Hostname: page-component-78c5997874-fbnjt Total loading time: 0 Render date: 2024-11-19T04:21:05.231Z Has data issue: false hasContentIssue false
  • English
  • Français

Screening of rhizosphere and soil bacteriafor transformability

Published online by Cambridge University Press:  26 October 2007

Babette Richter  and
Kornelia Smalla
Babette Richter
Affiliation:
Federal Biological Research Centre for Agriculture and Forestry, Institute for Plant Virology, Microbiology and Biosafety, Messeweg 11-12, 38104 Braunschweig, Germany
Kornelia Smalla
Affiliation:
Federal Biological Research Centre for Agriculture and Forestry, Institute for Plant Virology, Microbiology and Biosafety, Messeweg 11-12, 38104 Braunschweig, Germany

Abstract

Core share and HTML view are not available for this content. However, as you have access to this content, a full PDF is available via the ‘Save PDF’ action button.

Natural transformation is assumed to be the most likely mechanism by which DNA from transgenic plants could be horizontally transferred to bacteria. In order to determine the occurrence of naturally transformable bacteria amongst bulk and rhizosphere soil bacteria, different transformation strategies were employed using either plasmid DNA (IncQ plasmids pSM1890 and pSM1885, conferring GFP, Smr, Gmr and GFP, Smr, Tcr, respectively) or genomic DNA from rhizosphere isolates, which were chromosomally tagged with mini-Tn5~(GFP, Tcr, as transforming DNA. Transformation assays were done in microtiter plates (262 isolates and pSM1890 or pSM1885), on filters (i) with rhizosphere bacterial community mixed with pSM1890 or pSM1885, (ii) with 24 rhizosphere or soil bacterial isolates mixed with genomic DNA of the corresponding mini-Tn5-tagged strains, and in the rhizosphere of tobacco plants inoculated with rifampicin-resistant bacterial isolates and genomic DNA of the corresponding mini-Tn5-tagged strains added. One transformant colony was obtained when Brevundimonas vesicularis was transformed with genomic DNA of the corresponding mini-Tn5-tagged strain. Attempts to reproduce this result were unsuccessful. With this single exception, transformants were neither detected in the collection of isolates nor in the rhizosphere bacterial community. Acinetobacter baylyi BD413 used as a positive control showed drastically reduced transformation frequencies with plasmid pSM1890 as transforming DNA when mixed with the rhizosphere pellet. All transformants were characterized by BOX-PCR fingerprints, and three different BOX patterns were revealed. Sequencing the 16S rRNA gene showed that all transformants could be assigned to Acinetobacter sp. Since transformants were only observed in the positive control, the introduced BD413 either underwent genomic rearrangements, or competence of the Acinetobacter population present in the rhizosphere was stimulated by the introduction of BD413. The various transformation assays performed indicate that the proportion of rhizosphere or bulk soil bacteria which are naturally transformable is negligibly low.

Keywords

Horizontal gene transfertransformability screeningsoil bacteriarhizosphere bacteriaAcinetobacter baylyi BD413
Type
Research Article
Information
Environmental Biosafety Research , Volume 6 , Issue 1-2: Thematic Issue on Horizontal Gene Transfer , January 2007 , pp. 91 - 99
Copyright
© ISBR, EDP Sciences, 2007

References

Adesina, M, Lembke, A, Costa, R, Speksnijder, A, Smalla, K (2007) Screening of bacterial isolates from various European soils for in vitro antagonistic activity towards Rhizoctonia solani and Fusarium oxysporum: site-dependent composition and diversity revealed. Soil Biol. Biochem. 39: 2818–2828
Andersen JB, Sternberg C, Poulsen LK, Bjorn SP, Givskov M, Molin S (1998) New unstable variants of green fluorescent protein for studies of transient gene expression in bacteria. Appl. Environ. Microbiol. 64: 2240–2246
Berg G, Roskot N, Steidle A, Eberl L, Zock A, Smalla K (2002) Plant dependent genotypic and phenotypic diversity of antagonistic Rhizobacteria isolated from different Verticillium host plants. Appl. Environ. Microbiol. 68: 3328–3338
Berka, RM, Hahn, J, Albano, M, Draskovic, I, Persuh, M, Cui, X, Sloma, A, Widner, W, Dubnau, D (2002) Microarray analysis of the Bacillus subtilis K-state: genome-wide expression changes dependent on ComK. Mol. Microbiol. 43: 1331–1345 CrossRef
Bertolla, F, Pepin, R, Passelègue-Robe, E, Paget, E, Simkin, A, Nesme, X, Simonet, P (2000) Plant genome complexity may be a factor limiting in situ the transfer of transgenic plant genes to the phytopathogen Ralstonia solanacearum. Appl. Environ. Microbiol. 66: 4161–4167 CrossRef
Cérémonie, H, Buret, F, Simonet, P, Vogel, TM (2004) Isolation of lightning-competent soil bacteria. Appl. Environ. Microbiol. 70: 6342–6346 CrossRef
Cérémonie, H, Buret, F, Simonet, P, Vogel, TM (2006) Natural electrotransformation of lightning-competent Pseudomonas sp. strain N3 in artificial soil microcosms. Appl. Environ. Microbiol. 72: 2385–2389 CrossRef
de Vries J, Wackernagel W (1998) Detection of nptII (kanamycin resistance) genes in genomes of transgenic plants by marker-rescue transformation. Mol. Gen. Genet. 257: 606–613
de Vries, J, Wackernagel, W (2002) Integration of foreign DNA during natural transformation of Acinetobacter sp. by homology-facilitated illegitimate recombination. PNAS 99: 2094–2099 CrossRef
de Vries, J, Wackernagel, W (2004) Microbial horizontal gene transfer and the DNA release from transgenic crop plants. Plant Soil 266: 91–104 CrossRef
de Vries J, Meier P, Wackernagel, W (2001) The natural transformation of the soil bacteria Pseudomonas stutzeri and Acinetobacter sp. by transgenic plant DNA strictly depends on homologous sequences in the recipient cells. FEMS Microbiol. Lett. 195: 211–215
de Vries, J, Heine, M, Harms, K, Wackernagel, W (2003) Spread of recombinant DNA by roots and pollen of transgenic potato plants, identified by highly specific biomonitoring using natural transformation of an Acinetobacter sp. Appl. Environ. Microbiol. 69: 4455–4462 CrossRef
de Vries J, Herzfeld T, Wackernagel W (2004) Transfer of plastid DNA from tobacco to the soil bacterium Acinetobacter sp. by natural transformation. Mol. Microbiol. 53: 323–334
Demanèche, S, Kay, E, Gourbière, F, Simonet, P (2001) Natural transformation of Pseudomonas fluorescens and Agrobacterium tumefacien in soil. Appl. Environ. Microbiol. 67: 2617–2621 CrossRef
Dubnau D (1999) DNA uptake in bacteria. Annu. Rev. Microbiol. 53: 17–44
Gebhard, F, Smalla, K (1998) Transformation of Acinetobacter sp. strain BD 413 by transgenic sugar beet DNA. Appl. Environ. Microbiol. 64: 1550–1554
Gebhard F, Smalla K (1999) Monitoring field releases of genetically modified sugar beets for persistence of transgenic plant DNA and horizontal gene transfer. FEMS Microbiol. Ecol. 28: 261–272
Götz A, Pukall R, Tietze E, Prager R, Tschäpe H, van Elsas JD, Smalla K (1996) Detection and characterisation of broad-host-range plasmids in environmental bacteria by PCR. Appl. Environ. Microbiol. 62: 2621–2628
Haagensen, JAJ, Hansen, SK, Johansen, T, Molin, S (2002) In situ detection of horizontal transfer of mobile genetic elements. FEMS Microbiol Ecol 42: 261–268 CrossRef
Kay, E, Vogel, TM, Bertolla, F, Nalin, R, Simonet, P (2002) In situ transfer of antibiotic resistance genes from transgenic (transplastomic) tobacco plants to bacteria. Appl. Environ. Microbiol. 68: 3345–3351 CrossRef
Lorenz, MG, Wackernagel, W (1994) Bacterial gene transfer by natural genetic transformation in the environment. Microbiol. Rev. 58: 563–602
Maamar, H, Dubnau, D (2005) Bistability in the Bacillus subtilis K-state (competence) system requires a positive feedback loop. Mol. Microbiol. 56: 615–624 CrossRef
Meier P, Wackernagel W (2003) Mechanisms of homology-facilitated illegitimate recombination for foreign DNA acquisition in transformable Pseudomonas stutzeri. Mol. Microbiol. 48: 1107–1118
Milling, A, Smalla, K, Maidl, FX, Schloter, M, Munch, JC (2004) Effects of transgenic potatoes with an altered starch composition on the diversity of soil and rhizosphere bacteria and fungi. Plant Soil 266: 23–39 CrossRef
Nielsen KM, van Weerelt MD, Berg TN, Bones AM, Hagler AN, van Elsas JD (1997a) Natural transformation and availability of transforming DNA to Acinetobacter calcoaceticus in soil microcosms. Appl. Environ. Microbiol. 63: 1945–1952
Nielsen, KM, Bones, AM, van Elsas, JD (1997b) Induced natural transformation of Acinetobacter calcoaceticus in soil microcosms. Appl. Environ. Microbiol. 63: 3972–3977
Nielsen, KM, Bones, AM, Smalla, K, van Elsas, JD (1998) Horizontal gene transfer from transgenic plants to terrestrial bacteria - a rare event? FEMS Microbiol. Rev. 22: 79–103 CrossRef
Nielsen KM, Smalla K, van Elsas JD (2000a) Natural transformation of Acinetobacter sp. strain BD413 with cell lysates of Acinetobacter sp., Pseudomonas fluorescens and Burkholderia cepacia in soil microcosms. Appl. Environ. Microbiol. 66: 206–212
Nielsen, KM, van Elsas, JD, Smalla, K (2000b) Transformation of Acinetobacter sp. strain BD413 (pFG4 $\Delta $ nptII) with transgenic plant DNA in soil microcosms and effects of kanamycin on selection of transformants. Appl. Environ. Microbiol. 66: 1237–1242 CrossRef
Palmen R, Vosman B, Buijsman P, Breek CKD, Hellingwerf KJ (1993) Physiological characterization of natural transformation in Acinetobacter calcoaceticus. J. Gen. Microbiol. 139: 295–305
Rademaker JLW, de Bruijn FJ (1997) Characterization and classification of microbes by REP-PCR genomic fingerprinting and computer-assisted pattern analysis. In Gaetano-Anolles G, Gresshoff PM, eds, DNA markers: Protocols, Applications and Overviews, Wiley & Sons, Inc., New York, pp 151–171
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
Sikorski, J, Teschner, N, Wackernagel, W (2002) Highly different levels of natural transformation are associated with genomic subgroups within a local population of Pseudomonas stutzeri from soil. Appl. Environ. Microbiol. 68: 865–873 CrossRef
van den Eede, G, Aarts, H, Buhk, H-J, Corthier, G, Flint, HJ, Hammes, W, Jacobsen, B, Midtvedt, T, Van der Vossen, J, von Wright, A, Wackernagel, W, Wilcks, A (2004) The relevance of gene transfer to the safety of food and feed derived from genetically modified (GM) plants. Food Chem. Toxicol. 42: 1127–1156 CrossRef
Vaneechoutte, M, Young, DM, Ornston, LN, de Baere, T, Nemec, A, Van der Reijden, T, Carr, E, Tjernberg, I, Dijkshoorn, L (2006) Naturally transformable Acinetobacter sp. strain ADP1 belongs to the newly described species Acinetobacter baylyi. Appl. Environ. Microbiol. 72: 932–936 CrossRef