Characterization of boscalid-resistance conferring mutations in the SdhB subunit of respiratory complex II and impact on fitness and mycotoxin production in Penicillium expansum laboratory strains

Laboratory mutants of Penicillium expansum highly resistant (Rfs: 90 to >500, based on EC50s) to Succinate Dehydrogenase Inhibitors (SDHIs) were isolated after UV-mutagenesis and selection on media containing boscalid. A positive correlation was found between sensitivity of isolates to boscalid and other SDHIs such as isopyrazam and carboxin but not to fungicides affecting other cellular pathways or processes, such as the triazole flusilazole, the phenylpyrrole fludioxonil, the anilinopyrimidine cyprodinil and the benzimidazole benomyl. Most of the boscalid-resistant strains were more sensitive to the SDHI fluopyram and the QoI pyraclostrobin. In order to investigate the mechanism responsible for the observed resistance profiles, part of the SdhB subunit isolated the wild type and boscalid-resistant isolates, was genetically characterized. Comparison of the deduced amino-acid sequence between resistant and wild-type isolates revealed two point mutations at a position corresponding to codon 272 of the respective SdhB protein in Botrytis cinerea. The substitution of histidine by arginine was found in boscalid-resistant isolates which were equally sensitive to fluopyram compared with the wild-type whereas the replacement of histidine by tyrosine was found in strains with increased sensitivity to fluopyram. No adverse effects of resistance mutations were observed on fitness determining parameters such as osmotic sensitivity, sporulation and pathogenicity, while mycelial growth rate and spore germination was negatively affected in some of the mutants studied. P. expansum mutant strains displayed significantly perturbed patulin and citrinin levels as compared to the wild-type parent strain both in vitro and in vivo as revealed by thin layer (TLC) and high performance liquid chromatography (HPLC). (C) 2017 Elsevier Inc. All rights reserved.