Elimination of free-living amoebae in fresh water with pulsed electric fields.

This study investigates the effects of pulsed electric fields on the inactivation of trophozoite form of Naegleria lovaniensis Ar9M-1 in batch and flow processes, systematically examining the lethal effect of field strength, pulse duration, number of pulses, and pulse frequency. Our results show that amoebae eradication is modulated by pulse parameters, composition of the pulsing medium, and physiological state of the cells. Cell survival is not related to the energy delivered to the cell suspension during the electrical treatment. For a given energy a strong field applied for a short cumulative pulse duration affects viability more than a weak field with a long cumulative pulsation. We also determine the optimal electrical conditions to obtain an inactivation rate higher than 95% while using the least energy. Flow processes allow to treat large-scale volumes. Our results show that the most efficient flow process for amoeba eradication requires a field parallel to the flow. Pulsed electric fields are a new and attractive method for inactivating amoebae in large volumes of fresh water.

Recent biotechnological developments of electropulsation. A prospective review.

During the last 25 years, basic research has improved our knowledge on the molecular mechanisms triggered at the membrane level by electric pulses. Applied aspects may now be used under safe conditions. Electropulsation is known as a very efficient tool for obtaining gene transfer in many species to produce genetically modified organisms (GMO). This is routinely used for industrial purposes to transfer exogenous activities in bacteria, yeasts and plants. The method is simple and of a low cost. But electropulsation is not limited to this application for biotechnological purposes. It is known that the field-associated membrane alterations can be irreversible. The pulsed species cannot recover after the treatment. Their viability is strongly affected. This appears as a very promising technology for the eradication of pathogenic microorganisms. Recent developments are proposed for sterilization purposes. New flow technologies of field generation allow the treatment of large volumes of solution. When high flow rates are used, microorganisms are submitted both to a hydromechanical and to an electrical stress. The synergy of the two effects may be present when suitable pulsing conditions are chosen. Several examples for the treatment of domestic water and in the food industry are described. Walled microorganisms are affected not only at the membrane level. We observed that alterations are present on the cell wall. A very promising technology is the associated controlled leakage of the cytoplasmic soluble proteins. Large dimeric proteins such as beta-galactosidases can be extracted at a high yield. High volumes can be treated by using a flow process. Extraction of proteins is obtained with many systems including mammalian cells.

Chinese hamster ovary cells sensitivity to localized electrical stresses.

Application of an external electric field on a cell suspension induces an alteration in the membrane structure giving free access to the cell cytoplasm. Under mild pulsation conditions, permeabilization is a reversible process which weakly affects cell viability while drastic electrical conditions lead to cell death. The field pulse must be considered as a complex stress applied on the cell assembly. This study is a systematic investigation of the stress effects of field strength, pulse duration and number of pulses, at given joule energy. The loss in cell viability is not related to the energy delivered to the system. At a given joule energy, a strong field during a short cumulated pulse duration affects more viability than using a weak field associated with a long cumulated pulsation. At a given field strength and for a given cumulated pulse duration an accumulation of short pulses is also observed to be very damaging for cells. A control by the delay between the pulses suggests a memory effect. The field effect appears also to be vectorial in line with the known asymmetry of the membrane organization. These results suggest that processes at a cellular level are involved, either an activation of cell death or damage in cellular functions.

The interchain disulfide bond between TCR alpha beta heterodimers on human T cells is not required for TCR-CD3 membrane expression and signal transduction.

In the present paper, it was attempted to define the amino acids or regions on TCR beta molecules that determine the TCR alpha-TCR beta interaction. Sequence studies on HBP-ALL variant cells with an intrinsic deficiency in TCR alpha beta dimer formation elucidated a conserved amino acid motif in the TCR-C beta beta-strand E, = Y(C)(L)(S)SRLR(V)(S)(A); this motif seems to represent one interaction area for the TCR alpha-TCR beta interaction. In addition, amino acids in the connecting peptide may be shaped in a precise structure (by the interactions with CD3 molecules?) involved in TCR alpha-TCR beta dimerization. This result was supported by the finding that the interchain disulfide bond between TCR alpha and beta chains is not required for membrane expression or transmembrane signal transduction of TCR alpha beta-CD3 complexes. Finally, comparative results from two membrane TCR-CD3-negative Jurkat variants R4.9 and E6.E12 suggest that TCR-C beta exon 1- and 2-encoded amino acids are important for the TCR beta-CD3 gamma epsilon association.