Tight Modulation of Escherichia coli Bacterial Biofilm Formation through Controlled Expression of Adhesion Factors

Abstract

Despite the economic and sanitary problems caused by harmful biofilms, biofilms are nonetheless used empirically in industrial environmental and bioremediation processes and may be of potential use in medical settings for interfering with pathogen development. Escherichia coli is one of the bacteria with which biofilm formation has been studied in great detail, and it is especially appreciated for biotechnology applications because of its genetic amenability. Here we describe the development of two new genetic tools enabling the constitutive and inducible expression of any gene or operon of interest at its native locus. In addition to providing valuable tools for complementation and overexpression experiments, these two compact genetic cassettes were used to modulate the biofilm formation capacities of E. coli by taking control of two biofilmpromoting factors, autotransported antigen 43 adhesin and the bscABZC cellulose operon. The modulation of the biofilm formation capacities of E. coli or those of other bacteria capable of being genetically manipulated may be of use both for reducing and for improving the impact of biofilms in a number of industrial and medical applications.Most natural and artificial surfaces available in the environment are prone to bacterial colonization. Following the initial adhesion event, cell-to-cell adhesion and the secretion of an extracellular matrix rapidly lead to the formation of a surfaceattached multicellular structure known as a biofilm (3,43,75,84). Besides being resistant to environmental shear forces, biofilm communities are also phenotypically more resistant to antibiotic and biocide treatments, a trait that poses important sanitary and economic problems. Indeed, biofilms formed by bacterial pathogens on medically relevant surfaces are difficult to eradicate and are thus often involved in the development of infections (12,13,48). Moreover, industrial biofouling resulting from bacterial biofilm formation is a major cause of pipe biocorrosion and reduces the efficiency of pharmaceutical or food bioprocesses (2,10,70,71).While recent studies have focused mainly on the negative impact of biofilms, bacterial biofilms can also have valuable applications, including those involving bioremediation and wastewater treatment bioreactor processes and the improvement of biomineralization or plant-bacteria symbiosis (1,14,40,42,52,62,73,74,80). Beneficial biofilms may also have medical applications, and the use of protective innocuous bacterial biofilms that interfere with the development of bacterial pathogens is considered a promising approach (19).Escherichia coli is a gram-negative enterobacterium that has been used extensively as a model to study biofilm development due to its relevance to the human biotic environment and its genetic amenability. In E. coli, various cell surface appendages were shown to be necessary to achieve mature biofilm development (78). Flagella, type I fimbriae, and curli are implicated in early adhesion steps, while the production of a polysaccharide-rich m...

Molecular Characterization of UpaB and UpaC, Two New Autotransporter Proteins of Uropathogenic Escherichia coli CFT073

“…5B). In addition, we detected surface localization of both UpaB and UpaC in UpaB/UpaC overexpression strains constructed by chromosomal insertion of the inducible RExBAD expression cassette and a P R constitutive promoter (13,55) in front of each open reading frame, respectively (CFT073RexBADupaB and CFT073RexBADupaC, CFT073PcLupaB and CFT073PcLupaC) (Fig. 5).…”

“…In order to interrupt the AT-encoding genes in CFT073, create lacZ reporter transcriptional fusions, and place chromosomal target genes under the control of the RExBAD promoter or PcL promoter (13,14,55), we used homologous recombination mediated by -red recombinase expressed from the pKOBEG plasmid (15) and either a one-step PCR procedure with 40-bp homology arms for recombination or a three-step PCR procedure with 500-bp homology arms for recombination (15) (http://www.pasteur.fr /recherche/unites/Ggb/matmet.html). All mutants were confirmed via PCR and sequencing using primers Gene.ext 5= and Gene.ext 3=, either as a primer set or individually in combination with internal primers designed to bind within the inserted DNA fragments.…”

UpaG, a New Member of the Trimeric Autotransporter Family of Adhesins in UropathogenicEscherichia coli

“…DNA sequencing was performed by MWG Services. The construction of mutants and upaG-controlled-expression strains (PcL and RExBAD) (15,57) was performed using a three-step PCR procedure described previously (36; http://www.pasteur.fr/recherche/unites/Ggb /matmet.html).…”

Biofilm-Producing Ability of Bovine Extraintestinal Pathogenic Escherichia coli and Its Correlation with Attachment Factors

“…Production of these bacteria on medically relevant surfaces may be difficult to eradicate, resulting in the stability of infection (4).…”