Demethylation inhibitor (DMI)-resistant strains of the plant pathogenic fungus Penicillium digitatum were shown to be simultaneously resistant to cycloheximide, 4-nitroquinoline-N-oxide (4NQO), and acriflavine. APMR1 (Penicillium multidrug resistance) gene encoding an ATP-binding cassette (ABC) transporter (P-glycoprotein) was cloned from a genomic DNA library of a DMI-resistant strain (LC2) ofPenicillium digitatum by heterologous hybridization with a DNA fragment containing an ABC-encoding region from Botrytis cinerea. Sequence analysis revealed significant amino acid homology to the primary structures of PMR1 (protein encoded by thePMR1 gene) and ABC transporters of Saccharomyces cerevisiae (PDR5 and SNQ2), Schizosaccharomyces pombe(HBA2), Candida albicans (CDR1), and Aspergillus nidulans (AtrA and AtrB). Disruption of the PMR1 gene of P. digitatum DMI-resistant strain LC2 demonstrated that PMR1 was an important determinant of resistance to DMIs. The effective concentrations inhibiting radial growth by 50% (EC50s) and the MICs of fenarimol and bitertanol for the PMR1disruptants (Δpmr1 mutants) were equivalent to those for DMI-sensitive strains. Northern blot analysis indicated that severalfold more PMR1 transcript accumulated in the DMI-resistant strains compared with those in DMI-sensitive strains in the absence of fungicide. In both DMI-resistant and -sensitive strains, transcription of PMR1 was strongly enhanced within 10 min after treatment with the DMI fungicide triflumizole. These results suggested that the toxicant efflux system comprised of PMR1 participates directly in the DMI resistance of the fungus.

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The complete nucleotide sequence of the genomic RNA of cucumber green mottle mosaic virus watermelon strain SH (CGMMV-SH) was determined using cloned cDNA. This sequence is 6421 nucleotides long containing at least four open reading frames, which correspond to 186K, 129K, 29K and 17.3K proteins. The 17.3K protein is the coat protein. Sequence analysis shows that CGMMV-SH is very closely related to another watermelon strain, CGMMV-W, although three amino acid substitutions in the 29K protein were found between these strains. The sequence was also compared to those of other tobamoviruses, tobacco mosaic virus (TMV) vulgare, TMV-L (a tomato strain) and tobacco mild green mosaic virus reported by other groups. It shows 55 to 56% identity with these viruses. The size and location of the open reading frames are very similar to those of TMV but the 129K and 186K proteins are composed of 1142 and 1646 amino acids, being larger than those of TMV by 27 and 31 amino acids, respectively. The deduced amino acid sequences of these proteins are highly homologous to those of TMV, especially in the readthrough downstream region of the 186K protein.

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