Brettanomyces Medium

Mix and dissolve the following ingredients. Peptone Yeast extract 

If desired, cycloheximide can be added as a sterile filtered solution to the cooled media before solidification. Final concentration in the medium should be between 20 and lOOmg/L to inhibit Saccharomyces. 

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Rodrigues, N., Gongalves, G., Pereira-da-SUva, S., Malfeito-Ferreira, M., Loureiro, V. (2001). Development and use of a new medium to detect yeasts of the genera Dekkera/Brettanomyces spp. J. Appl. Microbiol., 90, 588-599. 

Agar gel is added to the nutrient medium to cul-mre yeast prior to counting. It is supplemented with biphenyl (0.015% in ethanol) to prevent mold development and 0.01% chloramphenicol to inhibit bacterial growth. Cycloheximide (0.1%) may be added to select non-Saccharomyces" Brettanomyces, etc.) yeasts, as they alone survive this treatment. The incubation period is around 2-3 days for the Saccharomyces genus and... 

Part of the caprylic flavor (several medium chain fatty acids associated (low levels) with pale lager beers (released during autolysis of yeast)) (Ttessl, Bahri, Kossa, 1980). May be produced by Brettanomyces (wet leather, goat-like, wet dog notes associated witb caprylic, capric, and caproic acids). 

Probably the most significant stumbling block in successful routine laboratory identification of Brettanomyces and Dekkera lies in the fundamental requirements of taxonomic guides to demonstrate the presence (or absence) of a sexual phase in the life cycle of the yeast. Dekkera requires a sporulation medium that includes augmentation with several vitamins (see Procedure 3.5.2). Required in microgram and milligram amounts, these nutrients are not easily and routinely supplied in most production-oriented laboratories. Hagan (1979) notes that even under ideal conditions, relatively poor sporulation (<1%) is observed. As a result, suspect isolates are often reported as Brett-like or Brettanomyces/Dekkera. ... 

Van der Walt (1970) suggests that in the case of fastidious yeast, such as Brettanomyces/Dekkera, cell suspensions be prepared in stock vitamin solution (see 3.5.2 for preparation). In this case, and in general, it is advisable to prepare a control to detect carryover of nitrogen or carbon from the original culture. The control, here, consists of only basal medium and yeast. 

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