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United States Department of Agriculture

Agricultural Research Service

Title: A Probe-Based Method for Rapid Identification and Enumeration of Brettanomyces in Wine

item Stender, Henrik - BOSTON PROBES, BEDFORD,MA
item Perry-O'keefe, Heather - BOSTON PROBES, BEDFORD,MA
item Hyldig-Nielsen, Jens - BOSTON PROBES, BEDFORD,MA
item Broomer, Adam - BOSTON PROBES, BEDFORD,MA
item Saracino, Misty - BOSTON PROBES, BEDFORD,MA
item Kurtzman, Cletus
item Young, Barbara - MILLIPORE, BEDFORD, MA

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: July 1, 1999
Publication Date: N/A

Technical Abstract: Brettanomyces (ascosporic state = Dekkera) are well recognized as spoilage yeasts in wine and cause 'mousiness', an undesirable odor and taste. Current methods for identification and enumeration take 1-2 weeks and rely on semi-selective culture medium followed by final identification from morphology and biochemical testing. A new in situ hybridization method using peptide nucleic acid (PNA) probes for simultaneous identification and enumeration of Brettanomyces within two days has been developed. The wine sample is filtered to isolate and separate individual microorganisms onto a membrane which is placed on a culture medium for up to 44 hours prior to testing. Microscopic colonies of Brettanomyces are detected on the membrane by hybridization with peroxidase-labeled PNA probes targeting Brettanomyces 26S rRNA. Excess probe is removed by washing and hybridized probe is visualized by a chemiluminescent reaction. Each Brettanomyces micro-colony is observed as a small dot providing simultaneous identification and enumeration. 100% sensitivity and 100% specificity of the probes have been determined using reference strains and wine isolates of Brettanomyces as well as other yeast species potentially found in wine. Results obtained using the method to detect Brettanomyces in wine samples will be presented.

Last Modified: 4/22/2015
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