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PMID: 9603843 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Establishment of new genetic traits in a microbial biofilm community.

Applied and environmental microbiology ·Vol. 64 ·No. 6 ·1998-06-00 ·Pages 2247-55

Christensen BB, Sternberg C, Andersen JB, Eberl L, Moller S, Givskov M, Molin S

Abstract

Conjugational transfer of the TOL plasmid (pWWO) was analyzed in a flow chamber biofilm community engaged in benzyl alcohol degradation. The community consisted of three species, Pseudomonas putida RI, Acinetobacter sp. strain C6, and an unidentified isolate, D8. Only P. putida RI could act as a recipient for the TOL plasmid. Cells carrying a chromosomally integrated lacIq gene and a lacp-gfp-tagged version of the TOL plasmid were introduced as donor strains in the biofilm community after its formation. The occurrence of plasmid-carrying cells was analyzed by viable-count-based enumeration of donors and transconjugants. Upon transfer of the plasmids to the recipient cells, expression of green fluorescence was activated as a result of zygotic induction of the gfp gene. This allowed a direct in situ identification of cells receiving the gfp-tagged version of the TOL plasmid. Our data suggest that the frequency of horizontal plasmid transfer was low, and growth (vertical transfer) of the recipient strain was the major cause of plasmid establishment in the biofilm community. Employment of scanning confocal laser microscopy on fixed biofilms, combined with simultaneous identification of P. putida cells and transconjugants by 16S rRNA hybridization and expression of green fluorescence, showed that transconjugants were always associated with noninfected P. putida RI recipient microcolonies. Pure colonies of transconjugants were never observed, indicating that proliferation of transconjugant cells preferentially took place on preexisting P. putida RI microcolonies in the biofilm.

MeSH Terms
Acinetobacter/genetics,metabolism Benzyl Alcohol/metabolism Biodegradation, Environmental Biofilms/growth & development Conjugation, Genetic DNA Primers/genetics Fluorescence Genes, Bacterial Green Fluorescent Proteins Luminescent Proteins/genetics Models, Biological Plasmids/genetics Pseudomonas putida/genetics,metabolism RNA, Bacterial/genetics RNA, Ribosomal, 16S/genetics
Chemicals
DNA Primers Luminescent Proteins RNA, Bacterial RNA, Ribosomal, 16S Green Fluorescent Proteins Benzyl Alcohol
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Christensen B B
Department of Microbiology, The Technical University of Denmark, DK-2800 Lyngby, Denmark.
Sternberg C
Andersen J B
Eberl L
Moller S
Givskov M
Molin S
References (43)
43 references, click to expand
  1. The specific growth rate of Pseudomonas putida PAW1 influences the conjugal transfer rate of the TOL plasmid.
    Appl Environ Microbiol. 1993 Oct;59(10):3430-7 PMID: 8250565
  2. New unstable variants of green fluorescent protein for studies of transient gene expression in bacteria.
    Appl Environ Microbiol. 1998 Jun;64(6):2240-6 PMID: 9603842
  3. Green fluorescent protein as a marker for gene expression and cell biology of mycobacterial interactions with macrophages.
    Mol Microbiol. 1995 Sep;17(5):901-12 PMID: 8596439
  4. 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):6568-72 PMID: 2172217
  5. A general system to integrate lacZ fusions into the chromosomes of gram-negative eubacteria: regulation of the Pm promoter of the TOL plasmid studied with all controlling elements in monocopy.
    Mol Gen Genet. 1992 May;233(1-2):293-301 PMID: 1318499
  6. Gene transfer of Alcaligenes eutrophus JMP134 plasmid pJP4 to indigenous soil recipients.
    Appl Environ Microbiol. 1996 Jul;62(7):2521-6 PMID: 8779592
  7. Use of rRNA fluorescence in situ hybridization for measuring the activity of single cells in young and established biofilms.
    Appl Environ Microbiol. 1993 May;59(5):1354-60 PMID: 7685999
  8. Promoters largely determine the efficiency of repressor action.
    Proc Natl Acad Sci U S A. 1988 Dec;85(23):8973-7 PMID: 3057497
  9. In situ gene expression in mixed-culture biofilms: evidence of metabolic interactions between community members.
    Appl Environ Microbiol. 1998 Feb;64(2):721-32 PMID: 9464414
  10. Green fluorescent protein as a marker for gene expression.
    Science. 1994 Feb 11;263(5148):802-5 PMID: 8303295
  11. Reasons for possible failure of inoculation to enhance biodegradation.
    Appl Environ Microbiol. 1985 Oct;50(4):977-83 PMID: 4083891
  12. Polymerase chain reaction and gene probe detection of the 2,4-dichlorophenoxyacetic acid degradation plasmid, pJP4.
    Appl Environ Microbiol. 1992 Apr;58(4):1271-5 PMID: 1599246
  13. The TOL plasmid is naturally derepressed for transfer.
    J Gen Microbiol. 1982 Dec;128(12):3019-24 PMID: 6134782
  14. A complementation analysis of the restriction and modification of DNA in Escherichia coli.
    J Mol Biol. 1969 May 14;41(3):459-72 PMID: 4896022
  15. Microbial exopolymers provide a mechanism for bioaccumulation of contaminants.
    Microb Ecol. 1994 May;27(3):279-91 PMID: 24190341
  16. Tracking the Response of Burkholderia cepacia G4 5223-PR1 in Aquifer Microcosms.
    Appl Environ Microbiol. 1995 Feb;61(2):448-55 PMID: 16534928
  17. Activity and three-dimensional distribution of toluene-degrading Pseudomonas putida in a multispecies biofilm assessed by quantitative in situ hybridization and scanning confocal laser microscopy.
    Appl Environ Microbiol. 1996 Dec;62(12):4632-40 PMID: 8953734
  18. Transfer and Expression of the Catabolic Plasmid pBRC60 in Wild Bacterial Recipients in a Freshwater Ecosystem.
    Appl Environ Microbiol. 1991 May;57(5):1546-53 PMID: 16348493
  19. DNA amplification to enhance detection of genetically engineered bacteria in environmental samples.
    Appl Environ Microbiol. 1988 Sep;54(9):2185-91 PMID: 3190225
  20. The RDP (Ribosomal Database Project).
    Nucleic Acids Res. 1997 Jan 1;25(1):109-11 PMID: 9016515
  21. Bacterial evolution.
    Microbiol Rev. 1987 Jun;51(2):221-71 PMID: 2439888
  22. Survival of Pseudomonas putida UWC1 containing cloned catabolic genes in a model activated-sludge unit.
    Appl Environ Microbiol. 1989 Oct;55(10):2627-34 PMID: 2604401
  23. Expression and transfer of engineered catabolic pathways harbored by Pseudomonas spp. introduced into activated sludge microcosms.
    Appl Environ Microbiol. 1992 Oct;58(10):3380-6 PMID: 1444370
  24. Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology.
    J Bacteriol. 1990 Feb;172(2):762-70 PMID: 1688842
  25. Phylogenetic identification and in situ detection of individual microbial cells without cultivation.
    Microbiol Rev. 1995 Mar;59(1):143-69 PMID: 7535888
  26. Multicellular organization in a degradative biofilm community.
    Appl Environ Microbiol. 1994 Feb;60(2):434-46 PMID: 16349173
  27. Metabolism of benzoate and the methylbenzoates by Pseudomonas putida (arvilla) mt-2: evidence for the existence of a TOL plasmid.
    J Bacteriol. 1974 Oct;120(1):416-23 PMID: 4418209
  28. Multicopy expression vectors carrying the lac repressor gene for regulated high-level expression of genes in Escherichia coli.
    Gene. 1987;51(2-3):255-67 PMID: 3110013
  29. FACS-optimized mutants of the green fluorescent protein (GFP).
    Gene. 1996;173(1 Spec No):33-8 PMID: 8707053
  30. 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):6557-67 PMID: 2172216
  31. Involvement of a chlorobenzoate-catabolic transposon, Tn5271, in community adaptation to chlorobiphenyl, chloroaniline, and 2,4-dichlorophenoxyacetic acid in a freshwater ecosystem.
    Appl Environ Microbiol. 1992 Jan;58(1):314-25 PMID: 1311543
  32. Environmental application of nucleic acid hybridization.
    Annu Rev Microbiol. 1990;44:625-48 PMID: 2252397
  33. Survival and activity of a 3-chlorobenzoate-catabolic genotype in a natural system.
    Appl Environ Microbiol. 1989 Jun;55(6):1584-90 PMID: 16347951
  34. Analysis and construction of stable phenotypes in gram-negative bacteria with Tn5- and Tn10-derived minitransposons.
    Methods Enzymol. 1994;235:386-405 PMID: 8057911
  35. Site-specific deletions of chromosomally located DNA segments with the multimer resolution system of broad-host-range plasmid RP4.
    J Bacteriol. 1995 Jan;177(1):52-8 PMID: 7798149
  36. The use of Mudlac transposons as tools for vital staining to visualize clonal and non-clonal patterns of organization in bacterial growth on agar surfaces.
    J Gen Microbiol. 1984 May;130(5):1169-81 PMID: 6206197
  37. Survival and activity of Pseudomonas sp. strain B13(FR1) in a marine microcosm determined by quantitative PCR and an rRNA-targeting probe and its effect on the indigenous bacterioplankton.
    Appl Environ Microbiol. 1995 Apr;61(4):1201-7 PMID: 7538271
  38. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors.
    Gene. 1985;33(1):103-19 PMID: 2985470
  39. Bacterial plasmid conjugation on semi-solid surfaces monitored with the green fluorescent protein (GFP) from Aequorea victoria as a marker.
    Gene. 1996;173(1 Spec No):59-65 PMID: 8707058
  40. Channel structures in aerobic biofilms of fixed-film reactors treating contaminated groundwater.
    Appl Environ Microbiol. 1995 Feb;61(2):769-77 PMID: 7574613
  41. Metabolism of toluene and xylenes by Pseudomonas (putida (arvilla) mt-2: evidence for a new function of the TOL plasmid.
    J Bacteriol. 1975 Oct;124(1):7-13 PMID: 1176436
  42. Use of green fluorescent protein for visualization of cell-specific gene expression and subcellular protein localization during sporulation in Bacillus subtilis.
    J Bacteriol. 1995 Oct;177(20):5906-11 PMID: 7592342
  43. Quantification of the effect of substrate concentration on the conjugal transfer rate of the TOL plasmid in short-term batch mating experiments.
    Lett Appl Microbiol. 1995 Sep;21(3):167-72 PMID: 7576502
Article Info
Journal
Applied and environmental microbiology
Abbr.
Appl Environ Microbiol
ISSN
0099-2240
DOI
10.1128/AEM.64.6.2247-2255.1998
Published
1998-06-00
Pages
2247-55
Language
English
Region
United States
NLM ID
7605801
PMCID
PMC106307
Subset
IM
Databases
GENBANK
Y11464, Y11465
External Links
PubMed source DOI: 10.1128/AEM.64.6.2247-2255.1998 PMC Full Text View journal details
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