Development of flow cytometry technique for detection of thinning of peptidoglycan layer as a result of solvent production by Clostridium pasteurianum.

Clostridium pasteurianum forms acetic and butyric acids in an initial growth phase, which is a typical feature of clostridial acetone-butanol fermentation where an initial accumulation of acids is followed by production of solvents 1-butanol, acetone and ethanol. The initiation of the solvent production coupled with endospore formation leads to decrease of cell-wall thickness; thinner cell wall is more resistant against solvents and dyes. These changes can be observed by the method based on adaptation of Gram staining. The cell wall of G+ bacteria allows the entry of hexidium iodide and rhodamine 123, whereas the outer membrane of G- bacteria does not allow the uptake and therefore G+ bacteria are stained with higher fluorescence intensity than G- bacteria. The ratio of fluorescence intensity (FI) to forward scatter (FSC) was determined to correspond to G+ bacteria when clostridia were producing less solvents. The significant drop of the ratio FI to FSC to the level corresponding to G- bacteria is detected after initiation of solvent production.

Lactic acid production in a cell retention continuous culture using lignocellulosic hydrolysate as a substrate.

The effect of lignocellulosic hydrolysate of crushed corn cobs on the kinetics of growth and lactic acid production of Lactobacillus casei and L. lactis in the cell retention continuous culture was studied. The continuous cultivations were carried out in a continuous flow stirred bioreactor combined in a recycle loop with an ultrafiltration module retaining all biomass and allowing the continuous removal of metabolites, including lactic acid, as a cell-free permeate. Based on computer-aided analysis of experimental data, a simple physiological model of lactic acid cultivation was developed. The parameters of the model were estimated by non-linear regression.