The effect of frequency and electric field strength on diffusion enhancement from cells and impedimetric signature were of considerable interest. The objectives were: to investigate the effects of moderate electric field (MEF, an electric field from 1-1000 V/cm) at different frequencies and stages of growth, on growth kinetics and metabolic activity of Lactobacillus acidophilus OSU 133, to investigate temporary cell membrane pore formation under the influence of MEF, and to develop a bio-detector chip to identify a single cell status based on impedimetric detection techniques. The effect of MEF at various frequency levels (45, 60, and 90 Hz) and stages of growth on microbial growth kinetics and bacteriocin (Lacidin A) production was investigated. MEF treatments with purely sinusoidal waveforms at all frequencies at suboptimum growth temperature produced a shorter lag phase than conventional fermentation. There was a significant increase in the bacteriocin production when MEF at 60 Hz with high frequency harmonics was applied at the early stage of the growth. We hypothesized that the effect of MEF on bacterial fermentation may be traced to temporary permeabilization of cells under electric fields.
To investigate temporary cell membrane pore formation, around the cell membrane at different MEF frequency (45, 60, 1000, 10000 Hz) and stages of growth (lag, exponential, stationary phases); we used fluorescent stains that binds to intracellular compounds. Propidium iodide (PI) is cell membranes impermeable; however when cells are made permeable, PI enters and binds to nucleic acid, exhibiting red fluorescence. SYTO 9 (green) is a cell permeable stain. Only cells under influence of 45 and 60 Hz MEF exhibit fluorescent red as evidence of temporary permeabilization. Cells at lag phase showed the greatest susceptibility to permeabilization followed by those at exponential phase. No evidence of electroporation was observed during the stationary phase.
Microfabrication techniques were used, to develop a device that contains wells that house individual microorganisms. The fundamental hypothesis here was that live and dead cells can be distinguished via an impedimetric signature, enabling rapid enumeration on a single cell basis. This device may potentially provide a rapid method for detection of microbial inactivation.
|Advisor:||Sastry, Sudhir K.|
|Commitee:||Hansford, Derek, Yousef, Ahmed E.|
|School:||The Ohio State University|
|Department:||Food, Agricultural, and Biological Engineering|
|School Location:||United States -- Ohio|
|Source:||DAI-A 78/11(E), Dissertation Abstracts International|
|Keywords:||Bacteriocin production, Fermentation, Frequency, Growth kinetics, Growth stage, Moderate electric field|
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