Skip to main content
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1995 Feb;61(2):481–486. doi: 10.1128/aem.61.2.481-486.1995

A stochastic killing system for biological containment of Escherichia coli.

P Klemm 1, L B Jensen 1, S Molin 1
PMCID: PMC167306  PMID: 7574584

Abstract

Bacteria with a stochastic conditional lethal containment system have been constructed. The invertible switch promoter located upstream of the fimA gene from Escherichia coli was inserted as expression cassette in front of the lethal gef gene deleted of its own natural promoter. The resulting fusion was placed on a plasmid and transformed to E. coli. The phenotype connected with the presence of such a plasmid was to reduce the population growth rate with increasing significance as the cell growth rate was reduced. In very fast growing cells, there was no measurable effect on growth rate. When a culture of E. coli harboring the plasmid comprising the containment system is left as stationary cells in suspension without nutrients, viability drops exponentially over a period of several days, in contrast to the control cells, which maintain viability nearly unaffected during the same period of time. Similar results were obtained with a strain in which the killing cassette was inserted in the chromosome. In competition with noncontained cells during growth, the contained cells are always outcompeted. Stochastic killing obtained by the fim-gef fusion is at present relevant only as a containment approach for E. coli, but the model may be mimicked in other organisms by using species-specific stochastic expression systems.

Full Text

The Full Text of this article is available as a PDF (200.9 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abraham J. M., Freitag C. S., Clements J. R., Eisenstein B. I. An invertible element of DNA controls phase variation of type 1 fimbriae of Escherichia coli. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5724–5727. doi: 10.1073/pnas.82.17.5724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Appleyard R K. Segregation of New Lysogenic Types during Growth of a Doubly Lysogenic Strain Derived from Escherichia Coli K12. Genetics. 1954 Jul;39(4):440–452. doi: 10.1093/genetics/39.4.440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. BERTANI G. Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli. J Bacteriol. 1951 Sep;62(3):293–300. doi: 10.1128/jb.62.3.293-300.1951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blomfield I. C., Calie P. J., Eberhardt K. J., McClain M. S., Eisenstein B. I. Lrp stimulates phase variation of type 1 fimbriation in Escherichia coli K-12. J Bacteriol. 1993 Jan;175(1):27–36. doi: 10.1128/jb.175.1.27-36.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Blomfield I. C., McClain M. S., Princ J. A., Calie P. J., Eisenstein B. I. Type 1 fimbriation and fimE mutants of Escherichia coli K-12. J Bacteriol. 1991 Sep;173(17):5298–5307. doi: 10.1128/jb.173.17.5298-5307.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  7. Casadaban M. J., Cohen S. N. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. J Mol Biol. 1980 Apr;138(2):179–207. doi: 10.1016/0022-2836(80)90283-1. [DOI] [PubMed] [Google Scholar]
  8. Contreras A., Molin S., Ramos J. L. Conditional-suicide containment system for bacteria which mineralize aromatics. Appl Environ Microbiol. 1991 May;57(5):1504–1508. doi: 10.1128/aem.57.5.1504-1508.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dorman C. J., Higgins C. F. Fimbrial phase variation in Escherichia coli: dependence on integration host factor and homologies with other site-specific recombinases. J Bacteriol. 1987 Aug;169(8):3840–3843. doi: 10.1128/jb.169.8.3840-3843.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dorman C. J., Ní Bhriain N. Thermal regulation of fimA, the Escherichia coli gene coding for the type 1 fimbrial subunit protein. FEMS Microbiol Lett. 1992 Dec 1;78(2-3):125–130. doi: 10.1016/0378-1097(92)90013-e. [DOI] [PubMed] [Google Scholar]
  11. Eisenstein B. I., Sweet D. S., Vaughn V., Friedman D. I. Integration host factor is required for the DNA inversion that controls phase variation in Escherichia coli. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6506–6510. doi: 10.1073/pnas.84.18.6506. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fulks K. A., Marrs C. F., Stevens S. P., Green M. R. Sequence analysis of the inversion region containing the pilin genes of Moraxella bovis. J Bacteriol. 1990 Jan;172(1):310–316. doi: 10.1128/jb.172.1.310-316.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gerdes K., Rasmussen P. B., Molin S. Unique type of plasmid maintenance function: postsegregational killing of plasmid-free cells. Proc Natl Acad Sci U S A. 1986 May;83(10):3116–3120. doi: 10.1073/pnas.83.10.3116. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Herrero M., de Lorenzo V., Timmis K. N. Transposon vectors containing non-antibiotic resistance selection markers for cloning and stable chromosomal insertion of foreign genes in gram-negative bacteria. J Bacteriol. 1990 Nov;172(11):6557–6567. doi: 10.1128/jb.172.11.6557-6567.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Higgins C. F., Dorman C. J., Stirling D. A., Waddell L., Booth I. R., May G., Bremer E. A physiological role for DNA supercoiling in the osmotic regulation of gene expression in S. typhimurium and E. coli. Cell. 1988 Feb 26;52(4):569–584. doi: 10.1016/0092-8674(88)90470-9. [DOI] [PubMed] [Google Scholar]
  16. Johnson R. C., Simon M. I. Hin-mediated site-specific recombination requires two 26 bp recombination sites and a 60 bp recombinational enhancer. Cell. 1985 Jul;41(3):781–791. doi: 10.1016/s0092-8674(85)80059-3. [DOI] [PubMed] [Google Scholar]
  17. Kawula T. H., Orndorff P. E. Rapid site-specific DNA inversion in Escherichia coli mutants lacking the histonelike protein H-NS. J Bacteriol. 1991 Jul;173(13):4116–4123. doi: 10.1128/jb.173.13.4116-4123.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Klemm P., Jørgensen B. J., van Die I., de Ree H., Bergmans H. The fim genes responsible for synthesis of type 1 fimbriae in Escherichia coli, cloning and genetic organization. Mol Gen Genet. 1985;199(3):410–414. doi: 10.1007/BF00330751. [DOI] [PubMed] [Google Scholar]
  19. Klemm P. Two regulatory fim genes, fimB and fimE, control the phase variation of type 1 fimbriae in Escherichia coli. EMBO J. 1986 Jun;5(6):1389–1393. doi: 10.1002/j.1460-2075.1986.tb04372.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. McClain M. S., Blomfield I. C., Eisenstein B. I. Roles of fimB and fimE in site-specific DNA inversion associated with phase variation of type 1 fimbriae in Escherichia coli. J Bacteriol. 1991 Sep;173(17):5308–5314. doi: 10.1128/jb.173.17.5308-5314.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McCormick B. A., Klemm P., Krogfelt K. A., Burghoff R. L., Pallesen L., Laux D. C., Cohen P. S. Escherichia coli F-18 phase locked 'on' for expression of type 1 fimbriae is a poor colonizer of the streptomycin-treated mouse large intestine. Microb Pathog. 1993 Jan;14(1):33–43. doi: 10.1006/mpat.1993.1004. [DOI] [PubMed] [Google Scholar]
  22. Molin S., Boe L., Jensen L. B., Kristensen C. S., Givskov M., Ramos J. L., Bej A. K. Suicidal genetic elements and their use in biological containment of bacteria. Annu Rev Microbiol. 1993;47:139–166. doi: 10.1146/annurev.mi.47.100193.001035. [DOI] [PubMed] [Google Scholar]
  23. Nevola J. J., Stocker B. A., Laux D. C., Cohen P. S. Colonization of the mouse intestine by an avirulent Salmonella typhimurium strain and its lipopolysaccharide-defective mutants. Infect Immun. 1985 Oct;50(1):152–159. doi: 10.1128/iai.50.1.152-159.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Olsen P. B., Klemm P. Localization of promoters in the fim gene cluster and the effect of H-NS on the transcription of fimB and fimE. FEMS Microbiol Lett. 1994 Feb 1;116(1):95–100. doi: 10.1111/j.1574-6968.1994.tb06681.x. [DOI] [PubMed] [Google Scholar]
  25. Pallesen L., Madsen O., Klemm P. Regulation of the phase switch controlling expression of type 1 fimbriae in Escherichia coli. Mol Microbiol. 1989 Jul;3(7):925–931. doi: 10.1111/j.1365-2958.1989.tb00242.x. [DOI] [PubMed] [Google Scholar]
  26. Plasterk R. H., van de Putte P. The invertible P-DNA segment in the chromosome of Escherichia coli. EMBO J. 1985 Jan;4(1):237–242. doi: 10.1002/j.1460-2075.1985.tb02341.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Poulsen L. K., Larsen N. W., Molin S., Andersson P. A family of genes encoding a cell-killing function may be conserved in all gram-negative bacteria. Mol Microbiol. 1989 Nov;3(11):1463–1472. doi: 10.1111/j.1365-2958.1989.tb00131.x. [DOI] [PubMed] [Google Scholar]
  28. Schneider K., Beck C. F. New expression vectors for identifying and testing signal structures for initiation and termination of transcription. Methods Enzymol. 1987;153:452–461. doi: 10.1016/0076-6879(87)53071-3. [DOI] [PubMed] [Google Scholar]
  29. de Lorenzo V., Herrero M., Jakubzik U., Timmis K. N. Mini-Tn5 transposon derivatives for insertion mutagenesis, promoter probing, and chromosomal insertion of cloned DNA in gram-negative eubacteria. J Bacteriol. 1990 Nov;172(11):6568–6572. doi: 10.1128/jb.172.11.6568-6572.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES