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Applied and Environmental Microbiology, December 1998, p. 4904-4911, Vol. 64, No. 12
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Characterization of Cell Lysis in Pseudomonas putida Induced upon Expression of Heterologous Killing Genes

M. Carmen Ronchel,1,2 Lázaro Molina,2 Angela Witte,3 Werner Lutbiz,3 Søren Molin,4 Juan L. Ramos,1 and Cayo Ramos1,4

Department of Biochemistry and Molecular and Cellular Biology of Plants, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas,1 and GX-Biosystems España S.L.,2 Granada, Spain; Department of Microbiology, University of Vienna, Vienna, Austria3; and Department of Microbiology, Technical University of Denmark, Lyngby, Denmark4

Received 15 June 1998/Accepted 25 September 1998

Active biological containment systems are based on the controlled expression of killing genes. These systems are of interest for the Pseudomonadaceae because of the potential applications of these microbes as bioremediation agents and biopesticides. The physiological effects that lead to cell death upon the induction of expression of two different heterologous killing genes in nonpathogenic Pseudomonas putida KT2440 derivatives have been analyzed. P. putida CMC4 and CMC12 carry in their chromosomes a fusion of the PA1-04/03 promoter to the Escherichia coli gef gene and the phi X174 lysis gene E, respectively. Expression of the killing genes is controlled by the LacI protein, whose expression is initiated from the XylS-dependent Pm promoter. Under induced conditions, killing of P. putida CMC12 cells mediated by phi X174 lysis protein E was faster than that observed for P. putida CMC4, for which the Gef protein was the killing agent. In both cases, cell death occurred as a result of impaired respiration, altered membrane permeability, and the release of some cytoplasmic contents to the extracellular medium.

Applied and Environmental Microbiology, December 1998, p. 4904-4911, Vol. 64, No. 12
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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