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Characterization of iprodione resistance in Botrytis cinerea from strawberry and blackberry.

Phytopathology (2013-10-26)
Anja Grabke, Dolores Fernández-Ortuño, Achour Amiri, Xingpeng Li, Natália A Peres, Powell Smith, Guido Schnabel
ABSTRACT

Gray mold, caused by the fungal pathogen Botrytis cinerea, is one of the most destructive diseases of strawberry. Control of the disease in commercial fields is largely dependent on the application of fungicides, including the dicarboximide iprodione. Single-spore isolates were collected from strawberry fields in Florida, North Carolina, and South Carolina and subjected to an assay using conidial germination that distinguished sensitive (S) isolates from isolates with various levels of resistance to iprodione. Of the 245 isolates, 1 was highly resistant (HR), 5 were moderately resistant (MR), and 43 had low resistance (LR) to iprodione. LR and MR strains were found in the Florida population and in 9 of 11 locations from North Carolina and South Carolina, indicating that resistance was widespread but accounted for only a relatively small percentage of the B. cinerea population. Sequence analysis of the target gene bos1, which codes for a class III histidine kinase, revealed that the MR phenotype was associated with Q369P and N373S mutations and that the LR phenotype was associated with either a I365S or a I365N mutation. The I365S and I365N mutations were also present in five additionally included HR isolates from North Carolina and South Carolina blackberry fields and one HR isolate from a Virginia strawberry field but no mutation or mutation combinations in bos1 were uniquely associated with the HR phenotype. Expression analysis of bos1 in S and HR isolates did not reveal convincing evidence of the gene's involvement in HR resistance either. The six HR isolates had three different phenotypes with respect to their sensitivity to fludioxonil; two were S, two were LR, and two were MR. The fludioxonil LR and MR isolates were also resistant to tolnaftate, an indication of multidrug efflux pump activity. These data suggest that, in addition to point mutations in bos1, drug efflux pump activity and potentially a third mechanism of resistance may be contributing to the iprodione HR phenotype. Detached fruit studies showed that field rates of Rovral 4 Flowable (iprodione) did not control iprodione MR and HR isolates.

MATERIALS
Product Number
Brand
Product Description

Supelco
Fludioxonil, PESTANAL®, analytical standard
Supelco
Iprodione, PESTANAL®, analytical standard
Sigma-Aldrich
5-Amino-4-imidazolecarboxamide, 95%
Sigma-Aldrich
Tolnaftate
Dacarbazine impurity B, European Pharmacopoeia (EP) Reference Standard
Tolnaftate for system suitability, European Pharmacopoeia (EP) Reference Standard
Tolnaftate, European Pharmacopoeia (EP) Reference Standard
USP
Tolnaftate, United States Pharmacopeia (USP) Reference Standard
Sigma-Aldrich
[3-(3,5-Dichlorophenyl)-2,4-dioxoimidazolidinyl]-N-(methylethyl)carboxamide, 97%