scholarly journals Basis for the Essentiality of H-NS Family Members in Pseudomonas aeruginosa

2012 ◽  
Vol 194 (18) ◽  
pp. 5101-5109 ◽  
Author(s):  
Sandra Castang ◽  
Simon L. Dove
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Family Members ◽  
Type Iv Pili ◽  
Replicative Form ◽  
Present Evidence ◽  
Content Type ◽  
Type Iv ◽  
Genes Encoding ◽  
And Function

ABSTRACTMembers of the histone-like nucleoid-structuring (H-NS) family of proteins have been shown to play important roles in silencing gene expression and in nucleoid compaction. InPseudomonas aeruginosa, the two H-NS family members MvaT and MvaU are thought to bind the same AT-rich regions of the chromosome and function coordinately to control a common set of genes. Here we present evidence that the loss of both MvaT and MvaU cannot be tolerated because it results in the production of Pf4 phage that superinfect and kill cells or inhibit their growth. Using a ClpXP-based protein depletion system in combination with transposon mutagenesis, we identify mutants ofP. aeruginosathat can tolerate the depletion of MvaT in an ΔmvaUmutant background. Many of these mutants contain insertions in genes encoding components, assembly factors, or regulators of type IV pili or contain insertions in genes of the prophage Pf4. We demonstrate that cells that no longer produce type IV pili or that no longer produce the replicative form of the Pf4 genome can tolerate the loss of both MvaT and MvaU. Furthermore, we show that the loss of both MvaT and MvaU results in an increase in expression of Pf4 genes and that cells that cannot produce type IV pili are resistant to infection by Pf4 phage. Our findings suggest that type IV pili are the receptors for Pf4 phage and that the essential activities of MvaT and MvaU are to repress the expression of Pf4 genes.

scholarly journals Vfr Directly ActivatesexsATranscription To Regulate Expression of the Pseudomonas aeruginosa Type III Secretion System

2016 ◽  
Vol 198 (9) ◽  
pp. 1442-1450 ◽  
Author(s):  
Anne E. Marsden ◽  
Peter J. Intile ◽  
Kayley H. Schulmeyer ◽  
Ethan R. Simmons-Patterson ◽  
Mark L. Urbanowski ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Type Iii Secretion ◽  
Virulence Gene ◽  
Type Iv Pili ◽  
Global Regulator ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Type Iii ◽  
Type Iv

ABSTRACTThePseudomonas aeruginosacyclic AMP (cAMP)-Vfr system (CVS) is a global regulator of virulence gene expression. Regulatory targets include type IV pili, secreted proteases, and the type III secretion system (T3SS). The mechanism by which CVS regulates T3SS gene expression remains undefined. Single-cell expression studies previously found that only a portion of the cells within a population express the T3SS under inducing conditions, a property known as bistability. We now report that bistability is altered in avfrmutant, wherein a substantially smaller fraction of the cells express the T3SS relative to the parental strain. Since bistability usually involves positive-feedback loops, we tested the hypothesis that virulence factor regulator (Vfr) regulates the expression ofexsA. ExsA is the central regulator of T3SS gene expression and autoregulates its own expression. AlthoughexsAis the last gene of theexsCEBApolycistronic mRNA, we demonstrate that Vfr directly activatesexsAtranscription from a second promoter (PexsA) located immediately upstream ofexsA. PexsApromoter activity is entirely Vfr dependent. Direct binding of Vfr to a PexsApromoter probe was demonstrated by electrophoretic mobility shift assays, and DNase I footprinting revealed an area of protection that coincides with a putative Vfr consensus-binding site. Mutagenesis of that site disrupted Vfr binding and PexsApromoter activity. We conclude that Vfr contributes to T3SS gene expression through activation of the PexsApromoter, which is internal to the previously characterizedexsCEBAoperon.IMPORTANCEVfr is a cAMP-dependent DNA-binding protein that functions as a global regulator of virulence gene expression inPseudomonas aeruginosa. Regulation by Vfr allows for the coordinate production of related virulence functions, such as type IV pili and type III secretion, required for adherence to and intoxication of host cells, respectively. Although the molecular mechanism of Vfr regulation has been defined for many target genes, a direct link between Vfr and T3SS gene expression had not been established. In the present study, we report that Vfr directly controlsexsAtranscription, the master regulator of T3SS gene expression, from a newly identified promoter located immediately upstream ofexsA.

scholarly journals The Swarming Motility of Pseudomonas aeruginosa Is Blocked by Cranberry Proanthocyanidins and Other Tannin-Containing Materials

2011 ◽  
Vol 77 (9) ◽  
pp. 3061-3067 ◽  
Author(s):  
Che O'May ◽  
Nathalie Tufenkji
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Adhesion ◽  
Type Iv Pili ◽  
Hydrolyzable Tannin ◽  
Swarming Motility ◽  
Subsequent Formation ◽  
Content Type ◽  
Type Iv ◽  
Derivatives Of ◽  
Growth Experiments

ABSTRACTBacterial motility plays a key role in the colonization of surfaces by bacteria and the subsequent formation of resistant communities of bacteria called biofilms. Derivatives of cranberry fruit, predominantly condensed tannins called proanthocyanidins (PACs) have been reported to interfere with bacterial adhesion, but the effects of PACs and other tannins on bacterial motilities remain largely unknown. In this study, we investigated whether cranberry PAC (CPAC) and the hydrolyzable tannin in pomegranate (PG; punicalagin) affected the levels of motilities exhibited by the bacteriumPseudomonas aeruginosa. This bacterium utilizes flagellum-mediated swimming motility to approach a surface, attaches, and then further spreads via the surface-associated motilities designated swarming and twitching, mediated by multiple flagella and type IV pili, respectively. Under the conditions tested, both CPAC and PG completely blocked swarming motility but did not block swimming or twitching motilities. Other cranberry-containing materials and extracts of green tea (also rich in tannins) were also able to block or impair swarming motility. Moreover, swarming bacteria were repelled by filter paper discs impregnated with many tannin-containing materials. Growth experiments demonstrated that the majority of these compounds did not impair bacterial growth. When CPAC- or PG-containing medium was supplemented with surfactant (rhamnolipid), swarming motility was partially restored, suggesting that the effective tannins are in part acting by a rhamnolipid-related mechanism. Further support for this theory was provided by demonstrating that the agar surrounding tannin-induced nonswarming bacteria was considerably less hydrophilic than the agar area surrounding swarming bacteria. This is the first study to show that natural compounds containing tannins are able to blockP. aeruginosaswarming motility and that swarming bacteria are repelled by such compounds.

scholarly journals Secretin channel-interactors prevent antibiotic influx during type IV pili assembly in P. aeruginosa

2021 ◽  
Author(s):  
Hongbaek Cho ◽  
Oh Hyun Kwon ◽  
Joel W Sher ◽  
Bi-o Kim ◽  
You-Hee Cho
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Virulence Factors ◽  
Bacterial Pathogenesis ◽  
Type Iv Pili ◽  
Permeability Barrier ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Type Iv ◽  
Periplasmic Proteins ◽  
And Function

Type IV pili (T4P) are important virulence factors involved in host attachment and other aspects of bacterial pathogenesis. In Gram-negative bacteria, the T4P filament is polymerized from pilin subunits at the platform complex in the inner membrane (IM) and exits the outer membrane (OM) through the OM secretin channel. Although it is essential for T4P assembly and function, the OM secretin complexes can potentially impair the permeability barrier function of the OM and allow the entry of antibiotics and other toxic molecules. The mechanism by which Gram-negative bacteria prevent secretin-mediated OM leakage is currently not well understood. Here, we report a discovery of SlkA and SlkB (PA5122 and PA5123) that prevent permeation of several classes of antibiotics through the secretin channel of Pseudomonas aeruginosa type IV pili. We found these periplasmic proteins interact with the OM secretin complex and prevent toxic molecules from entering through the channel when there is a problem in the assembly of the T4P IM subcomplexes or when docking between the OM and IM complexes is defective.

scholarly journals ThePseudomonas aeruginosaPilSR Two-Component System Regulates Both Twitching and Swimming Motilities

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Sara L. N. Kilmury ◽  
Lori L. Burrows
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iv Pili ◽  
Geobacter Sulfurreducens ◽  
Type Iv Pilus ◽  
Two Component System ◽  
Component System ◽  
Content Type ◽  
Swimming Motility ◽  
Type Iv ◽  
Two Component

ABSTRACTMotility is an important virulence trait for many bacterial pathogens, allowing them to position themselves in appropriate locations at appropriate times. The motility structures type IV pili and flagella are also involved in sensing surface contact, which modulates pathogenicity. InPseudomonas aeruginosa, the PilS-PilR two-component system (TCS) regulates expression of the type IV pilus (T4P) major subunit PilA, while biosynthesis of the single polar flagellum is regulated by a hierarchical system that includes the FleSR TCS. Previous studies ofGeobacter sulfurreducensandDichelobacter nodosusimplicated PilR in regulation of non-T4P-related genes, including some involved in flagellar biosynthesis. Here we used transcriptome sequencing (RNA-seq) analysis to identify genes in addition topilAwith changes in expression in the absence ofpilR. Among the genes identified were 10 genes whose transcription increased in thepilAmutant but decreased in thepilRmutant, despite both mutants lacking T4P and pilus-related phenotypes. The products of these inversely dysregulated genes, many of which were hypothetical, may be important for virulence and surface-associated behaviors, as mutants had altered swarming motility, biofilm formation, type VI secretion system expression, and pathogenicity in a nematode model. Further, the PilSR TCS positively regulated transcription offleSR, and thus many genes in the FleSR regulon. As a result,pilSRdeletion mutants had defects in swimming motility that were independent of the loss of PilA. Together, these data suggest that in addition to controlling T4P expression, PilSR could have a broader role in the regulation ofP. aeruginosamotility and surface sensing behaviors.IMPORTANCESurface appendages such as type IV pili and flagella are important for establishing surface attachment and infection in a host in response to appropriate cues. The PilSR regulatory system that controls type IV pilus expression inPseudomonas aeruginosahas an established role in expression of the major pilin PilA. Here we provide evidence supporting a new role for PilSR in regulating flagellum-dependent swimming motility in addition to pilus-dependent twitching motility. Further, even though bothpilAandpilRmutants lack PilA and pili, we identified sets of genes downregulated in thepilRmutant and upregulated in apilAmutant as well as genes downregulated only in apilRmutant, independent of pilus expression. This finding suggests that change in the inner membrane levels of PilA is only one of the cues to which PilR responds to modulate gene expression. Identification of PilR as a regulator of multiple motility pathways may make it an interesting therapeutic target for antivirulence compounds.

scholarly journals Swarming of Pseudomonas aeruginosa Is Controlled by a Broad Spectrum of Transcriptional Regulators, Including MetR

2009 ◽  
Vol 191 (18) ◽  
pp. 5592-5602 ◽  
Author(s):  
Amy T. Y. Yeung ◽  
Ellen C. W. Torfs ◽  
Farzad Jamshidi ◽  
Manjeet Bains ◽  
Irith Wiegand ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Virulence Gene ◽  
Swarming Motility ◽  
Regulatory Mutants ◽  
Virulence Gene Expression ◽  
Type Iv ◽  
Genes Encoding ◽  
Regulator Genes ◽  
Transposon Insertions

ABSTRACT Pseudomonas aeruginosa exhibits swarming motility on semisolid surfaces (0.5 to 0.7% agar). Swarming is a more than just a form of locomotion and represents a complex adaptation resulting in changes in virulence gene expression and antibiotic resistance. In this study, we used a comprehensive P. aeruginosa PA14 transposon mutant library to investigate how the complex swarming adaptation process is regulated. A total of 233 P. aeruginosa PA14 transposon mutants were verified to have alterations in swarming motility. The swarming-associated genes functioned not only in flagellar or type IV pilus biosynthesis but also in processes as diverse as transport, secretion, and metabolism. Thirty-three swarming-deficient and two hyperswarming mutants had transposon insertions in transcriptional regulator genes, including genes encoding two-component sensors and response regulators; 27 of these insertions were newly identified. Of the 25 regulatory mutants whose swarming motility was highly impaired (79 to 97%), only 1 (a PA1458 mutant) had a major defect in swimming, suggesting that this regulator might influence flagellar synthesis or function. Twitching motility, which requires type IV pili, was strongly affected in only two regulatory mutants (pilH and PA2571 mutants) and was moderately affected in three other mutants (algR, ntrB, and nosR mutants). Microarray analyses were performed to compare the gene expression profile of a swarming-deficient PA3587 mutant to that of the wild-type PA14 strain under swarming conditions. PA3587 showed 63% homology to metR, which encodes a regulator of methionine biosynthesis in Escherichia coli. The observed dysregulation in the metR mutant of nine different genes required for swarming motility provided a possible explanation for the swarming-deficient phenotype of this mutant.

scholarly journals Mucin Promotes Rapid Surface Motility in Pseudomonas aeruginosa

mBio ◽  
2012 ◽  
Vol 3 (3) ◽  
Author(s):  
Amy T. Y. Yeung ◽  
Alicia Parayno ◽  
Robert E. W. Hancock
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type Iv Pili ◽  
Transcriptional Analysis ◽  
Bacterial Cells ◽  
Content Type ◽  
Type Iv ◽  
Sensing Systems ◽  
Bright Green ◽  
Surface Motility

ABSTRACTAn important environmental factor that determines the mode of motility adopted byPseudomonas aeruginosais the viscosity of the medium, often provided by adjusting agar concentrationsin vitro. However, the viscous gel-like property of the mucus layer that overlays epithelial surfaces is largely due to the glycoprotein mucin.P. aeruginosais known to swim within 0.3% (wt/vol) agar and swarm on the surface at 0.5% (wt/vol) agar with amino acids as a weak nitrogen source. When physiological concentrations or as little as 0.05% (wt/vol) mucin was added to the swimming agar, in addition to swimming,P. aeruginosawas observed to undergo highly accelerated motility on the surface of the agar. The surface motility colonies in the presence of mucin appeared to be circular, with a bright green center surrounded by a thicker white edge. While intact flagella were required for the surface motility in the presence of mucin, type IV pili and rhamnolipid production were not. Replacement of mucin with other wetting agents indicated that the lubricant properties of mucin might contribute to the surface motility. Based on studies with mutants, the quorum-sensing systems (lasandrhl) and the orphan autoinducer receptor QscR played important roles in this form of surface motility. Transcriptional analysis of cells taken from the motility zone revealed the upregulation of genes involved in virulence and resistance. Based on these results, we suggest that mucin may be promoting a new or highly modified form of surface motility, which we propose should be termed “surfing.”IMPORTANCEAn important factor that dictates the mode of motility adopted byP. aeruginosais the viscosity of the medium, often provided by adjusting agar concentrationsin vitro. However, the gel-like properties of the mucous layers that overlay epithelial surfaces, such as those of the lung, a major site ofPseudomonasinfection, are contributed mostly by the production of the glycoprotein mucin. In this study, we added mucin to swimming media and found that it promoted the ability ofP. aeruginosato exhibit rapid surface motility. These motility colonies appeared in a circular form, with a bright green center surrounded by a thicker white edge. Interestingly, bacterial cells at the thick edge appeared piled up and lacked flagella, while cells at the motility center had flagella. Our data from various genetic and phenotypic studies suggest that mucin may be promoting a modified form of swarming or a novel form of surface motility inP. aeruginosa.

scholarly journals Flagellar Mutants Have Reduced Pilus Synthesis in Caulobacter crescentus

2019 ◽  
Vol 201 (18) ◽  
Author(s):  
Courtney K. Ellison ◽  
Douglas B. Rusch ◽  
Yves V. Brun
Keyword(s):  
Cell Biology ◽  
Regulatory Networks ◽  
Caulobacter Crescentus ◽  
Single Cells ◽  
Bacterial Species ◽  
Type Iv Pili ◽  
Content Type ◽  
Transcription Profiles ◽  
Type Iv ◽  
Genes Encoding

ABSTRACT Surface appendages, such as flagella and type IV pili, mediate a broad range of bacterial behaviors, including motility, attachment, and surface sensing. While many species harbor both flagella and type IV pili, little is known about how or if their syntheses are coupled. Here, we show that deletions of genes encoding different flagellum machinery components result in a reduction of pilus synthesis in Caulobacter crescentus. First, we show that different flagellar mutants exhibit different levels of sensitivity to a pilus-dependent phage and that fewer cells within populations of flagellar mutants make pili. Furthermore, we find that single cells within flagellar mutant populations produce fewer pili per cell. We demonstrate that these gene deletions result in reduced transcription of pilus-associated genes and have a slight but significant effect on general transcription profiles. Finally, we show that the decrease in pilus production is due to a reduction in the pool of pilin subunits that are polymerized into pilus fibers. These data demonstrate that mutations in flagellar gene components not only affect motility but also can have considerable and unexpected consequences for other aspects of cell biology. IMPORTANCE Most bacterial species synthesize surface-exposed appendages that are important for environmental interactions and survival under diverse conditions. It is often assumed that these appendages act independently of each other and that mutations in either system can be used to assess functionality in specific processes. However, we show that mutations in flagellar genes can impact the production of type IV pili, as well as alter general RNA transcriptional profiles compared to a wild-type strain. These data demonstrate that seemingly simple mutations can broadly affect cell-regulatory networks.

scholarly journals Novel Role for PilNO in Type IV Pilus Retraction Revealed by Alignment Subcomplex Mutations

2015 ◽  
Vol 197 (13) ◽  
pp. 2229-2238 ◽  
Author(s):  
Tiffany L. Leighton ◽  
Neha Dayalani ◽  
Liliana M. Sampaleanu ◽  
P. Lynne Howell ◽  
Lori L. Burrows
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Small Molecule ◽  
Small Molecule Inhibitors ◽  
Type Iv Pili ◽  
Site Directed Mutagenesis ◽  
Twitching Motility ◽  
Content Type ◽  
The Core ◽  
Type Iv ◽  
Two Hybrid

ABSTRACTType IV pili (T4P) are dynamic protein filaments that mediate bacterial adhesion, biofilm formation, and twitching motility. The highly conserved PilMNOP proteins form an inner membrane alignment subcomplex required for function of the T4P system, though their exact roles are unclear. Three potential interaction interfaces for PilNO were identified: core-core, coiled coils (CC), and the transmembrane segments (TMSs). A high-confidence PilNO heterodimer model was used to select key residues for mutation, and the resulting effects on protein-protein interactions were examined both in a bacterial two-hybrid (BTH) system and in their nativePseudomonas aeruginosacontext. Mutations in the oppositely charged CC regions or the TMS disrupted PilNO heterodimer formation in the BTH assay, while up to six combined mutations in the core failed to disrupt the interaction. When the mutations were introduced into theP. aeruginosachromosome at thepilNorpilOlocus, specific changes at each of the three interfaces—including core mutations that failed to disrupt interactions in the BTH system—abrogated surface piliation and/or impaired twitching motility. Unexpectedly, specific CC mutants were hyperpiliated but nonmotile, a hallmark of pilus retraction defects. These data suggest that PilNO participate in both the extension and retraction of T4P. Our findings support a model of multiple, precise interaction interfaces between PilNO; emphasize the importance of studying protein function in a minimally perturbed context and stoichiometry; and highlight potential target sites for development of small-molecule inhibitors of the T4P system.IMPORTANCEPseudomonas aeruginosais an opportunistic pathogen that uses type IV pili (T4P) for host attachment. The T4P machinery is composed of four cell envelope-spanning subcomplexes. PilN and PilO heterodimers are part of the alignment subcomplex and essential for T4P function. Three potential PilNO interaction interfaces (the core-core, coiled-coil, and transmembrane segment interfaces) were probed using site-directed mutagenesis followed by functional assays in anEscherichia colitwo-hybrid system and inP. aeruginosa. Several mutations blocked T4P assembly and/or motility, including two that revealed a novel role for PilNO in pilus retraction, while other mutations affected extension dynamics. These critical PilNO interaction interfaces represent novel targets for small-molecule inhibitors with the potential to disrupt T4P function.

scholarly journals Type IV Pili Can Mediate Bacterial Motility within Epithelial Cells

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Vincent Nieto ◽  
Abby R. Kroken ◽  
Melinda R. Grosser ◽  
Benjamin E. Smith ◽  
Matteo M. E. Metruccio ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Host Cell ◽  
Type Iv Pili ◽  
Fluorescent Imaging ◽  
Bacterial Motility ◽  
Twitching Motility ◽  
Wild Type ◽  
Content Type ◽  
Type Iv

ABSTRACT Pseudomonas aeruginosa is among bacterial pathogens capable of twitching motility, a form of surface-associated movement dependent on type IV pili (T4P). Previously, we showed that T4P and twitching were required for P. aeruginosa to cause disease in a murine model of corneal infection, to traverse human corneal epithelial multilayers, and to efficiently exit invaded epithelial cells. Here, we used live wide-field fluorescent imaging combined with quantitative image analysis to explore how twitching contributes to epithelial cell egress. Results using time-lapse imaging of cells infected with wild-type PAO1 showed that cytoplasmic bacteria slowly disseminated throughout the cytosol at a median speed of >0.05 μm s−1 while dividing intracellularly. Similar results were obtained with flagellin (fliC) and flagellum assembly (flhA) mutants, thereby excluding swimming, swarming, and sliding as mechanisms. In contrast, pilA mutants (lacking T4P) and pilT mutants (twitching motility defective) appeared stationary and accumulated in expanding aggregates during intracellular division. Transmission electron microscopy confirmed that these mutants were not trapped within membrane-bound cytosolic compartments. For the wild type, dissemination in the cytosol was not prevented by the depolymerization of actin filaments using latrunculin A and/or the disruption of microtubules using nocodazole. Together, these findings illustrate a novel form of intracellular bacterial motility differing from previously described mechanisms in being directly driven by bacterial motility appendages (T4P) and not depending on polymerized host actin or microtubules. IMPORTANCE Host cell invasion can contribute to disease pathogenesis by the opportunistic pathogen Pseudomonas aeruginosa. Previously, we showed that the type III secretion system (T3SS) of invasive P. aeruginosa strains modulates cell entry and subsequent escape from vacuolar trafficking to host lysosomes. However, we also showed that mutants lacking either type IV pili (T4P) or T4P-dependent twitching motility (i) were defective in traversing cell multilayers, (ii) caused less pathology in vivo, and (iii) had a reduced capacity to exit invaded cells. Here, we report that after vacuolar escape, intracellular P. aeruginosa can use T4P-dependent twitching motility to disseminate throughout the host cell cytoplasm. We further show that this strategy for intracellular dissemination does not depend on flagellin and resists both host actin and host microtubule disruption. This differs from mechanisms used by previously studied pathogens that utilize either host actin or microtubules for intracellular dissemination independently of microbe motility appendages.

scholarly journals Role of Cyclic Di-GMP and Exopolysaccharide in Type IV Pilus Dynamics

2017 ◽  
Vol 199 (8) ◽  
Author(s):  
Jan Ribbe ◽  
Amy E. Baker ◽  
Sebastian Euler ◽  
George A. O'Toole ◽  
Berenike Maier
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Molecular Motor ◽  
Type Iv Pili ◽  
Force Generation ◽  
Surface Attachment ◽  
Content Type ◽  
Type Iv ◽  
Critical Components ◽  
Planktonic State

ABSTRACT For Pseudomonas aeruginosa, levels of cyclic di-GMP (c-di-GMP) govern the transition from the planktonic state to biofilm formation. Type IV pili (T4P) are crucial determinants of biofilm structure and dynamics, but it is unknown how levels of c-di-GMP affect pilus dynamics. Here, we scrutinized how c-di-GMP affects molecular motor properties and adhesive behavior of T4P. By means of retraction, T4P generated forces of ∼30 pN. Deletion mutants in the proteins with known roles in biofilm formation, swarming motility, and exopolysaccharide (EPS) production (specifically, the diguanylate cyclases sadC and roeA or the c-di-GMP phosphodiesterase bifA) showed only modest effects on velocity or force of T4P retraction. At high levels of c-di-GMP, the production of exopolysaccharides, particularly of Pel, is upregulated. We found that Pel production strongly enhances T4P-mediated surface adhesion of P. aeruginosa, suggesting that T4P-matrix interactions may be involved in biofilm formation by P. aeruginosa. Finally, our data support the previously proposed model of slingshot-like “twitching” motility of P. aeruginosa. IMPORTANCE Type IV pili (T4P) play various important roles in the transition of bacteria from the planktonic state to the biofilm state, including surface attachment and surface sensing. Here, we investigate adhesion, dynamics, and force generation of T4P after bacteria engage a surface. Our studies showed that two critical components of biofilm formation by Pseudomonas aeruginosa, T4P and exopolysaccharides, contribute to enhanced T4P-mediated force generation by attached bacteria. These data indicate a crucial role for the coordinated impact of multiple biofilm-promoting factors during the early stages of attachment to a surface. Our data are also consistent with a previous model explaining why pilus-mediated motility in P. aeruginosa results in characteristic “twitching” behavior.

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