ARS | Publication request: Dual fluorochrome flow cytometric assessment of yeast viability

Submitted to: Current Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 9, 2009
Publication Date: January 5, 2010
Citation: Hernlem, B.J., Hua, S.T. 2010. Dual fluorochrome flow cytometric assessment of yeast viability. Current Microbiology. 61:57-63.

Interpretive Summary: Some yeasts are useful to control unwanted (aflatoxin producing) fungi when applied to crops (including nut crops such as pistachio, almond, walnut &c.). It is important to be able to monitor the condition of these yeast in storage and in the environment where sunlight and temperature can affect their survival. The authors, collaborating as members of the Foodborne Contaminants and Plant Mycotoxin Research Units of the ARS-WRRC, found and developed a test method shown to be useful. It is expected that this method will aid development of longer-lived and more effective yeast formulations that will promote food safety and marketability of crops.

Technical Abstract: A novel staining protocol is reported for the assessment of viability in yeast, specifically the biocontrol yeast, Pichia anomala. Employing both the red fluorescent membrane potential sensitive oxonol stain DiBAC4(5) (Bis-(1,3-dibutylbarbituric acid)pentamethine oxonol), a structural analog of the commonly used DiBAC4(3) (Bis-(1,3-dibutylbarbituric acid)trimethine oxonol), with one of the esterase dependent green fluorogenic probes such as CFDA-AM (5-Carboxyfluorescein diacetate, acetoxymethyl ester) or Calcein-AM (Calcein acetoxymethyl ester), a two-color flow cytometric method was developed, which yields rapid quantitative information on the vitality and vigor of yeast cell cultures. The method was validated by cell sorting and analysis of live, stored, heat killed and UV-treated yeast.