| Research Abstract |
1. Mechanisms of benomyl fungicide-resistance in Botrytis cinerea. The pulsed-field gel electrophoresis (PFGE) pattern of 5 benomyl-resistant strains (IHES-3, IHES-4, CAES-7, PHDF-6 and PHGF-1) and a sensitive strain (IPCR-1) of B. cinerea were examined The four resistant strains (IHES-4, CAES-7, PHDF-6 and PHGF-1) yielded patterns that were identical to that of the sensitive strain. The resistant strain IHES-3, which contained a lot of small nuclei in the hyphae, yielded the pattern that was almost identical to that of the above strains, but still more contained a low molecular band. For all the strains analyzed, the genome size was ca. 10 Mb, with no apparent difference among the teat strains of B. cinerea. Genomic DNAs from the 6 strains were surveyed for restriction fragment length polymorphisms (RFLPs) by digestion with the restriction enzymes (BamHI, BglII, EcoRI, HindIII, NotI and ShpI) and analyzed southern blotting against a puc18-DNA, yeast-RNA or E. coli-rRNA probe labelled
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with biotin. The different patterns of rflp between the resistant and the sensitive strains was not observed. From all the strains analyzed, mitochondorial DNA (ca. 2.15kb) was Isolated, and the quantities from the resistant strains were larger than that from the sensitive strain.
2. Induction mechanisms of dicarboximide fungicide-resistance in Botrytis cinerea. Dicarboximide and/or benomyl fungicide-resistant strains (CAES and IHES series) of B. cinerea were pre-shade cultured in MS media, and after removal of the mycelia, the sensitive isolate (DS-2) of Saccharomyces cerevisiae was post-shake cultured in the respective filtrates. The DS-2 cells cultured in the filtrates were induced resistance to dicarboximide fungicides, but not to benomyl. The induced fungicide-resistance had been maintained through more than five times of consecutive subcultures. The respective patterns of PFGE and RFLP of the resistant isolates (DR-C2 and DR-C4) induced by the filtrates of B. cinerea were examined. The resistant isolates yielded patterns that were identical to that of the sensitive isolate. From the DNA band patterns by PFGE and RFLP, genetic difference the resistant and the sensitive isolates of S. cerevisiae was not found. Less
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