Archives of Microbiology

, Volume 171, Issue 6, pp 424–429

Generation and initial characterization of Pseudomonas stutzeri KC mutants with impaired ability to degrade carbon tetrachloride

  • Lycely Del C. Sepúlveda-Torres
  • Narayanan Rajendran
  • Michael J. Dybas
  • Craig S. Criddle
Original paper

DOI: 10.1007/s002030050729

Cite this article as:
Del C. Sepúlveda-Torres, L., Rajendran, N., Dybas, M. et al. Arch Microbiol (1999) 171: 424. doi:10.1007/s002030050729

Abstract

Under iron-limiting conditions, Pseudomonas stutzeri KC secretes a small but as yet unidentified factor that transforms carbon tetrachloride (CT) to CO2 and nonvolatile products when activated by reduction at cell membranes. Pseudomonas fluorescens and other cell types activate the factor. Triparental mating was used to generate kanamycin-resistant lux::Tn5 recombinants of strain KC. Recombinants were streaked onto the surface of agar medium plugs in microtiter plates and were then screened for carbon tetrachloride degradation by exposing the plates to gaseous 14C-carbon tetrachloride. CT+ recombinants generated nonvolatile 14C-labeled products, but four CT recombinants did not generate significant nonvolatile 14C-labeled products and had lost the ability to degrade carbon tetrachloride. When colonies of P. fluorescens were grown next to colonies of CT+ recombinants and were exposed to gaseous 14C-carbon tetrachloride, 14C-labeled products accumulated around the P. fluorescens colonies, indicating that the factor secreted by CT+ colonies had diffused through the agar and become activated. When P. fluorescens was grown next to CT colonies, little carbon tetrachloride transformation was observed, indicating a lack of active factor. Expression of lux reporter genes in three of the CT mutants was regulated by added iron and was induced under the same iron-limiting conditions that induce carbon tetrachloride transformation in the wild-type.

Key words Transposon mutagenesisCarbontetrachlorideBiotransformationBiodegradationLuciferasePseudomonas stutzeri KCReporter genesMutants

Copyright information

© Springer-Verlag Berlin Heidelberg 1999

Authors and Affiliations

  • Lycely Del C. Sepúlveda-Torres
    • 1
  • Narayanan Rajendran
    • 3
  • Michael J. Dybas
    • 2
  • Craig S. Criddle
    • 2
  1. 1.Department of Microbiology, Michigan State University, East Lansing, MI 48824, USAUS
  2. 2.Center for Microbial Ecology, Michigan State University, East Lansing, MI 48824, USAUS
  3. 3.Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI 48824, USAUS
  4. 4.Department of Civil and Environmental Engineering, Stanford University, Stanford, CA 94305-4020, USA e-mail: criddle@ce.stanford.edu, Tel.: +1-650-723-9032, Fax: +1-650-725-9474US