Mutation Incyp51band Overexpression Ofcyp51aandcyp51bconfer Multiple Resistant Todmisfungicide Prochloraz Infusarium Fujikuroi

BACKGROUND Fusarium fujikuroiis a plant pathogen that causes rice bakanae disease. Prochloraz is an imidazole-class sterol, 14 alpha-demethylase inhibitor (DMI), which has been in use for several years as a foliar spray to controlFusariumspp.on agriculturally important monocot crops.F. fujikuroiis highly resistant to prochloraz treatment, and the aim of this study was to clarify the mechanism by whichF. fujikuroirenders itself resistant to prochloraz. RESULTS Recently, prochloraz-resistant strains were identified over a vast geographical area in the agricultural regions of Zhejiang Province, China. It was found that 21.13% and 3.96% of the strains examined were highly resistant (HR) to prochloraz during 2017 to 2018. The HR strains contained a point mutation (S312T) in the FfCYP51B protein, while the strains identified with prochloraz susceptibility had no such point mutation in FfCYP51A/B/C. To confirm whether the mutations in FfCYP51B confer resistance to prochloraz, we exchanged the CYP51B locus between the sensitive strain and the resistant strain by homologous double exchange. The transformed mutants with a copy of the resistant fragment exhibited resistance to prochloraz, and the transformed mutants with a copy of the sensitive fragment exhibited sensitivity to prochloraz. Furthermore, qRT-PCR analysis ofFfcyp51a/b/cgene expression revealed thatFfcyp51aandFfcyp51bwere significantly up-regulated in the prochloraz-resistant strains relative to the sensitive strains inF. fujikuroi. Contrary to our expectation, docking of prochloraz into the modeled binding pocket of FfCYP51B indicated that the affinity between prochloraz and the FfCYP51B increased after the amino acid at codon 312 changed to Thr. CONCLUSION The point mutation S312T in FfCYP51B and overexpression ofFfcyp51aandFfcyp51btogether lead to the prochloraz-resistant phenotype inF. fujikuroi.