Fluorescence-based screening of microbial status during bioprocesses

Fluorescence-based screening of microbial status during bioprocesses

[Abstract]: During microbial fermentations and other bioprocesses, monitoring of potential contamination,
cell viability, physiological status and performance is of prime importance. As a given bioprocess
advances, changes in population dynamics, cell physiology or process constituents will likely
have impacts that require intervention. Various off-line procedures have been employed to
estimate status of cells, for example identity, viability, stress tolerance and vitality. However the
current techniques are both protracted and labour-intensive, so are not useful for monitoring
rapidly changing processes. We are developing fluorescence-based techniques to provide
feedback on cell status in rapid timeframes that enable realistic on-line control of process
parameters. Spectroscopic approaches enable on-line analysis of cell populations using simple
and robust instrumentation. While microscopy-based approaches remain labour intensive and
must be carried out off-line, they provide important information on differential individual cell
responses and spatial relationships that may be critical to overall performance. In parallel
studies fluorescence-based approaches have been established or are under development to
determine viability and identity, respectively. The major focus of investigations described here is
assessment in whole cell systems of modulation of cell membrane fluidity, as an indicator of
adaptability and vitality. Membrane fluidity is determined by fluorescence Polarization and
Generalized Polarization of the membrane-localising probe 6-lauroyl-2-dimethylamino
naphthalene (laurdan). Fluidity modulation has been observed in relation to physiological state
(growth phase and glucose repression), temperature up shift and increased concentrations of
ethanol. Furthermore, we detected effects on cell membranes of process additives that may
potentially be toxic or membrane-active, in both yeast and bacterial systems.

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