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OmpR and RcsB abolish temporal and spatial changes in expression of flhD in Escherichia coli biofilm.

Samanta P, Clark ER, Knutson K, Horne SM, Prüß BM - BMC Microbiol. (2013)

Bottom Line: Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well.This increase was paralleled by reductions in biofilm amounts at four tested time points.Our data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: Biofilms are communities of bacteria that are characterized by specific phenotypes, including an increased resistance towards anti-microbials and the host immune system. This calls for the development of novel biofilm prevention and treatment options to combat infectious disease. In Escherichia coli, numerous global regulators have been implicated in the control of biofilm associated cell surface organelles. These include the flagellar regulator FlhD/FlhC, the osmoregulator EnvZ/OmpR, and the colanic acid activator RcsCDB. Using flow cell technology and fluorescence microscopy, we determined the temporal expression from flhD::gfp, ompR::gfp, and rcsB::gfp in E. coli biofilm, as well as the impact of the negative regulation of flhD by OmpR and RcsB. Spatial gene expression was investigated from flhD::gfp.

Results: The temporal gene expression profile for flhD yielded an early peak at 12 h, a minimum of expression at 35 h, and a second increase in expression towards 51 h of biofilm development. In contrast, the ompR profile showed a peak at 35 h. A mutation in ompR abolished time dependence of flhD expression after the initial growth period of 12 h. Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well. Spatially, expression of flhD was highest in the outermost layer of the biofilm in the parent strain. In ompR and rcsB mutants, flhD was expressed throughout the biofilm. Mutations in both, ompR and rcsB increased flhD expression throughout all temporal and spatial experiments. This increase was paralleled by reductions in biofilm amounts at four tested time points.

Conclusion: Our data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

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Related in: MedlinePlus

Fluorescence images of flhD::gfp, ompR::gfp, rcsB::gfp in AJW678 and flhD in the ompR and rcsB mutant strains. Biofilms of BP1470, BP1432, BP1462, BP1531, and BP1532 were grown in flow cells and subjected to fluorescence microscopy. Four time points were selected for each strain; these are printed on top of the respective images. At the very top of each column, promoter names are printed. Images were taken at 1,000 fold magnification.
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Figure 1: Fluorescence images of flhD::gfp, ompR::gfp, rcsB::gfp in AJW678 and flhD in the ompR and rcsB mutant strains. Biofilms of BP1470, BP1432, BP1462, BP1531, and BP1532 were grown in flow cells and subjected to fluorescence microscopy. Four time points were selected for each strain; these are printed on top of the respective images. At the very top of each column, promoter names are printed. Images were taken at 1,000 fold magnification.

Mentions: Fluorescence microscopy images were produced from flow cell grown biofilm of the E. coli genetic parent strain AJW678 that contained the flhD::gfp fusion plasmid, called pPS71. Fluorescence signals obtained from these biofilms were highest at 12 h, lowest at 35 h, and then increased again towards 51 h of biofilm formation. This was seen in all four time series of images that had been taken from four independently formed biofilms. A selection of images from one of these experiments is shown in the left column of Figure 1. Occasionally, we observed high signals in individual bacteria of the 3 h sample, but the number of bacteria on the slides was not indicative of a biofilm at that point in time.


OmpR and RcsB abolish temporal and spatial changes in expression of flhD in Escherichia coli biofilm.

Samanta P, Clark ER, Knutson K, Horne SM, Prüß BM - BMC Microbiol. (2013)

Fluorescence images of flhD::gfp, ompR::gfp, rcsB::gfp in AJW678 and flhD in the ompR and rcsB mutant strains. Biofilms of BP1470, BP1432, BP1462, BP1531, and BP1532 were grown in flow cells and subjected to fluorescence microscopy. Four time points were selected for each strain; these are printed on top of the respective images. At the very top of each column, promoter names are printed. Images were taken at 1,000 fold magnification.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750693&req=5

Figure 1: Fluorescence images of flhD::gfp, ompR::gfp, rcsB::gfp in AJW678 and flhD in the ompR and rcsB mutant strains. Biofilms of BP1470, BP1432, BP1462, BP1531, and BP1532 were grown in flow cells and subjected to fluorescence microscopy. Four time points were selected for each strain; these are printed on top of the respective images. At the very top of each column, promoter names are printed. Images were taken at 1,000 fold magnification.
Mentions: Fluorescence microscopy images were produced from flow cell grown biofilm of the E. coli genetic parent strain AJW678 that contained the flhD::gfp fusion plasmid, called pPS71. Fluorescence signals obtained from these biofilms were highest at 12 h, lowest at 35 h, and then increased again towards 51 h of biofilm formation. This was seen in all four time series of images that had been taken from four independently formed biofilms. A selection of images from one of these experiments is shown in the left column of Figure 1. Occasionally, we observed high signals in individual bacteria of the 3 h sample, but the number of bacteria on the slides was not indicative of a biofilm at that point in time.

Bottom Line: Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well.This increase was paralleled by reductions in biofilm amounts at four tested time points.Our data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: Biofilms are communities of bacteria that are characterized by specific phenotypes, including an increased resistance towards anti-microbials and the host immune system. This calls for the development of novel biofilm prevention and treatment options to combat infectious disease. In Escherichia coli, numerous global regulators have been implicated in the control of biofilm associated cell surface organelles. These include the flagellar regulator FlhD/FlhC, the osmoregulator EnvZ/OmpR, and the colanic acid activator RcsCDB. Using flow cell technology and fluorescence microscopy, we determined the temporal expression from flhD::gfp, ompR::gfp, and rcsB::gfp in E. coli biofilm, as well as the impact of the negative regulation of flhD by OmpR and RcsB. Spatial gene expression was investigated from flhD::gfp.

Results: The temporal gene expression profile for flhD yielded an early peak at 12 h, a minimum of expression at 35 h, and a second increase in expression towards 51 h of biofilm development. In contrast, the ompR profile showed a peak at 35 h. A mutation in ompR abolished time dependence of flhD expression after the initial growth period of 12 h. Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well. Spatially, expression of flhD was highest in the outermost layer of the biofilm in the parent strain. In ompR and rcsB mutants, flhD was expressed throughout the biofilm. Mutations in both, ompR and rcsB increased flhD expression throughout all temporal and spatial experiments. This increase was paralleled by reductions in biofilm amounts at four tested time points.

Conclusion: Our data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

Show MeSH
Related in: MedlinePlus