APPLICATION OF AN AUTOMATED QUANTITATIVE METHOD TO DETERMINE FUNGICIDE RESISTANCE IN BOTRYTIS-CINEREA

An automated quantitative (AQ) assay used for measuring fungal growth with a microplate reader was compared with the linear growth method to determine fungicide resistance in Botrytis cinerea. The AQ assay uses absorbance in the range of 0.0-0.6 units as a measure of fungal bioma ss. This technique was successfully used to establish EC(50) values (t he concentration of fungicide that reduces absorbance by half) to ipro dione and a mixture (1:5) of carbendazim and diethofencarb in an econo mical and rapid way. The AQ assay used 100 times less medium than did the conventional method of measuring radial growth (RG) of mycelium on fungicide-amended medium, and up to 96 samples (one microplate) could be processed at once. The assay was also performed with conidia store d at -20 C in a 20% glycerol solution for 4 mo, and EC(50) values did not differ significantly from EC(50) values using fresh conidia. Growt h inhibition was measured most accurately when the spore concentration in a well of the microplate was 10 or 20 spores per microliter. EC(50 ) values determined by the AQ assay were compared with EC(50) values o btained by the RG method, and both were positively correlated. Regress ion lines to predict EC(50) values by the RG method from AQ values wer e: logY = -0.392 + 1.04 logX for iprodione, and logY 0.742 + 1.38 logX for the mixture of carbendazim and diethofencarb where Y = EC(50) val ue determined by the RG method and X = EC(50) value determined by the AQ assay.