54F-15

New non thermal preservation processes, as the application of pulsed electric fields (PEF), are being developed with the aim of inactivating spoilage or pathogenic microorganisms without affecting the quality and nutritional characteristics of foods. Although the mechanism of microbial inactivation by PEF treatment is not completely elucidated, permeabilization of the cell membrane is generally considered as the main factor. Therefore, PEF treatments may facilitate the passage of both small solutes and macromolecules across the membrane. In fact, this feature has long been applied in molecular biology for bacterial transformation.

The aim of this work was a contribution to determine whether PEF treatments could lead to electrotransformation of cells.

For this purpose, Escherichia coli DH5a, a laboratory strains widely used in molecular biology research, was chosen as a model organism in order to assess the chance of recovery of electrotransformants under several conditions. Cells prepared using procedures designed for electrotransformation in laboratory were subjected to PEF treatments in the presence of plasmid DNA using an OSU-4D bench-scale continuous PEF system. Bipolar, square wave, 20 ms width pulses were applied at a flow rate of 60 ml/min.

Our results showed that, when subjected to a mild PEF treatment (15 kV/cm, 700 µs), a low number of electrotransformants could be isolated. Temperature had a major effect on the frequency of transformation: the frequency obtained was an order of magnitude higher at 25°C than at 15 or 1.5°C. When the electric field strength applied was raised to 20 kV or higher, no transformants were isolated.

These results indicate that electrotransformation can occur as a consequence of PEF treatment, when mild treatments were applied. Therefore, for a safe use of this technology, the risk of transformation by PEF should be evaluated when setting up a low intensity PEF process.