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Identification and Characterization of Benzimidazole Resistance in.pdf

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Identification and Characterization of Benzimidazole Resistance in

APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Dec. 2003, p. 7145–7152 Vol. 69, No. 12 0099-2240/03/$08.000 DOI: 10.1128/AEM.69.12.7145–7152.2003 Copyright ? 2003, American Society for Microbiology. All Rights Reserved. Identification and Characterization of Benzimidazole Resistance in Monilinia fructicola from Stone Fruit Orchards in California Zhonghua Ma, Michael A. Yoshimura, and Themis J. Michailides* Department of Plant Pathology, University of California, Kearney Agricultural Center, Parlier, California 93648 Received 1 May 2003/Accepted 4 September 2003 Low and high levels of resistance to the benzimidazole fungicides benomyl and thiophanate-methyl were observed in field isolates of Monilinia fructicola, which is the causative agent of brown rot of stone fruit. Isolates that had low levels of resistance (hereafter referred to as LR isolates) and high levels of resistance (hereafter referred to as HR isolates) were also cold and heat sensitive, respectively. Results from microsatellite DNA fingerprints showed that genetic identities among the populations of sensitive (S), LR, and HR isolates were very high (0.96). Analysis of DNA sequences of the -tubulin gene showed that the LR isolates had a point mutation at codon 6, causing a replacement of the amino acid histidine by tyrosine. Codon 198, which encodes a glutamic acid in S and LR isolates, was converted to a codon for alanine in HR isolates. Based on these point mutations in the -tubulin gene, allele-specific PCR assays were developed for rapid detection of benzimidazole- resistant isolates of M. fructicola from stone fruit. Brown rot, caused by the fungal pathogen Monilinia fructi- cola (G. Wint.) Honey, is one of the most destructive diseases of stone fruits. Chemical control of brown rot is an important strategy for the management of this disease. The benzimid- azole fungicides thiophanate-methyl and benomyl have been used widely for controlling the disease. In California, benomyl was used almost exclusive