were lower by 100- to 1000-fold than those of the RES isolates.
Despite the variations in EC50 values, results from both the Petri dish and the microplate assays were consistent with those obtained from the molecular assay.
The microplate assay was more sensitive in detecting growth inhibition and multiple isolates could be screened for resistance simultaneously demonstrating how the 96-well microplate method could be utilised for monitoring fungicide resistance in Colletotrichum spp.
Authors
I. Nokdy, L.S. Pretorius, J.E. Thomas, E.K. Dann
Keywords
microplate assay, in vitro methods, fungicide resistance, strobilurin, azoxystrobin, Colletotrichum, anthracnose
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