scholarly journals Role of DLP12 lysis genes in Escherichia coli biofilm formation

Microbiology ◽  
2011 ◽  
Vol 157 (6) ◽  
pp. 1640-1650 ◽  
Author(s):  
Faustino A. Toba ◽  
Mitchell G. Thompson ◽  
Bryan R. Campbell ◽  
Lauren M. Junker ◽  
Karl-Gustav Rueggeberg ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Wall ◽  
Biofilm Formation ◽  
Wall Stress ◽  
Wild Type ◽  
Lambda Phage ◽  
Kinase Gene ◽  
Abiotic Surfaces ◽  
K 12 ◽  
Biofilm Development

Phages have recently been implicated as important in biofilm development, although the mechanisms whereby phages impact biofilms remain unclear. One defective lambdoid phage carried by Escherichia coli K-12 is DLP12. Among the genes found in DLP12 are essD, ybcS and rzpD/rzoD, which are homologues of the Lambda phage genes encoding cell-lysis proteins (S, R and Rz/Rz1). The role that these DLP12 lysis genes play in biofilm formation was examined in deletion mutants of E. coli PHL628, a curli-overproducing, biofilm-forming K-12 derivative. Strains lacking essD, ybcS and rzpD/rzoD were unable to form wild-type biofilms. While all mutants were compromised in attachment to abiotic surfaces and aggregated less well than the wild-type, the effect of the essD knockout on biofilm formation was less dramatic than that of deleting ybcS or rzpD/rzoD. These results were consistent with electron micrographs of the mutants, which showed a decreased number of curli fibres on cell surfaces. Also consistent with this finding, we observed that expression from the promoter of csgB, which encodes the curli subunits, was downregulated in the mutants. As curli production is transcriptionally downregulated in response to cell wall stress, we challenged the mutants with SDS and found them to be more sensitive to the detergent than the wild-type. We also examined the release of 14C-labelled peptidoglycan from the mutants and found that they did not lose labelled peptidoglycan to the same extent as the wild-type. Given that curli production is known to be suppressed by N-acetylglucosamine 6-phosphate (NAG-6P), a metabolite produced during peptidoglycan recycling, we deleted nagK, the N-acetylglucosamine kinase gene, from the lysis mutants and found that this restored curli production. This suggested that deletion of the lysis genes affected cell wall status, which was transduced to the curli operon by NAG-6P via an as yet unknown mechanism. These observations provide evidence that the S, R and Rz/Rz1 gene homologues encoded by DLP12 are not merely genetic junk, but rather play an important, though undefined, role in cell wall maintenance.

scholarly journals Mat fimbriae promote biofilm formation by meningitis-associated Escherichia coli

Microbiology ◽  
2010 ◽  
Vol 156 (8) ◽  
pp. 2408-2417 ◽  
Author(s):  
Timo A. Lehti ◽  
Philippe Bauchart ◽  
Johanna Heikkinen ◽  
Jörg Hacker ◽  
Timo K. Korhonen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Early Stage ◽  
Surface Expression ◽  
Growth Media ◽  
Biofilm Growth ◽  
Abiotic Surfaces ◽  
K 12 ◽  
Biofilm Development

The mat (or ecp) fimbrial operon is ubiquitous and conserved in Escherichia coli, but its functions remain poorly described. In routine growth media newborn meningitis isolates of E. coli express the meningitis-associated and temperature-regulated (Mat) fimbria, also termed E. coli common pilus (ECP), at 20 °C, and here we show that the six-gene (matABCDEF)-encoded Mat fimbria is needed for temperature-dependent biofilm formation on abiotic surfaces. The matBCDEF deletion mutant of meningitis E. coli IHE 3034 was defective in an early stage of biofilm development and consequently unable to establish a detectable biofilm, contrasting with IHE 3034 derivatives deleted for flagella, type 1 fimbriae or S-fimbriae, which retained the wild-type biofilm phenotype. Furthermore, induced production of Mat fimbriae from expression plasmids enabled biofilm-deficient E. coli K-12 cells to form biofilm at 20 °C. No biofilm was detected with IHE 3034 or MG1655 strains grown at 37 °C. The surface expression of Mat fimbriae and the frequency of Mat-positive cells in the IHE 3034 population from 20 °C were high and remained unaltered during the transition from planktonic to biofilm growth and within the matured biofilm community. Considering the prevalence of the highly conserved mat locus in E. coli genomes, we hypothesize that Mat fimbria-mediated biofilm formation is an ancestral characteristic of E. coli.

scholarly journals Bundle-Forming Pili and EspA Are Involved in Biofilm Formation by Enteropathogenic Escherichia coli

2006 ◽  
Vol 188 (11) ◽  
pp. 3952-3961 ◽  
Author(s):  
Cristiano G. Moreira ◽  
Kelli Palmer ◽  
Marvin Whiteley ◽  
Marcelo P. Sircili ◽  
Luiz R. Trabulsi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Signaling Mechanism ◽  
Bacterial Aggregation ◽  
Abiotic Surfaces ◽  
Genes Encoding ◽  
Continuous Culture System ◽  
Static Conditions ◽  
Biofilm Development ◽  
Isogenic Mutants

ABSTRACT Microcolony formation is one of the initial steps in biofilm development, and in enteropathogenic Escherichia coli (EPEC) it is mediated by several adhesins, including the bundle-forming pilus (BFP) and the EspA filament. Here we report that EPEC forms biofilms on plastic under static conditions and a flowthrough continuous culture system. The abilities of several EPEC isogenic mutants to form biofilms were assessed. Adhesins such as BFP and EspA, important in microcolony formation on epithelial cells, are also involved in bacterial aggregation during biofilm formation on abiotic surfaces. Mutants that do not express BFP or EspA form more-diffuse biofilms than does the wild type. We also determined, using gfp transcriptional fusions, that, consistent with the role of these adhesins in biofilms, the genes encoding BFP and EspA are expressed during biofilm formation. Finally, expression of espA is controlled by a quorum-sensing (QS) regulatory mechanism, and the EPEC qseA QS mutant also forms altered biofilms, suggesting that this signaling mechanism plays an important role in EPEC biofilm development. Taken together, these studies allowed us to propose a model of EPEC biofilm formation.

scholarly journals The Transcriptional Antiterminator RfaH Represses Biofilm Formation in Escherichia coli

2006 ◽  
Vol 188 (4) ◽  
pp. 1316-1331 ◽  
Author(s):  
Christophe Beloin ◽  
Kai Michaelis ◽  
Karin Lindner ◽  
Paolo Landini ◽  
Jörg Hacker ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Wild Type Strain ◽  
Wild Type ◽  
Strain Mg1655 ◽  
Upec Strain ◽  
E Coli ◽  
Initial Adhesion ◽  
State Production ◽  
K 12

ABSTRACT We investigated the influence of regulatory and pathogenicity island-associated factors (Hha, RpoS, LuxS, EvgA, RfaH, and tRNA5 Leu) on biofilm formation by uropathogenic Escherichia coli (UPEC) strain 536. Only inactivation of rfaH, which encodes a transcriptional antiterminator, resulted in increased initial adhesion and biofilm formation by E. coli 536. rfaH inactivation in nonpathogenic E. coli K-12 isolate MG1655 resulted in the same phenotype. Transcriptome analysis of wild-type strain 536 and an rfaH mutant of this strain revealed that deletion of rfaH correlated with increased expression of flu orthologs. flu encodes antigen 43 (Ag43), which mediates autoaggregation and biofilm formation. We confirmed that deletion of rfaH leads to increased levels of flu and flu-like transcripts in E. coli K-12 and UPEC. Supporting the hypothesis that RfaH represses biofilm formation through reduction of the Ag43 level, the increased-biofilm phenotype of E. coli MG1655rfaH was reversed upon inactivation of flu. Deletion of the two flu orthologs, however, did not modify the behavior of mutant 536rfaH. Our results demonstrate that the strong initial adhesion and biofilm formation capacities of strain MG1655rfaH are mediated by both increased steady-state production of Ag43 and likely increased Ag43 presentation due to null rfaH-dependent lipopolysaccharide depletion. Although the roles of rfaH in the biofilm phenotype are different in UPEC strain 536 and K-12 strain MG1655, this study shows that RfaH, in addition to affecting the expression of bacterial virulence factors, also negatively controls expression and surface presentation of Ag43 and possibly another Ag43-independent factor(s) that mediates cell-cell interactions and biofilm formation.

scholarly journals O-Mannosylation in Candida albicans Enables Development of Interkingdom Biofilm Communities

mBio ◽  
2014 ◽  
Vol 5 (2) ◽  
Author(s):  
Lindsay C. Dutton ◽  
Angela H. Nobbs ◽  
Katy Jepson ◽  
Mark A. Jepson ◽  
M. Margaret Vickerman ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Biofilm Formation ◽  
Early Stage ◽  
Growth Conditions ◽  
Wild Type ◽  
Content Type ◽  
Subsequent Performance ◽  
Biofilm Development ◽  
Mutant Cells

ABSTRACTCandida albicansis a fungus that colonizes oral cavity surfaces, the gut, and the genital tract.Streptococcus gordoniiis a ubiquitous oral bacterium that has been shown to form biofilm communities withC. albicans. Formation of dual-speciesS. gordonii-C. albicansbiofilm communities involves interaction of theS. gordoniiSspB protein with the Als3 protein on the hyphal filament surface ofC. albicans. Mannoproteins comprise a major component of theC. albicanscell wall, and in this study we sought to determine if mannosylation in cell wall biogenesis ofC. albicanswas necessary for hyphal adhesin functions associated with interkingdom biofilm development. AC. albicans mnt1Δmnt2Δ mutant, with deleted α-1,2-mannosyltransferase genes and thus defective inO-mannosylation, was abrogated in biofilm formation under various growth conditions and produced hyphal filaments that were not recognized byS. gordonii. Cell wall proteomes of hypha-formingmnt1Δmnt2Δ mutant cells showed growth medium-dependent alterations, compared to findings for the wild type, in a range of protein components, including Als1, Als3, Rbt1, Scw1, and Sap9. Hyphal filaments formed bymnt1Δmnt2Δ mutant cells, unlike wild-type hyphae, did not interact withC. albicansAls3 or Hwp1 partner cell wall proteins or withS. gordoniiSspB partner adhesin, suggesting defective functionality of adhesins on themnt1Δmnt2Δ mutant. These observations imply that early stageO-mannosylation is critical for activation of hyphal adhesin functions required for biofilm formation, recognition by bacteria such asS. gordonii, and microbial community development.IMPORTANCEIn the human mouth, microorganisms form communities known as biofilms that adhere to the surfaces present.Candida albicansis a fungus that is often found within these biofilms. We have focused on the mechanisms by whichC. albicansbecomes incorporated into communities containing bacteria, such asStreptococcus. We find that impairment of early stage addition of mannose sugars toC. albicanshyphal filament proteins deleteriously affects their subsequent performance in mediating formation of polymicrobial biofilms. Our analyses provide new understanding of the way that microbial communities develop, and of potential means to controlC. albicansinfections.

scholarly journals Biofilm Formation and Dispersal under the Influence of the Global Regulator CsrA of Escherichia coli

2002 ◽  
Vol 184 (1) ◽  
pp. 290-301 ◽  
Author(s):  
Debra W. Jackson ◽  
Kazushi Suzuki ◽  
Lawrence Oakford ◽  
Jerry W. Simecka ◽  
Mark E. Hart ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Rna Binding ◽  
Glycogen Synthase ◽  
Regulatory Protein ◽  
Ectopic Expression ◽  
Knockout Mutant ◽  
E Coli ◽  
K 12 ◽  
Biofilm Development

ABSTRACT The predominant mode of growth of bacteria in the environment is within sessile, matrix-enclosed communities known as biofilms. Biofilms often complicate chronic and difficult-to-treat infections by protecting bacteria from the immune system, decreasing antibiotic efficacy, and dispersing planktonic cells to distant body sites. While the biology of bacterial biofilms has become a major focus of microbial research, the regulatory mechanisms of biofilm development remain poorly defined and those of dispersal are unknown. Here we establish that the RNA binding global regulatory protein CsrA (carbon storage regulator) of Escherichia coli K-12 serves as both a repressor of biofilm formation and an activator of biofilm dispersal under a variety of culture conditions. Ectopic expression of the E. coli K-12 csrA gene repressed biofilm formation by related bacterial pathogens. A csrA knockout mutation enhanced biofilm formation in E. coli strains that were defective for extracellular, surface, or regulatory factors previously implicated in biofilm formation. In contrast, this csrA mutation did not affect biofilm formation by a glgA (glycogen synthase) knockout mutant. Complementation studies with glg genes provided further genetic evidence that the effects of CsrA on biofilm formation are mediated largely through the regulation of intracellular glycogen biosynthesis and catabolism. Finally, the expression of a chromosomally encoded csrA′-′lacZ translational fusion was dynamically regulated during biofilm formation in a pattern consistent with its role as a repressor. We propose that global regulation of central carbon flux by CsrA is an extremely important feature of E. coli biofilm development.

scholarly journals Role ofrpoSin Escherichia coli O157:H7 Strain H32 Biofilm Development and Survival

2012 ◽  
Vol 78 (23) ◽  
pp. 8331-8339 ◽  
Author(s):  
Jessica R. Sheldon ◽  
Mi-Sung Yim ◽  
Jessica H. Saliba ◽  
Wai-Hong Chung ◽  
Kwok-Yin Wong ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Lake Water ◽  
Biofilm Formation ◽  
Survival Rates ◽  
Confocal Scanning Laser Microscopy ◽  
Wild Type ◽  
Minimal Growth ◽  
Content Type ◽  
Biofilm Development

ABSTRACTThe protein RpoS is responsible for mediating cell survival during the stationary phase by conferring cell resistance to various stressors and has been linked to biofilm formation. In this study, the role of therpoSgene inEscherichia coliO157:H7 biofilm formation and survival in water was investigated. Confocal scanning laser microscopy of biofilms established on coverslips revealed a nutrient-dependent role ofrpoSin biofilm formation, where the biofilm biomass volume of therpoSmutant was 2.4- to 7.5-fold the size of itsrpoS+wild-type counterpart in minimal growth medium. The enhanced biofilm formation of therpoSmutant did not, however, translate to increased survival in sterile double-distilled water (ddH2O), filter-sterilized lake water, or unfiltered lake water. TherpoSmutant had an overall reduction of 3.10 and 5.30 log10in sterile ddH2O and filter-sterilized lake water, respectively, while only minor reductions of 0.53 and 0.61 log10in viable counts were observed for the wild-type form in the two media over a 13-day period, respectively. However, the survival rates of the detached biofilm-derivedrpoS+andrpoSmutant cells were comparable. Under the competitive stress conditions of unfiltered lake water, the advantage conferred by the presence ofrpoSwas lost, and both the wild-type and knockout forms displayed similar declines in viable counts. These results suggest thatrpoSdoes have an influence on both biofilm formation and survival ofE. coliO157:H7 and that the advantage conferred byrpoSis contingent on the environmental conditions.

scholarly journals Chaperone DnaJ Influences the Formation of Biofilm by Escherichia coli

2015 ◽  
Vol 64 (3) ◽  
pp. 279-283 ◽  
Author(s):  
Anna Grudniak ◽  
Jolanta Włodkowska ◽  
Krystyna Wolska
Keyword(s):  
Escherichia Coli ◽  
Heat Shock ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Cell Physiology ◽  
Wild Type ◽  
Type Gene ◽  
Biofilm Development

DnaJ chaperone, a member of the so called DnaK-DnaJ-GrpE chaperone machine plays an important role in cell physiology. The ability of Escherichia coli ΔdnaJ mutant to form biofilm was studied. It was shown that this mutant is impaired in biofilm development when exposed to 42°C for 2 h. The impairment in biofilm development was observed when the heat shock was applied either at the onset of biofilm formation or 2 h later. The biofilm formed was thinner and its structure was changed as compared to wild-type strain. This defect could be complemented by the introduction of a wild-type gene on a low-copy plasmid.

scholarly journals Escherichia coli Biofilms Have an Organized and Complex Extracellular Matrix Structure

mBio ◽  
2013 ◽  
Vol 4 (5) ◽  
Author(s):  
Chia Hung ◽  
Yizhou Zhou ◽  
Jerome S. Pinkner ◽  
Karen W. Dodson ◽  
Jan R. Crowley ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Extracellular Matrix ◽  
Biofilm Formation ◽  
Developmental Stages ◽  
Three Dimensional ◽  
Bacterial Biofilms ◽  
Content Type ◽  
Abiotic Surfaces ◽  
Biochemical Analyses ◽  
Biofilm Development

ABSTRACTBacterial biofilms are ubiquitous in nature, and their resilience is derived in part from a complex extracellular matrix that can be tailored to meet environmental demands. Although common developmental stages leading to biofilm formation have been described, how the extracellular components are organized to allow three-dimensional biofilm development is not well understood. Here we show that uropathogenicEscherichia coli(UPEC) strains produce a biofilm with a highly ordered and complex extracellular matrix (ECM). We used electron microscopy (EM) techniques to image floating biofilms (pellicles) formed by UPEC. EM revealed intricately constructed substructures within the ECM that encase individual, spatially segregated bacteria with a distinctive morphology. Mutational and biochemical analyses of these biofilms confirmed curli as a major matrix component and revealed important roles for cellulose, flagella, and type 1 pili in pellicle integrity and ECM infrastructure. Collectively, the findings of this study elucidated that UPEC pellicles have a highly organized ultrastructure that varies spatially across the multicellular community.IMPORTANCEBacteria can form biofilms in diverse niches, including abiotic surfaces, living cells, and at the air-liquid interface of liquid media. Encasing these cellular communities is a self-produced extracellular matrix (ECM) that can be composed of proteins, polysaccharides, and nucleic acids. The ECM protects biofilm bacteria from environmental insults and also makes the dissolution of biofilms very challenging. As a result, formation of biofilms within humans (during infection) or on industrial material (such as water pipes) has detrimental and costly effects. In order to combat bacterial biofilms, a better understanding of components required for biofilm formation and the ECM is required. This study defined the ECM composition and architecture of floating pellicle biofilms formed byEscherichia coli.

scholarly journals YliH (BssR) and YceP (BssS) Regulate Escherichia coli K-12 Biofilm Formation by Influencing Cell Signaling

2006 ◽  
Vol 72 (4) ◽  
pp. 2449-2459 ◽  
Author(s):  
Joanna Domka ◽  
Jintae Lee ◽  
Thomas K. Wood
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Cell Signaling ◽  
Biofilm Formation ◽  
Differentially Expressed ◽  
Wild Type ◽  
Autoinducer 2 ◽  
Expression Of Genes ◽  
Differential Gene ◽  
K 12

ABSTRACT We previously discovered that yliH and yceP are induced in Escherichia coli biofilms (D. Ren, L. A. Bedzyk, S. M. Thomas, R. W. Ye, and T. K. Wood, Appl. Microbiol. Biotechnol. 64:515-524, 2004). Here, it is shown that deletion of yceP (b1060) and yliH (b0836) increases biofilm formation in continuous-flow chambers with minimal glucose medium by increasing biofilm mass (240- to 290-fold), surface coverage (16- to 31-fold), and mean thickness (2,800-fold). To determine the genetic basis of the increase in biofilm formation, we examined the differential gene expression profile in biofilms for both the mutants relative to the wild-type strain in rich medium with glucose and found that 372 to 882 genes were induced and that 76 to 337 were repressed consistently >2-fold (P ≤ 0.05). The increase in biofilm formation was related to differential expression of genes related to stress response (8 to 64 genes) for both mutants, including rpoS and sdiA. More importantly, 42 to 130 genes related to autoinducer 2 cell signaling were also differentially expressed, including gadAB and flgBCEGHIJLMN, as well as signaling through indole, since 17 to 26 indole-related genes were differentially expressed, including phoAER, gltBD, mtr (encodes protein for indole import), and acrEF (encodes proteins for indole export). Increased biofilm formation in the yliH and yceP mutants in LB supplemented with 0.2% glucose (LB glu) occurred through a reduction in extracellular and intracellular indole concentrations in both mutants (50- to 140-fold), and the addition of indole to the culture restored the wild-type biofilm phenotype; hence, indole represses biofilms. Additionally, both mutants regulate biofilms through quorum sensing, since deletion of either yliH or yceP increased extracellular autoinducer 2 concentrations 50-fold when grown in complex medium (most notably in the stationary phase). Both proteins are involved in motility regulation, since YliH (127 amino acids) and YceP (84 amino acids) repressed motility two to sevenfold (P ≤ 0.05) in LB, and YceP repressed motility sevenfold (P ≤ 0.05) in LB glu. Heightened motility in the yceP mutant occurred, due to increased transcription of the flagella and motility loci, including fliC, motA, and qseB (3- to 86-fold). We propose new names for these two loci: bssR for yliH and bssS for yceP, based on the phrase “regulator of biofilm through signal secretion.”

scholarly journals Autoinducer 2 Controls Biofilm Formation in Escherichia coli through a Novel Motility Quorum-Sensing Regulator (MqsR, B3022)

2006 ◽  
Vol 188 (1) ◽  
pp. 305-316 ◽  
Author(s):  
Andrés F. González Barrios ◽  
Rongjun Zuo ◽  
Yoshifumi Hashimoto ◽  
Li Yang ◽  
William E. Bentley ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Continuous Flow ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Flow Cells ◽  
Autoinducer 2 ◽  
K 12

ABSTRACT The cross-species bacterial communication signal autoinducer 2 (AI-2), produced by the purified enzymes Pfs (nucleosidase) and LuxS (terminal synthase) from S-adenosylhomocysteine, directly increased Escherichia coli biofilm mass 30-fold. Continuous-flow cells coupled with confocal microscopy corroborated these results by showing the addition of AI-2 significantly increased both biofilm mass and thickness and reduced the interstitial space between microcolonies. As expected, the addition of AI-2 to cells which lack the ability to transport AI-2 (lsr null mutant) failed to stimulate biofilm formation. Since the addition of AI-2 increased cell motility through enhanced transcription of five motility genes, we propose that AI-2 stimulates biofilm formation and alters its architecture by stimulating flagellar motion and motility. It was also found that the uncharacterized protein B3022 regulates this AI-2-mediated motility and biofilm phenotype through the two-component motility regulatory system QseBC. Deletion of b3022 abolished motility, which was restored by expressing b3022 in trans. Deletion of b3022 also decreased biofilm formation significantly, relative to the wild-type strain in three media (46 to 74%) in 96-well plates, as well as decreased biomass (8-fold) and substratum coverage (19-fold) in continuous-flow cells with minimal medium (growth rate not altered and biofilm restored by expressing b3022 in trans). Deleting b3022 changed the wild-type biofilm architecture from a thick (54-μm) complex structure to one that contained only a few microcolonies. B3022 positively regulates expression of qseBC, flhD, fliA, and motA, since deleting b3022 decreased their transcription by 61-, 25-, 2.4-, and 18-fold, respectively. Transcriptome analysis also revealed that B3022 induces crl (26-fold) and flhCD (8- to 27-fold). Adding AI-2 (6.4 μM) increased biofilm formation of wild-type K-12 MG1655 but not that of the isogenic b3022, qseBC, fliA, and motA mutants. Adding AI-2 also increased motA transcription for the wild-type strain but did not stimulate motA transcription for the b3022 and qseB mutants. Together, these results indicate AI-2 induces biofilm formation in E. coli through B3022, which then regulates QseBC and motility; hence, b3022 has been renamed the motility quorum-sensing regulator gene (the mqsR gene).

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