Faculty Opinions recommendation of Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants.

ABSTRACT The opportunistic pathogen Pseudomonas aeruginosa produces multifunctional, polar, filamentous appendages termed type IV pili. Type IV pili are involved in colonization during infection, twitching motility, biofilm formation, bacteriophage infection, and natural transformation. Electrostatic surface analysis of modeled pilus fibers generated from P. aeruginosa strain PAK, K122-4, and KB-7 pilin monomers suggested that a solvent-exposed band of positive charge may be a common feature of all type IV pili. Several functions of type IV pili, including natural transformation and biofilm formation, involve DNA. We investigated the ability of P. aeruginosa type IV pili to bind DNA. Purified PAK, K122-4, and KB-7 pili were observed to bind both bacterial plasmid and salmon sperm DNA in a concentration-dependent and saturable manner. PAK pili had the highest affinity for DNA, followed by K122-4 and KB-7 pili. DNA binding involved backbone interactions and preferential binding to pyrimidine residues even though there was no evidence of sequence-specific binding. Pilus-mediated DNA binding was a function of the intact pilus and thus required elements present in the quaternary structure. However, binding also involved the pilus tip as tip-specific, but not base-specific, antibodies inhibited DNA binding. The conservation of a Thr residue in all type IV pilin monomers examined to date, along with the electrostatic data, implies that DNA binding is a conserved function of type IV pili. Pilus-mediated DNA binding could be important for biofilm formation both in vivo during an infection and ex vivo on abiotic surfaces.

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Type IV Pili Are Required for Virulence, Twitching Motility, and Biofilm Formation of Acidovorax avenae subsp. citrulli

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-22-8-0909 ◽

2009 ◽

Vol 22 (8) ◽

pp. 909-920 ◽

Cited By ~ 84

Author(s):

Ofir Bahar ◽

Tal Goffer ◽

Saul Burdman

Keyword(s):

Biofilm Formation ◽

Host Plants ◽

Seed Transmission ◽

Type Iv Pili ◽

Twitching Motility ◽

Wild Type ◽

Type Iv ◽

Group I ◽

Tn5 Mutant

Acidovorax avenae subsp. citrulli is the causal agent of bacterial fruit blotch (BFB), a threatening disease of watermelon, melon, and other cucurbits. Despite the economic importance of BFB, relatively little is known about basic aspects of the pathogen's biology and the molecular basis of its interaction with host plants. To identify A. avenae subsp. citrulli genes associated with pathogenicity, we generated a transposon (Tn5) mutant library on the background of strain M6, a group I strain of A. avenae subsp. citrulli, and screened it for reduced virulence by seed-transmission assays with melon. Here, we report the identification of a Tn5 mutant with reduced virulence that is impaired in pilM, which encodes a protein involved in assembly of type IV pili (TFP). Further characterization of this mutant revealed that A. avenae subsp. citrulli requires TFP for twitching motility and wild-type levels of biofilm formation. Significant reductions in virulence and biofilm formation as well as abolishment of twitching were also observed in insertional mutants affected in other TFP genes. We also provide the first evidence that group I strains of A. avenae subsp. citrulli can colonize and move through host xylem vessels.

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Non-motile subpopulations of Pseudomonas aeruginosa repress flagellar motility in motile cells through a type IV pili- and Pel-dependent mechanism

10.1101/2021.10.21.465385 ◽

2021 ◽

Author(s):

Kimberley A. Lewis ◽

Danielle M Vermilyea ◽

Shanice S Webster ◽

Jaime de Anda ◽

Gerard Wong ◽

...

Keyword(s):

Biofilm Formation ◽

Total Population ◽

Type Iv Pili ◽

External Factors ◽

Flagellar Motility ◽

Wild Type ◽

Type Iv ◽

Motile Cells ◽

Flagellum Biosynthesis ◽

Dependent Mechanism

The downregulation of P. aeruginosa flagellar motility is a key event in biofilm formation, host-colonization, and the formation of microbial communities, but the external factors that repress motility are not well understood. Here, we report that under swarming conditions, motility can be repressed by cells that are non-motile due to the absence of a flagellum or flagellar rotation. Non-motile cells, due to mutations that prevent either flagellum biosynthesis or rotation, present at 5% of the total population suppressed swarming of wild-type cells under the conditions tested in this study. Non-motile cells required functional type IV pili and the ability to produce the Pel exopolysaccharide to suppress swarming by the motile wild type. In contrast, motile cells required only type IV pili, but not Pel production, in order for swarming to be repressed by non-motile cells. We hypothesize that interactions between motile and non-motile cells may enhance the formation of sessile communities including those involving multiple genotypes, phenotypically-diverse cells, and perhaps other species.

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Pseudomonas aeruginosa Exhibits Directed Twitching Motility Up Phosphatidylethanolamine Gradients

Journal of Bacteriology ◽

10.1128/jb.183.2.763-767.2001 ◽

2001 ◽

Vol 183 (2) ◽

pp. 763-767 ◽

Cited By ~ 70

Author(s):

Daniel B. Kearns ◽

Jayne Robinson ◽

Lawrence J. Shimkets

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Solid Surfaces ◽

Twitching Motility ◽

Wild Type ◽

Directed Movement ◽

Endogenous Factors ◽

Type Iv ◽

Cell Velocity ◽

Dependent Form

ABSTRACT Pseudomonas aeruginosa translocates over solid surfaces by a type IV pilus-dependent form of multicellular motility known as twitching. We wondered whether cells utilize endogenous factors to organize twitching, and we purified from wild-type cells a lipid that caused directed movement. Wild-type P. aeruginosa, but not a pilJ pilus-deficient mutant, showed biased movement up gradients of phosphatidylethanolamine (PE) established in agar. Activity was related to the fatty acid composition of the lipid, as two synthetic PE species, dilauroyl and dioleoyl PE, were capable of directing P. aeruginosa motility while many other species were inactive. P. aeruginosa PE did not contain either laurate or oleate, implying that the native attractant species contains different fatty acids. Uniform concentrations of PE increased cell velocity, suggesting that chemokinesis may be at least partly responsible for directed movement. We speculate that PE-directed twitching motility may be involved in biofilm formation and pathogenesis.

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Type IV Pili Can Mediate Bacterial Motility within Epithelial Cells

mBio ◽

10.1128/mbio.02880-18 ◽

2019 ◽

Vol 10 (4) ◽

Cited By ~ 6

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.

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

Journal of Bacteriology ◽

10.1128/jb.00859-16 ◽

2017 ◽

Vol 199 (8) ◽

Cited By ~ 12

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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tonB3 Is Required for Normal Twitching Motility and Extracellular Assembly of Type IV Pili

Journal of Bacteriology ◽

10.1128/jb.186.13.4387-4389.2004 ◽

2004 ◽

Vol 186 (13) ◽

pp. 4387-4389 ◽

Cited By ~ 32

Author(s):

Bixing Huang ◽

Kelin Ru ◽

Zheng Yuan ◽

Cynthia B. Whitchurch ◽

John S. Mattick

Keyword(s):

Pseudomonas Aeruginosa ◽

Type Iv Pili ◽

Twitching Motility ◽

Wild Type ◽

Type Iv

ABSTRACT Three mutants with Tn5-B21 insertion in tonB3 (PA0406) of Pseudomonas aeruginosa exhibited defective twitching motility and reduced assembly of extracellular pili. These defects could be complemented with wild-type tonB3.

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Faculty Opinions recommendation of Distinct function of Pseudomonas aeruginosa type IV pili disclosed in the bacterial pass-through of membrane filter with smaller pore sizes.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1081001.533960 ◽

2007 ◽

Author(s):

Esther Bullitt

Keyword(s):

Pseudomonas Aeruginosa ◽

Membrane Filter ◽

Type Iv Pili ◽

Pore Sizes ◽

Type Iv ◽

Pass Through

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Aztreonam Lysine Increases the Activity of Phages E79 and phiKZ against Pseudomonas aeruginosa PA01

Microorganisms ◽

10.3390/microorganisms9010152 ◽

2021 ◽

Vol 9 (1) ◽

pp. 152

Author(s):

Carly M. Davis ◽

Jaclyn G. McCutcheon ◽

Jonathan J. Dennis

Keyword(s):

Pseudomonas Aeruginosa ◽

Morphological Changes ◽

Burst Size ◽

Type Iv Pili ◽

Biofilm Growth ◽

Promising Alternative ◽

Type Iv ◽

Alternative Treatment Option ◽

One Step ◽

Step Growth

Pseudomonas aeruginosa is a pernicious bacterial pathogen that is difficult to treat because of high levels of antibiotic resistance. A promising alternative treatment option for such bacteria is the application of bacteriophages; the correct combination of phages plus antibiotics can produce synergistic inhibitory effects. In this study, we describe morphological changes induced by sub-MIC levels of the antibiotic aztreonam lysine (AzLys) on P. aeruginosa PA01, which may in part explain the observed phage–antibiotic synergy (PAS). One-step growth curves for phage E79 showed increased adsorption rates, decreased infection latency, accelerated time to lysis and a minor reduction in burst size. Phage E79 plus AzLys PAS was also able to significantly reduce P. aeruginosa biofilm growth over 3-fold as compared to phage treatment alone. Sub-inhibitory AzLys-induced filamentation of P. aeruginosa cells resulted in loss of twitching motility and a reduction in swimming motility, likely due to a reduction in the number of polar Type IV pili and flagella, respectively, on the filamented cell surfaces. Phage phiKZ, which uses Type IV pili as a receptor, did not exhibit increased activity with AzLys at lower sub-inhibitory levels, but still produced phage–antibiotic synergistic killing with sub-inhibitory AzLys. A one-step growth curve indicates that phiKZ in the presence of AzLys also exhibits a decreased infection latency and moderately undergoes accelerated time to lysis. In contrast to prior PAS studies demonstrating that phages undergo delayed time to lysis with cell filamentation, these PAS results show that phages undergo accelerated time to lysis, which therefore suggests that PAS is dependent upon multiple factors, including the type of phages and antibiotics used, and the bacterial host being tested.

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