Flow cytometric applications in microbiological research: fermentation, biosensors, and food safety.
K. R. Harkins, A. Pometto III, G. A. Beattie, R. J. Venugopal and J. S. Dickson.

Collaborations with several microbiologists at Iowa State University have led to the development of an array of rapid cytometry applications for analyzing microbes using the Coulter XL in the cytometry core facility. 1) The counting and viability state of laboratory grown cultures of Escherichia, Lactobacillus, and Salmonella can be accurately determined
with a detection limit of 1,000 bacteria/ml of culture using the BacLightTM kit from Molecular Probes and the addition of 6 micron beads. This method has proven useful in counting microorganisms used in phagocytosis and stress test studies and in evaluating stabilizers used in the process of long term preservation of fermentation microorganisms. 2) We have initiated studies to evaluate the ability of bacterial "biosensors" to sense the environmental conditions present in microhabitats on plant surfaces. The biosensors contain fusions of environmentally responsive promoters to the gene encoding the green fluorescent protein (GFP). After introduction of these biosensors to plant surfaces, their exposure to environmental signals can be assessed based on cytometric measurements of the fluorescence and light scatter properties of the bacteria recovered from the plant surfaces. 3) Finally, clarified suspensions of bacteria isolated from spoiling ground pork can be counted using the bead addition method in conjunction with the BacLightTM stains. Live bacteria can be readily distinguished from meat tissue debris. The cytometric bacterial counts correlated with standard plate counts at concentrations of 1 05 microorganisms and above per gram of meat sample.