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In situ analyses of single Listeria monocytogens cells at subinhibitory concentrations of leucocin 4010 and nisin revealed two subpopulations when measured by fluorescence ratio imaging microscopy (FRIM). The study thus highlights the important practical implications of applying subinhibitory concentrations of bacteriocins, which results in unaffected target cells which makes it suitable for use in food preservation.

For optimization of propagation conditions for an industrially usedBacillus licheniformis, this study examines the effect of transferring cells at the early-stationary growth phase (pH 5.3) to fresh growth medium at pH 5.0-8.0. Intracellular pH (pHi) was measured on a single-cell level, using fluorescence ratio imaging microscopy after staining with 5(6)-carboxyfluorescein diacetate succinimidyl ester. Transcription profiles were determined using a genome DNA microarray. The optimum extracellular pH (pHex) value for growth ofB. licheniformiswas found to be pH 7.0, resulting in the shortest lag phase, highest maximum specific growth rate and maximum biomass formation. An average pH gradient (?pH = pHi - pHex) of approx. 1.0 was found inB. licheniformis15 min after transfer to pHex 5.0-8.0. Up-regulation of genes involved in sucrose uptake at pH 7.0 could be related to the optimum growth observed. Transcription profiles indicated that the organism was experiencing phosphate starvation upon transfer to pH 7.0 and pH 8.0. Mechanisms involved in pHi regulation appeared to include changes in fatty acid synthesis to yield a more rigid cell membrane structure at low pHex values and conversion of pyruvate to acetoin instead of acetate for neutralization of low pHex values. [ABSTRACT FROM AUTHOR]

Shorter lag phases were obtained in cultivations of Bacillus licheniformis using early-compared to late-stationary growth phase inocula and using liquid versus solid propagation medium. Flow cytometry and fluorescence ratio imaging microscopy (FRIM) after staining with 5(6)-carboxyfluorescein diacetate succinimidyl ester (CFDA-SE), confirmed that liquid early-stationary growth phase inoculum had a higher vitality and was more homogeneous than solid late-stationary growth phase inoculum. DNA-microarray analyses indicated that liquid early-stationary growth phase inoculum was in a more active state in terms of cell multiplication whereas solid late-stationary growth phase inoculum was induced to some spore formation potentially causing delayed growth initiation. [Copyright &y& Elsevier]

For Bacillus licheniformis SJ4628, an organism widely used in the enzyme industry, methods for determination of cell vitality at a single cell level using 5(6)-carboxyfluorescein diacetate succinimidyl ester in combination with fluorescence ratio imaging microscopy and flow cytometry were developed. Immediately after inoculation and during growth, changes in intracellular pH values determined by fluorescence ratio imaging microscopy and in green fluorescence intensities determined by flow cytometry were observed. Correlations between the capacity to multiply and intracellular pH or green fluorescence intensity were demonstrated. Populations of cells not having a pH gradient or exhibiting low fluorescence intensities had significantly longer lag phases than populations of cells with a pH gradient and high fluorescence intensities. [Copyright &y& Elsevier]

For Bacillus licheniformis SJ4628, an organism widely used in the enzyme industry, methods for determination of cell vitality at a single cell level using 5(6)-carboxyfluorescein diacetate succinimidyl ester in combination with fluorescence ratio imaging microscopy and flow cytometry were developed. Immediately after inoculation and during growth, changes in intracellular pH values determined by fluorescence ratio imaging microscopy and in green fluorescence intensities determined by flow cytometry were observed. Correlations between the capacity to multiply and intracellular pH or green fluorescence intensity were demonstrated. Populations of cells not having a pH gradient or exhibiting low fluorescence intensities had significantly longer lag phases than populations of cells with a pH gradient and high fluorescence intensities.