scholarly journals Requirement of the Pseudomonas aeruginosa tonB Gene for High-Affinity Iron Acquisition and Infection

2000 ◽  
Vol 68 (8) ◽  
pp. 4498-4504 ◽  
Author(s):  
Hiroyuki Takase ◽  
Hironobu Nitanai ◽  
Kazuki Hoshino ◽  
Tsuyoshi Otani
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Iron Acquisition ◽  
Animal Experiments ◽  
Strain Pao1 ◽  
Free Iron ◽  
High Affinity ◽  
Tonb Protein ◽  
In Vivo Growth ◽  
Immunosuppressed Mice

ABSTRACT To investigate the contribution of the TonB protein to high-affinity iron acquisition in Pseudomonas aeruginosa, we constructed tonB-inactivated mutants from strain PAO1 and its derivative deficient in producing the siderophores pyoverdin and pyochelin. The tonB mutants could not grow in a free-iron-restricted medium prepared by apotransferrin addition, even though the medium was supplemented with each purified siderophore or with a heme source (hemoglobin or hemin). The tonBinactivation was shown to make P. aeruginosa unable to acquire iron from the transferrin with either siderophore. Introduction of a plasmid carrying the intact tonB gene restored growth of the tonB mutant of PAO1 in the free-iron-restricted medium without any supplements and restored growth of thetonB mutant of the siderophore-deficient derivative in the medium supplemented with pyoverdin, pyochelin, hemoglobin, or hemin. In addition, animal experiments showed that, in contrast to PAO1, thetonB mutant of PAO1 could not grow in vivo, such as in the muscles and lungs of immunosuppressed mice, and could not kill any of the animals. The in vivo growth ability and lethal virulence were also restored by introduction of the tonB-carrying plasmid in the tonB mutant. These results indicate clearly that the intact tonB gene—and, therefore, the TonB protein encoded by it—is essential for iron acquisition mediated by pyoverdin and pyochelin and via heme uptake in P. aeruginosa and suggest that the TonB-dependent iron acquisition may be essential for P. aeruginosa to infect the animal host.

scholarly journals Staphylococcus aureus Serves as an Iron Source for Pseudomonas aeruginosa during In Vivo Coculture

2005 ◽  
Vol 187 (2) ◽  
pp. 554-566 ◽  
Author(s):  
Lauren M. Mashburn ◽  
Amy M. Jett ◽  
Darrin R. Akins ◽  
Marvin Whiteley
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Pathogenic Bacteria ◽  
Iron Acquisition ◽  
Low Oxygen ◽  
Dialysis Membrane ◽  
Opportunistic Human Pathogen ◽  
The Impact

ABSTRACT Pseudomonas aeruginosa is a gram-negative opportunistic human pathogen often infecting the lungs of individuals with the heritable disease cystic fibrosis and the peritoneum of individuals undergoing continuous ambulatory peritoneal dialysis. Often these infections are not caused by colonization with P. aeruginosa alone but instead by a consortium of pathogenic bacteria. Little is known about growth and persistence of P. aeruginosa in vivo, and less is known about the impact of coinfecting bacteria on P. aeruginosa pathogenesis and physiology. In this study, a rat dialysis membrane peritoneal model was used to evaluate the in vivo transcriptome of P. aeruginosa in monoculture and in coculture with Staphylococcus aureus. Monoculture results indicate that approximately 5% of all P. aeruginosa genes are differentially regulated during growth in vivo compared to in vitro controls. Included in this analysis are genes important for iron acquisition and growth in low-oxygen environments. The presence of S. aureus caused decreased transcription of P. aeruginosa iron-regulated genes during in vivo coculture, indicating that the presence of S. aureus increases usable iron for P. aeruginosa in this environment. We propose a model where P. aeruginosa lyses S. aureus and uses released iron for growth in low-iron environments.

scholarly journals Identification and Characterization of the PutP Proline Permease That Contributes to In Vivo Survival ofStaphylococcus aureus in Animal Models

1998 ◽  
Vol 66 (2) ◽  
pp. 567-572 ◽  
Author(s):  
William R. Schwan ◽  
Silvija N. Coulter ◽  
Eva Y. W. Ng ◽  
Michael H. Langhorne ◽  
Heather D. Ritchie ◽  
...  
Keyword(s):  
Animal Models ◽  
Reading Frame ◽  
High Affinity ◽  
Wound Model ◽  
Infection Models ◽  
Uptake Assay ◽  
Entire Open Reading Frame ◽  
In Vivo Growth

ABSTRACT Staphylococcus aureus is an important pathogen of humans and other animals, causing bacteremia, abscesses, endocarditis, and other infectious syndromes. A signature-tagged mutagenesis (STM) system was adapted for use in studying the genes required for in vivo survival of S. aureus. An STM library was ultimately created in S. aureus RN6390, with Tn917 being used to create the transposon mutations. Pools of S. aureusRN6390 mutants were screened in mouse abscess, bacteremia, and wound infection models for growth attenuation after in vivo passage. One of the mutants that was identified displayed marked attenuation following large-pool screening in all three animal models, which was confirmed in bacteremia and endocarditis models of infection with a smaller pool of mutants. Sequence analysis of the entire open reading frame showed a 99% identity to the high-affinity proline permease (putP) gene characterized in another strain of S. aureus. In wound and murine abscess infection models, the putP mutant was approximately 10-fold more attenuated than was wild-type strain RN6390. Another S. aureus strain transduced with theputP mutation also displayed an attenuated phenotype after passage in the wound model. A [3H]proline uptake assay showed that less proline was specifically transported into theputP mutant than into strain RN6390. The reduced viability of the bacteria possessing the mutation in the S. aureushigh-affinity proline permease suggests that proline scavenging by the bacteria is important for in vivo growth and proliferation and that analogs of proline may serve as potential antistaphylococcal therapeutic agents.

scholarly journals Clearance of Pseudomonas aeruginosa from a Healthy Ocular Surface Involves Surfactant Protein D and Is Compromised by Bacterial Elastase in a Murine Null-Infection Model

2009 ◽  
Vol 77 (6) ◽  
pp. 2392-2398 ◽  
Author(s):  
James J. Mun ◽  
Connie Tam ◽  
David Kowbel ◽  
Samuel Hawgood ◽  
Mitchell J. Barnett ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Ocular Surface ◽  
Surfactant Protein ◽  
Tear Fluid ◽  
Bacterial Invasion ◽  
Surfactant Protein D ◽  
Infection Model ◽  
Strain Pao1 ◽  
Corneal Epithelial

ABSTRACT Our previous studies showed that surfactant protein D (SP-D) is present in human tear fluid and that it can protect corneal epithelial cells against bacterial invasion. Here we developed a novel null-infection model to test the hypothesis that SP-D contributes to the clearance of viable Pseudomonas aeruginosa from the healthy ocular surface in vivo. Healthy corneas of Black Swiss mice were inoculated with 107 or 109 CFU of invasive (PAO1) or cytotoxic (6206) P. aeruginosa. Viable counts were performed on tear fluid collected at time points ranging from 3 to 14 h postinoculation. Healthy ocular surfaces cleared both P. aeruginosa strains efficiently, even when 109 CFU was used: e.g., <0.01% of the original inoculum was recoverable after 3 h. Preexposure of eyes to bacteria did not enhance clearance. Clearance of strain 6206 (low protease producer), but not strain PAO1 (high protease producer), was delayed in SP-D gene-targeted (SP-D−/−) knockout mice. A protease mutant of PAO1 (PAO1 lasA lasB aprA) was cleared more efficiently than wild-type PAO1, but this difference was negligible in SP-D−/− mice, which were less able to clear the protease mutant. Experiments to study mechanisms for these differences revealed that purified elastase could degrade tear fluid SP-D in vivo. Together, these data show that SP-D can contribute to the clearance of P. aeruginosa from the healthy ocular surface and that proteases can compromise that clearance. The data also suggest that SP-D degradation in vivo is a mechanism by which P. aeruginosa proteases could contribute to virulence.

Substrate specificity of the high-affinity glucose transport system of Pseudomonas aeruginosa

1993 ◽  
Vol 39 (7) ◽  
pp. 722-725 ◽  
Author(s):  
John L. Wylie ◽  
Elizabeth A. Worobec
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Glucose Transport ◽  
Transport System ◽  
Binding Protein ◽  
High Affinity ◽  
Glucose Binding ◽  
Glucose Transport System ◽  
Glucose Binding Protein

Specificity of the high-affinity glucose transport system of Pseudomonas aeruginosa was examined. At a concentration of [14C]glucose near the Vmax of the system, inhibition by maltose, galactose, and xylose was detected. This inhibition is similar to that detected in earlier in vivo studies and correlates with the known specificity of OprB, a glucose-specific porin of P. aeruginosa. At a level of [14C]glucose 100 times lower, only unlabelled glucose inhibited uptake to any extent. This matches the known in vitro specificity of the periplasmic glucose binding protein. These findings were used to explain the discrepancy between earlier in vivo and in vitro results reported in the literature.Key words: Pseudomonas aeruginosa, glucose transport, OprB, glucose binding protein.

scholarly journals Yersiniabactin Is a Virulence Factor for Klebsiella pneumoniae during Pulmonary Infection

2007 ◽  
Vol 75 (3) ◽  
pp. 1463-1472 ◽  
Author(s):  
Matthew S. Lawlor ◽  
Christopher O'Connor ◽  
Virginia L. Miller
Keyword(s):  
Klebsiella Pneumoniae ◽  
Iron Acquisition ◽  
Infection Model ◽  
Mutagenesis Screen ◽  
Starting Point ◽  
In Vivo Growth ◽  
Isogenic Mutants ◽  
Isogenic Strains

ABSTRACT Iron acquisition systems are essential for the in vivo growth of bacterial pathogens. Despite the epidemiological importance of Klebsiella pneumoniae, few experiments have examined the importance of siderophores in the pathogenesis of this species. A previously reported signature-tagged mutagenesis screen identified an attenuated strain that featured an insertional disruption in ybtQ, which encodes a transporter for the siderophore yersiniabactin. We used this finding as a starting point to evaluate the importance of siderophores in the physiology and pathogenesis of K. pneumoniae. Isogenic strains carrying in-frame deletions in genes required for the synthesis of either enterobactin or yersiniabactin were constructed, and the growth of these mutants was examined both in vitro and in vivo using an intranasal infection model. The results suggest divergent functions for each siderophore in different environments, with enterobactin being more important for growth in vitro under iron limitation than in vivo and the reverse being true for the yersiniabactin locus. These observations represent the first examination of isogenic mutants in iron acquisition systems for K. pneumoniae and may indicate that the acquisition of nonenterobactin siderophores is an important step in the evolution of virulent enterobacterial strains.

scholarly journals Adaptation-Based Resistance to Siderophore-Conjugated Antibacterial Agents by Pseudomonas aeruginosa

2013 ◽  
Vol 57 (9) ◽  
pp. 4197-4207 ◽  
Author(s):  
Andrew P. Tomaras ◽  
Jared L. Crandon ◽  
Craig J. McPherson ◽  
Mary Anne Banevicius ◽  
Steven M. Finegan ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibacterial Agents ◽  
Iron Acquisition ◽  
Content Type ◽  
Multiple Strains ◽  
Resistance Rates ◽  
Assay Conditions

ABSTRACTMultidrug resistance in Gram-negative bacteria has become so threatening to human health that new antibacterial platforms are desperately needed to combat these deadly infections. The concept of siderophore conjugation, which facilitates compound uptake across the outer membrane by hijacking bacterial iron acquisition systems, has received significant attention in recent years. While standardin vitroMIC and resistance frequency methods demonstrate that these compounds are potent, broad-spectrum antibacterial agents whose activity should not be threatened by unacceptably high spontaneous resistance rates, recapitulation of these results in animal models can prove unreliable, partially because of the differences in iron availability in these different methods. Here, we describe the characterization of MB-1, a novel siderophore-conjugated monobactam that demonstrates excellentin vitroactivity againstPseudomonas aeruginosawhen tested using standard assay conditions. Unfortunately, thein vitrofindings did not correlate with thein vivoresults we obtained, as multiple strains were not effectively treated by MB-1 despite having low MICs. To address this, we also describe the development of newin vitroassays that were predictive of efficacy in mouse models, and we provide evidence that competition with native siderophores could contribute to the recalcitrance of someP. aeruginosaisolatesin vivo.

scholarly journals Priming with intranasal lactobacilli prevents Pseudomonas aeruginosa acute pneumonia in mice.

2020 ◽  
Author(s):  
Marie-Sarah FANGOUS ◽  
Philippe Gosset ◽  
Nicolas Galakhoff ◽  
Stéphanie Gouriou ◽  
Charles-Antoine Guilloux ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Virulence Factors ◽  
Saline Solution ◽  
Clinical Isolates ◽  
Control Groups ◽  
Strain Pao1 ◽  
Log Reduction ◽  
Animal Survival

Abstract Background : Increasing resistance to antibiotics of Pseudomonas aeruginosa leads to therapeutic deadlock and alternative therapies are needed. We aimed to evaluate the effects of Lactobacillus clinical isolates in vivo, through intranasal administration on a murine model of Pseudomonas aeruginosa pneumonia.Results : We screened in vitro 50 pulmonary clinical isolates of Lactobacillus for their ability to decrease the synthesis of two QS dependent-virulence factors (elastase and pyocyanin) produced by Pseudomonas aeruginosa strain PAO1.Two blends of three Lactobacillus isolates were then tested in vivo: one with highly effective anti-PAO1 virulence factors properties (blend named L.rff for L. rhamnosus, two L. fermentum strains), and the second with no properties (blend named L.psb, for L. paracasei, L. salivarius and L. brevis). Each blend was administered intranasally to mice 18h prior to PAO1 pulmonary infection. Animal survival, bacterial loads, cytological analysis, and cytokines secretion in the lungs were evaluated at 6 or 24h post infection with PAO1. Intranasal priming with both lactobacilli blends significantly improved 7-day mice survival from 12% for the control PAO1 group to 71% and 100% for the two groups receiving L.rff and L.psb respectively. No mortality was observed for both control groups receiving either L.rff or L.psb. Additionally, the PAO1 lung clearance was significantly enhanced at 24h. A 2-log and 4-log reduction was observed in the L.rff+PAO1 and L.psb+PAO1 groups respectively, compared to the control PAO1 group. Significant reductions in neutrophil recruitment and proinflammatory cytokine and chemokine secretion were observed after lactobacilli administration compared to saline solution, whereas IL-10 production was increased. Conclusions : These results demonstrate that intranasal priming with lactobacilli acts as a prophylaxis, and avoids fatal complications caused by Pseudomonas aeruginosa pneumonia in mice. These results were independent of in vitro anti-Pseudomonas aeruginosa activity on QS-dependent virulence factors. Further experiments are required to identify the immune mechanism before initiating clinical trials.

scholarly journals Increased production of the extracellular polysaccharide Psl can give a growth advantage to Pseudomonas aeruginosa in low-iron conditions

2018 ◽  
Author(s):  
Jaime Hutchison ◽  
Karishma Kaushik ◽  
Christopher A. Rodesney ◽  
Thomas Lilieholm ◽  
Layla Bakhtiari ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Iron Acquisition ◽  
Single Cells ◽  
Extracellular Polysaccharide ◽  
Time Lapse ◽  
Evolutionary Development ◽  
Intercellular Signaling ◽  
Biofilm Growth

AbstractIn infections, biofilm formation is associated with a number of fitness advantages, such as resistance to antibiotics and to clearance by the immune system. Biofilm formation has also been linked to fitness advantages in environments other than in vivo infections; primarily, biofilms are thought to help constituent organisms evade predation and to promote intercellular signaling. The opportunistic human pathogen Pseudomonas aeruginosa forms biofilm infections in lungs, wounds, and on implants and medical devices. However, the tendency toward biofilm formation originated in this bacterium’s native environment, primarily plants and soil. Such environments are polymicrobial and often resource-limited. Other researchers have recently shown that the P. aeruginosa extracellular polysaccharide Psl can bind iron. For the lab strain PA01, Psl is also the dominant adhesive and cohesive “glue” holding together multicellular aggregates and biofilms. Here, we perform quantitative time-lapse confocal microscopy and image analysis of early biofilm growth by PA01. We find that aggregates of P. aeruginosa have a growth advantage over single cells of P. aeruginosa in the presence of Staphylococcus aureus in low-iron environments. Our results suggest the growth advantage of aggregates is linked to their high Psl content and to the production of an active factor by S. aureus. We posit that the ability of Psl to promote iron acquisition may have been linked to the evolutionary development of the strong biofilm-forming tendencies of P. aeruginosa.

scholarly journals Constitutive High Expression of Chromosomal β-Lactamase in Pseudomonas aeruginosa Caused by a New Insertion Sequence (IS1669) Located in ampD

2002 ◽  
Vol 46 (11) ◽  
pp. 3406-3411 ◽  
Author(s):  
Niels Bagge ◽  
Oana Ciofu ◽  
Morten Hentzer ◽  
Joan I. A. Campbell ◽  
Michael Givskov ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Insertion Sequence ◽  
Enterobacter Cloacae ◽  
High Level Expression ◽  
Dramatic Decrease ◽  
Strain Pao1 ◽  
Genetic Changes ◽  
High Level

ABSTRACT The expression of chromosomal AmpC β-lactamase in Pseudomonas aeruginosa is negatively regulated by the activity of an amidase, AmpD. In the present study we examined resistant clinical P. aeruginosa strains and several resistant variants isolated from in vivo and in vitro biofilms for mutations in ampD to find evidence for the genetic changes leading to high-level expression of chromosomal β-lactamase. A new insertion sequence, IS1669, was found located in the ampD genes of two clinical P. aeruginosa isolates and several biofilm-isolated variants. The presence of IS1669 in ampD resulted in the expression of high levels of AmpC β-lactamase. Complementation of these isolates with ampD from the reference P. aeruginosa strain PAO1 caused a dramatic decrease in the expression of AmpC β-lactamase and a parallel decrease of the MIC of ceftazidime to a level comparable to that of PAO1. One highly resistant, constitutive β-lactamase-producing variant contained no mutations in ampD, but a point mutation was observed in ampR, resulting in an Asp-135→Asn change. An identical mutation of AmpR in Enterobacter cloacae has been reported to cause a 450-fold higher AmpC expression. However, in many of the isolates expressing high levels of chromosomal β-lactamase, no changes were found in either ampD, ampR, or in the promoter region of ampD, ampR, or ampC. Our results suggest that multiple pathways may exist leading to increased antimicrobial resistance due to chromosomal β-lactamase.

scholarly journals Iron Acquisition from Pseudomonas aeruginosa Siderophores by Human Phagocytes: an Additional Mechanism of Host Defense through Iron Sequestration?

2000 ◽  
Vol 68 (3) ◽  
pp. 1271-1275 ◽  
Author(s):  
Bradley E. Britigan ◽  
George T. Rasmussen ◽  
Oyebode Olakanmi ◽  
Charles D. Cox
Keyword(s):  
Molecular Weight ◽  
Pseudomonas Aeruginosa ◽  
Host Defense ◽  
Iron Acquisition ◽  
High Capacity ◽  
Human Neutrophils ◽  
Low Molecular Weight ◽  
Additional Mechanism ◽  
Low Molecular Weight Iron

ABSTRACT Chelation of iron to iron-binding proteins is a strategy of host defense. Some pathogens counter this via the secretion of low-molecular-weight iron-chelating agents (siderophores). Human phagocytes possess a high-capacity mechanism for iron acquisition from low-molecular-weight iron chelates. Efficient acquisition and sequestration of iron bound to bacterial siderophores by host phagocytes could provide a secondary mechanism to limit microbial access to iron. In the present work we report that human neutrophils, macrophages, and myeloid cell lines can acquire iron from the twoPseudomonas aeruginosa siderophores. Analogous to iron acquisition from other low-molecular-weight chelates, iron acquisition from the siderophores is ATP independent, induced by multivalent cationic metals, and unaffected by inhibitors of endocytosis and pinocytosis. In vivo, this process could serve as an additional mechanism of host defense to limit iron availability to invading siderophore-producing microbes.

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