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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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Spatial gene expression of flhD in the ompR and rcsB mutant strains. (A) is the 3D image of the 33 h biofilm from BP1531 (ompR::Tn10 pPS71), (B) is the respective image from the 51 h biofilm from BP1532 (rcsB::Tn5 pKK12). (C) is the quantitative representation of the spatial gene expression of flhD in the ompR mutant (red line) and the rcsB mutant (orange line) at the times points represented in A and B.
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Figure 4: Spatial gene expression of flhD in the ompR and rcsB mutant strains. (A) is the 3D image of the 33 h biofilm from BP1531 (ompR::Tn10 pPS71), (B) is the respective image from the 51 h biofilm from BP1532 (rcsB::Tn5 pKK12). (C) is the quantitative representation of the spatial gene expression of flhD in the ompR mutant (red line) and the rcsB mutant (orange line) at the times points represented in A and B.

Mentions: The temporal gene expression study had determined that the expression of flhD in the ompR and rcsB mutant strains was constitutively high throughout the experiment after a primary increase during the initial time period of biofilm formation. As time points for the spatial experiment, we selected 33 h for the ompR mutant (Figure 4A) and 51 h for the rcsB mutant (Figure 4B). Interestingly, expression of flhD in both mutants was high across all layers of the biofilm. Fluorescence was between 80 and 95% coverage across the entire biofilm of both mutants (Figure 4C). By all appearances, both OmpR and RcsB abolished spatial differences in flhD expression together with temporal ones, while increasing overall expression.


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)

Spatial gene expression of flhD in the ompR and rcsB mutant strains. (A) is the 3D image of the 33 h biofilm from BP1531 (ompR::Tn10 pPS71), (B) is the respective image from the 51 h biofilm from BP1532 (rcsB::Tn5 pKK12). (C) is the quantitative representation of the spatial gene expression of flhD in the ompR mutant (red line) and the rcsB mutant (orange line) at the times points represented in A and B.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 4: Spatial gene expression of flhD in the ompR and rcsB mutant strains. (A) is the 3D image of the 33 h biofilm from BP1531 (ompR::Tn10 pPS71), (B) is the respective image from the 51 h biofilm from BP1532 (rcsB::Tn5 pKK12). (C) is the quantitative representation of the spatial gene expression of flhD in the ompR mutant (red line) and the rcsB mutant (orange line) at the times points represented in A and B.
Mentions: The temporal gene expression study had determined that the expression of flhD in the ompR and rcsB mutant strains was constitutively high throughout the experiment after a primary increase during the initial time period of biofilm formation. As time points for the spatial experiment, we selected 33 h for the ompR mutant (Figure 4A) and 51 h for the rcsB mutant (Figure 4B). Interestingly, expression of flhD in both mutants was high across all layers of the biofilm. Fluorescence was between 80 and 95% coverage across the entire biofilm of both mutants (Figure 4C). By all appearances, both OmpR and RcsB abolished spatial differences in flhD expression together with temporal ones, while increasing overall expression.

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