ABSTRACT: Botrytis cinerea (gray mold) is one of the most destructive pathogens of cherry tomatoes, causing fruit decay and economic loss. Fludioxonil is an effective fungicide widely used for crop protec-tion and is essential for controlling tomato gray mold. The emergence of fungicide-resistant strains has made the control of Botrytis cinerea more difficult. While the genome of Botrytis cinerea is available, there are few reports regarding the large-scale functional annotation of the genome using expressed genes derived from transcriptomes, and the mechanism(s) underlying such flu-dioxonil resistance remain unclear. The present study prepared RNA-sequencing (RNA-seq) li-braries for three Botrytis cinerea strains [two highly resistant (LR and FR) versus one highly sen-sitive (S) to fludioxonil], with and without fludioxonil treatment, to identify fludioxonil responsive genes that facilitate fungicide resistance. Functional enrichment analysis identified nine resistant related DEGs in the fludioxonil-induced LR and FR transcriptome that were simultaneously up regulated, and seven resistant related DEGs down regulated. These included adenosine tri-phosphate (ATP)-binding cassette (ABC) transporter-encoding genes, major facilitator super-family (MFS) transporter-encoding genes, and the high-osmolarity glycerol (HOG) pathway homologues or related genes. The expression patterns of twelve out of the sixteen fludioxo-nil-responsive genes, obtained from the RNA-sequence data sets were validated using quantita-tive real-time PCR (qRT-PCR). Based on RNA-sequence analysis it was found that fugal HHKs, like BOS1, BcHHK2, and Bchhk17, were in some way involved in the fludioxonil resistance of B. cinerea, in addition, a number of ABC and MFS transporter genes that were not reported before, such as BcATRO, BMR1, BMR3, BcNMT1, BcAMF1, BcTOP1, BcVBA2, and BcYHK8 were differen-tially expressed in the fludioxonil-resistant strains, indicating that overexpression of these efflux transporters located in the plasma membranes played a crucial role in the fludioxonil resistant mechanism of B. cinerea. These lines of evidence together allowed us to draw a general portrait of the anti-fludioxonil mechanisms for Botrytis cinerea, and the assembled and annotated transcrip-tome data provide valuable genomic resources for further study of the molecular mechanisms of B. cinerea resistance to fludioxonil.
