Development of engineered genomic DNA to monitor the natural transformation ofPseudomonas stutzeriin soil-like microcosms

1997 ◽  
Vol 43 (1) ◽  
pp. 78-84 ◽  
Author(s):  
Eric Paget ◽  
Pascal Simonet
Keyword(s):  
Humic Acids ◽  
Genomic Dna ◽  
Natural Transformation ◽  
Pseudomonas Stutzeri ◽  
Viable Cell ◽  
Extracellular Dna ◽  
Sterile Soil ◽  
Soil Matrix ◽  
Soil Microcosms ◽  
Suicide Plasmid

The goal of this paper was to demonstrate whether natural transformation could occur in the environment to promote horizontal gene transfer between bacteria. Microcosms consisting of clay, clay and humic acids, or sterile soil were compared with respect to the natural transformation of Pseudomonas stutzeri by mineral-adsorbed DNA. Genes conferring resistance to tetracycline and ampicillin were first inserted in P. stutzeri pp100 chromosome via the pSUP202 suicide plasmid. Then, DNA extracted from the engineered P. stutzeri strain was used for transformation experiments, allowing the new transformed cells to be detected by hybridization with a tet probe. It turned out that DNA adsorbed on clay or soil particles and in presence of humic acids still transformed competent cells with frequencies up to 10−8transformants/viable cell. Finally, natural transformation assays involving two different DNAs were carried out in sterile soil microcosms. The use of nonisogenic DNA extracted from a rifampicin-resistant Pseudomonas fluorescens strain resulted in production of transformants, while isogenic DNA from our engineered strain failed to produce any. These observations confirmed that extracellular DNA adsorbed on a soil matrix composed of minerals and organic matter could still transform competent bacteria under environmental conditions.Key words: transformation, Pseudomonas stutzeri, soil microcosm, DNA, suicide plasmid.

scholarly journals Natural Transformation of Pseudomonas fluorescens and Agrobacterium tumefaciens in Soil

2001 ◽  
Vol 67 (6) ◽  
pp. 2617-2621 ◽  
Author(s):  
Sandrine Demanèche ◽  
Elisabeth Kay ◽  
François Gourbière ◽  
Pascal Simonet
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Pseudomonas Fluorescens ◽  
Natural Transformation ◽  
Physiological State ◽  
Extracellular Dna ◽  
Soil Microcosms ◽  
Do So

ABSTRACT Little information is available concerning the occurrence of natural transformation of bacteria in soil, the frequency of such events, and the actual role of this process on bacterial evolution. This is because few bacteria are known to possess the genes required to develop competence and because the tested bacteria are unable to reach this physiological state in situ. In this study we found that two soil bacteria, Agrobacterium tumefaciens and Pseudomonas fluorescens, can undergo transformation in soil microcosms without any specific physical or chemical treatment. Moreover, P. fluorescens produced transformants in both sterile and nonsterile soil microcosms but failed to do so in the various in vitro conditions we tested. A. tumefaciens could be transformed in vitro and in sterile soil samples. These results indicate that the number of transformable bacteria could be higher than previously thought and that these bacteria could find the conditions necessary for uptake of extracellular DNA in soil.

scholarly journals Changes in natural transformation after salt adaptation

2021 ◽  
Author(s):  
HA Kittredge ◽  
SE Evans
Keyword(s):  
Growth Rate ◽  
Driving Force ◽  
Transformation Efficiency ◽  
Natural Transformation ◽  
Pseudomonas Stutzeri ◽  
Serial Passage ◽  
Extracellular Dna ◽  
Salt Adaptation ◽  
Serial Transfer ◽  
Evolutionary Origins

AbstractThe exchange of genes between potentially unrelated bacteria is termed horizontal gene transfer (HGT) and is a driving force in bacterial evolution. Natural transformation is one mechanism of HGT where extracellular DNA (eDNA) from the environment is recombined into a host genome. The widespread conservation of transformation in bacterial lineages implies there is a fitness benefit. However, the nature of these benefits and the evolutionary origins of transformation are still unknown. Here, I examine how ∼330 generations or 100 days of serial passage in either constant or increasing salinities impacts the growth rate and transformation efficiency of Pseudomonas stutzeri. While the growth rate generally improved in response to serial transfer, the transformation efficiency of the evolved lineages varied extensively, with only 39-64% of populations undergoing transformation at the end of adaptive evolution. In comparison, 100% of the ancestral populations were able to undergo natural transformation. I also found that evolving P. stutzeri with different cell lysates (or populations of dead cells) minimally affected the growth rate and transformation efficiency, especially in comparison to the pervasiveness with which transformation capacity was lost across the evolved populations. Taken together, I show that the efficiency of eDNA uptake changes over relatively rapid timescales, suggesting that transformation is an adaptive and selectable trait that could be lost in environments where it is not beneficial.

scholarly journals Biology of Pseudomonas stutzeri

2006 ◽  
Vol 70 (2) ◽  
pp. 510-547 ◽  
Author(s):  
Jorge Lalucat ◽  
Antoni Bennasar ◽  
Rafael Bosch ◽  
Elena García-Valdés ◽  
Norberto J. Palleroni
Keyword(s):  
Natural Transformation ◽  
Model Organism ◽  
Pseudomonas Stutzeri ◽  
Opportunistic Pathogen ◽  
Denitrifying Bacterium ◽  
The Past ◽  
History Of ◽  
Metabolic Properties ◽  
Degradation Of Pollutants ◽  
Made In

SUMMARY Pseudomonas stutzeri is a nonfluorescent denitrifying bacterium widely distributed in the environment, and it has also been isolated as an opportunistic pathogen from humans. Over the past 15 years, much progress has been made in elucidating the taxonomy of this diverse taxonomical group, demonstrating the clonality of its populations. The species has received much attention because of its particular metabolic properties: it has been proposed as a model organism for denitrification studies; many strains have natural transformation properties, making it relevant for study of the transfer of genes in the environment; several strains are able to fix dinitrogen; and others participate in the degradation of pollutants or interact with toxic metals. This review considers the history of the discovery, nomenclatural changes, and early studies, together with the relevant biological and ecological properties, of P. stutzeri.

scholarly journals Pseudomonas stutzeri and related species undergo natural transformation.

1983 ◽  
Vol 153 (1) ◽  
pp. 93-99 ◽  
Author(s):  
C A Carlson ◽  
L S Pierson ◽  
J J Rosen ◽  
J L Ingraham
Keyword(s):  
Related Species ◽  
Natural Transformation ◽  
Pseudomonas Stutzeri

scholarly journals Evaluation of Different Methods for Extracting Extracellular DNA from the Biofilm Matrix

2009 ◽  
Vol 75 (16) ◽  
pp. 5390-5395 ◽  
Author(s):  
Jianfeng Wu ◽  
Chuanwu Xi
Keyword(s):  
Chemical Treatment ◽  
Genomic Dna ◽  
Cell Lysis ◽  
Extraction Methods ◽  
Enzymatic Treatment ◽  
Extracellular Dna ◽  
Extracellular Matrices ◽  
Treatment Methods ◽  
Biofilm Development ◽  
Biofilm Matrix

ABSTRACT The occurrence of high concentrations of extracellular DNA (eDNA) in the extracellular matrices of biofilms plays an important role in biofilm formation and development and possibly in horizontal gene transfer through natural transformation. Studies have been conducted to characterize the nature of eDNA and its potential function in biofilm development, but it is difficult to extract eDNA from the extracellular matrices of biofilms without any contamination from genomic DNA released by cell lysis during the extraction process. In this report, we compared several different extraction methods in order to obtain highly pure eDNA from different biofilm samples. After different extraction methods were explored, it was concluded that using chemical treatment or enzymatic treatment of biofilm samples may obtain larger amounts of eDNA than using the simple filtration method. There was no detectable cell lysis when the enzymatic treatment methods were used, but substantial cell lysis was observed when the chemical treatment methods were used. These data suggest that eDNA may bind to other extracellular polymers in the biofilm matrix and that enzymatic treatment methods are effective and favorable for extracting eDNA from biofilm samples. Moreover, randomly amplified polymorphic DNA analysis of eDNA in Acinetobacter sp. biofilms and Acinetobacter sp. genomic DNA and DNA sequencing analysis revealed that eDNA originated from genomic DNA but was not structurally identical to the genomic DNA.

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