scholarly journals Transcriptional Response of Mucoid Pseudomonas aeruginosa to Human Respiratory Mucus

mBio ◽  
2012 ◽  
Vol 3 (6) ◽  
Author(s):  
V. Cattoir ◽  
G. Narasimhan ◽  
D. Skurnik ◽  
H. Aschard ◽  
D. Roux ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Opportunistic Pathogen ◽  
Secretion System ◽  
Type Vi Secretion System ◽  
Transcript Levels ◽  
Content Type ◽  
Type Vi Secretion ◽  
Small Regulatory Rnas ◽  
Regulatory Rnas

ABSTRACTAdaptation of bacterial pathogens to a host can lead to the selection and accumulation of specific mutations in their genomes with profound effects on the overall physiology and virulence of the organisms. The opportunistic pathogenPseudomonas aeruginosais capable of colonizing the respiratory tract of individuals with cystic fibrosis (CF), where it undergoes evolution to optimize survival as a persistent chronic human colonizer. The transcriptome of a host-adapted, alginate-overproducing isolate from a CF patient was determined following growth of the bacteria in the presence of human respiratory mucus. This stable mucoid strain responded to a number of regulatory inputs from the mucus, resulting in an unexpected repression of alginate production. Mucus in the medium also induced the production of catalases and additional peroxide-detoxifying enzymes and caused reorganization of pathways of energy generation. A specific antibacterial type VI secretion system was also induced in mucus-grown cells. Finally, a group of small regulatory RNAs was identified and a fraction of these were mucus regulated. This report provides a snapshot of responses in a pathogen adapted to a human host through assimilation of regulatory signals from tissues, optimizing its long-term survival potential.IMPORTANCEThe basis for chronic colonization of patients with cystic fibrosis (CF) by the opportunistic pathogenPseudomonas aeruginosacontinues to represent a challenging problem for basic scientists and clinicians. In this study, the host-adapted, alginate-overproducingPseudomonas aeruginosa2192 strain was used to assess the changes in its transcript levels following growth in respiratory CF mucus. Several significant and unexpected discoveries were made: (i) although the alginate overproduction in strain 2192 was caused by a stable mutation, a mucus-derived signal caused reduction in the transcript levels of alginate biosynthetic genes; (ii) mucus activated the expression of the type VI secretion system, a mechanism for killing of other bacteria in a mixed population; (iii) expression of a number of genes involved in respiration was altered; and (iv) several small regulatory RNAs were identified, some being mucus regulated. This work highlights the strong influence of the host environment in shaping bacterial survival strategies.

scholarly journals Type VI Secretion System Dynamics Reveals a Novel Secretion Mechanism in Pseudomonas aeruginosa

2018 ◽  
Vol 200 (11) ◽  
Author(s):  
Jacqueline Corbitt ◽  
Jun Seok Yeo ◽  
C. Ian Davis ◽  
Michele LeRoux ◽  
Paul A. Wiggins
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Vibrio Cholerae ◽  
Conformational Change ◽  
Secretion System ◽  
Force Generation ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Evolutionary Diversification ◽  
Type Vi Secretion ◽  
Generation Mechanisms

ABSTRACT The type VI secretion system (T6SS) inhibits the growth of neighboring bacterial cells through a contact-mediated mechanism. Here, we describe a detailed characterization of the protein localization dynamics in the Pseudomonas aeruginosa T6SS. It has been proposed that the type VI secretion process is driven by a conformational-change-induced contraction of the T6SS sheath. However, although the contraction of an optically resolvable TssBC sheath and the subsequent localization of ClpV are observed in Vibrio cholerae , coordinated assembly and disassembly of TssB and ClpV are observed without TssB contraction in P. aeruginosa . These dynamics are inconsistent with the proposed contraction sheath model. Motivated by the phenomenon of dynamic instability, we propose a new model in which ATP hydrolysis, rather than conformational change, generates the force for secretion. IMPORTANCE The type VI secretion system (T6SS) is widely conserved among Gram-negative bacteria and is a central determinant of bacterial fitness in polymicrobial communities. The secretion system targets bacteria and secretes effectors that inhibit the growth of neighboring cells, using a contact-mediated-delivery system. Despite significant homology to the previously characterized Vibrio cholerae T6SS, our analysis reveals that effector secretion is driven by a distinct force generation mechanism in Pseudomonas aeruginosa . The presence of two distinct force generation mechanisms in T6SS represents an example of the evolutionary diversification of force generation mechanisms.

scholarly journals TheprrF-Encoded Small Regulatory RNAs Are Required for Iron Homeostasis and Virulence of Pseudomonas aeruginosa

2014 ◽  
Vol 83 (3) ◽  
pp. 863-875 ◽  
Author(s):  
Alexandria A. Reinhart ◽  
Daniel A. Powell ◽  
Angela T. Nguyen ◽  
Maura O'Neill ◽  
Louise Djapgne ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Iron Homeostasis ◽  
Opportunistic Pathogen ◽  
Wild Type ◽  
Content Type ◽  
Genes Encoding ◽  
Small Regulatory Rnas ◽  
Regulatory Rnas ◽  
Heme Regulation ◽  
Heme Binding Protein

Pseudomonas aeruginosais an opportunistic pathogen that requires iron to cause infection, but it also must regulate the uptake of iron to avoid iron toxicity. The iron-responsive PrrF1 and PrrF2 small regulatory RNAs (sRNAs) are part ofP. aeruginosa'siron regulatory network and affect the expression of at least 50 genes encoding iron-containing proteins. The genes encoding the PrrF1 and PrrF2 sRNAs are encoded in tandem inP. aeruginosa, allowing for the expression of a distinct, heme-responsive sRNA named PrrH that appears to regulate genes involved in heme metabolism. Using a combination of growth, mass spectrometry, and gene expression analysis, we showed that the ΔprrF1,2mutant, which lacks expression of the PrrF and PrrH sRNAs, is defective for both iron and heme homeostasis. We also identifiedphuS, encoding a heme binding protein involved in heme acquisition, andvreR, encoding a previously identified regulator ofP. aeruginosavirulence genes, as novel targets ofprrF-mediated heme regulation. Finally, we showed that theprrFlocus encoding the PrrF and PrrH sRNAs is required forP. aeruginosavirulence in a murine model of acute lung infection. Moreover, we showed that inoculation with a ΔprrF1,2deletion mutant protects against future challenge with wild-typeP. aeruginosa. Combined, these data demonstrate that theprrF-encoded sRNAs are critical regulators ofP. aeruginosavirulence.

scholarly journals The Type VI Secretion System of Pseudomonas aeruginosa: a gun loaded with antimicrobial bullets

2020 ◽  
Vol 2 (7A) ◽  
Author(s):  
Sophie Howard ◽  
Chris Furniss ◽  
Dora Bonini ◽  
Himani Amin ◽  
Patricia Paracuellos-Torrecilla ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Respiratory Infections ◽  
Opportunistic Pathogen ◽  
Secretion System ◽  
Type Vi Secretion System ◽  
Type Vi Secretion ◽  
Interaction Partners ◽  
Bacterial Cytoplasm ◽  
Bacterial Effectors ◽  
Future Work

Pseudomonas aeruginosa is an opportunistic pathogen that can cause severe respiratory infections in people who are immunocompromised. P. aeruginosa possesses the Type VI Secretion System (T6SS), a bacterial weapon that injects effectors into neighbouring prokaryotes and eukaryotes. The T6SS is crucial for bacterial warfare, allowing P. aeruginosa to kill its competitors, which promotes its dominance in mixed microbial environments. P. aeruginosa has three T6SSs, H1/2/3-T6SS, these are structural homologs but deliver unique effectors. Effectors are delivered via the secreted component, a Hcp tube topped with a VgrG and PAAR spike. Only the first three identified effectors are delivered by Hcp1. Since then, there has been a bias in identification of VgrG or PAAR delivered effectors, mostly as these are encoded next to the spike proteins. Some P. aeruginosa effectors not only kill bacteria but have a dual role in pathogenesis. Our aim was to identify a comprehensive set of Hcp-delivered effectors for all three systems. Using Hcp1/2/3, systematic pull-down screens were performed to identify novel interaction partners. After confirming interaction, antibacterial toxicity was evaluated, identifying new Hcp delivered T6SS effectors for Hcp2 and Hcp3, which are toxic in the bacterial cytoplasm. These new anti-bacterial effectors may kill bacteria in novel ways, which could lead to novel antibiotics. Additionally, a toxin fusion proved too large for secretion and blocked the T6SS, revealing a Hcp-delivered effector size limit. Future work will focus on fully characterising these new toxins, as well as to look into the potential eukaryotic role of other interaction partners.

scholarly journals Effectors of the Stenotrophomonas maltophilia Type IV Secretion System Mediate Killing of Clinical Isolates of Pseudomonas aeruginosa

mBio ◽  
2021 ◽  
Author(s):  
Megan Y. Nas ◽  
Jeffrey Gabell ◽  
Nicholas P. Cianciotto
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Stenotrophomonas Maltophilia ◽  
Opportunistic Pathogen ◽  
Lung Infection ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Content Type ◽  
Type Iv

S. maltophilia is an increasingly important opportunistic pathogen. Inherently resistant to many antibiotics, S. maltophilia is often associated with lung infection, being, among other things, a complicating factor in cystic fibrosis patients.

scholarly journals Chelation of Membrane-Bound Cations by Extracellular DNA Activates the Type VI Secretion System in Pseudomonas aeruginosa

2016 ◽  
Vol 84 (8) ◽  
pp. 2355-2361 ◽  
Author(s):  
Mike Wilton ◽  
Megan J. Q. Wong ◽  
Le Tang ◽  
Xiaoye Liang ◽  
Richard Moore ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Adaptation Strategy ◽  
Secretion System ◽  
Extracellular Dna ◽  
Target Cells ◽  
Interspecies Interactions ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Membrane Bound

Pseudomonas aeruginosaemploys its type VI secretion system (T6SS) as a highly effective and tightly regulated weapon to deliver toxic molecules to target cells. T6SS-secreted proteins ofP. aeruginosacan be detected in the sputum of cystic fibrosis (CF) patients, who typically present a chronic and polymicrobial lung infection. However, the mechanism of T6SS activation in the CF lung is not fully understood. Here we demonstrate that extracellular DNA (eDNA), abundant within the CF airways, stimulates the dynamics of the H1-T6SS cluster apparatus inPseudomonas aeruginosaPAO1. Addition of Mg2+or DNase with eDNA abolished such activation, while treatment with EDTA mimicked the eDNA effect, suggesting that the eDNA-mediated effect is due to chelation of outer membrane-bound cations. DNA-activated H1-T6SS enablesP. aeruginosato nonselectively attack neighboring species regardless of whether or not it was provoked. Because of the importance of the T6SS in interspecies interactions and the prevalence of eDNA in the environments thatP. aeruginosainhabits, our report reveals an important adaptation strategy that likely contributes to the competitive fitness ofP. aeruginosain polymicrobial communities.

scholarly journals Differential Modulation of Quorum Sensing Signaling through QslA in Pseudomonas aeruginosa Strains PAO1 and PA14

2019 ◽  
Vol 201 (21) ◽  
Author(s):  
T. G. Sana ◽  
R. Lomas ◽  
M. R. Gimenez ◽  
A. Laubier ◽  
C. Soscia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Molecular Mechanisms ◽  
Opportunistic Pathogen ◽  
Secretion System ◽  
Type Ii Secretion ◽  
Type Vi Secretion System ◽  
Content Type

ABSTRACT Two clinical isolates of the opportunist pathogen Pseudomonas aeruginosa named PAO1 and PA14 are commonly studied in research laboratories. Despite the isolates being closely related, PA14 exhibits increased virulence compared to that of PAO1 in various models. To determine which players are responsible for the hypervirulence phenotype of the PA14 strain, we elected a transcriptomic approach through RNA sequencing. We found 2,029 genes that are differentially expressed between the two strains, including several genes that are involved with or regulated by quorum sensing (QS), known to control most of the virulence factors in P. aeruginosa. Among them, we chose to focus our study on QslA, an antiactivator of QS whose expression was barely detectable in the PA14 strain according our data. We hypothesized that lack of expression of qslA in PA14 could be responsible for higher QS expression in the PA14 strain, possibly explaining its hypervirulence phenotype. After confirming that QslA protein was highly produced in PAO1 but not in the PA14 strain, we obtained evidence showing that a PAO1 deletion strain of qslA has faster QS gene expression kinetics than PA14. Moreover, known virulence factors activated by QS, such as (i) pyocyanin production, (ii) H2-T6SS (type VI secretion system) gene expression, and (iii) Xcp-T2SS (type II secretion system) machinery production and secretion, were all lower in PAO1 than in PA14, due to higher qslA expression. However, biofilm formation and cytotoxicity toward macrophages, although increased in PA14 compared to PAO1, were independent of QslA control. Together, our findings implicated differential qslA expression as a major determinant of virulence factor expression in P. aeruginosa strains PAO1 and PA14. IMPORTANCE Pseudomonas aeruginosa is an opportunistic pathogen responsible for acute nosocomial infections and chronic pulmonary infections. P. aeruginosa strain PA14 is known to be hypervirulent in different hosts. Despite several studies in the field, the underlining molecular mechanisms sustaining this phenotype remain enigmatic. Here we provide evidence that the PA14 strain has faster quorum sensing (QS) kinetics than the PAO1 strain, due to the lack of QslA expression, an antiactivator of QS. QS is a major regulator of virulence factors in P. aeruginosa; therefore, we propose that the hypervirulent phenotype of the PA14 strain is, at least partially, due to the lack of QslA expression. This mechanism could be of great importance, as it could be conserved among other P. aeruginosa isolates.

scholarly journals Internalization of Pseudomonas aeruginosa Strain PAO1 into Epithelial Cells Is Promoted by Interaction of a T6SS Effector with the Microtubule Network

mBio ◽  
2015 ◽  
Vol 6 (3) ◽  
Author(s):  
Thibault G. Sana ◽  
Christoph Baumann ◽  
Andreas Merdes ◽  
Chantal Soscia ◽  
Thomas Rattei ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Bacterial Pathogen ◽  
Secretion System ◽  
Bacterial Protein ◽  
Type Vi Secretion System ◽  
Strain Pao1 ◽  
Content Type ◽  
Type Vi Secretion ◽  
Ring Complex

ABSTRACTInvasion of nonphagocytic cells through rearrangement of the actin cytoskeleton is a common immune evasion mechanism used by most intracellular bacteria. However, some pathogens modulate host microtubules as well by a still poorly understood mechanism. In this study, we aim at deciphering the mechanisms by which the opportunistic bacterial pathogenPseudomonas aeruginosainvades nonphagocytic cells, although it is considered mainly an extracellular bacterium. Using confocal microscopy and immunofluorescence, we show that the evolved VgrG2b effector ofP. aeruginosastrain PAO1 is delivered into epithelial cells by a type VI secretion system, called H2-T6SS, involving the VgrG2a component. Anin vivointeractome of VgrG2b in host cells allows the identification of microtubule components, including the γ-tubulin ring complex (γTuRC), a multiprotein complex catalyzing microtubule nucleation, as the major host target of VgrG2b. This interaction promotes a microtubule-dependent internalization of the bacterium since colchicine and nocodazole, two microtubule-destabilizing drugs, prevent VgrG2b-mediatedP. aeruginosaentry even if the invasion still requires actin. We further validate our findings by demonstrating that the type VI injection step can be bypassed by ectopic production of VgrG2b inside target cells prior to infection. Moreover, such uncoupling between VgrG2b injection and bacterial internalization also reveals that they constitute two independent steps. With VgrG2b, we provide the first example of a bacterial protein interacting with the γTuRC. Our study offers key insight into the mechanism of self-promoting invasion ofP. aeruginosainto human cells via a directed and specific effector-host protein interaction.IMPORTANCEInnate immunity and specifically professional phagocytic cells are key determinants in the ability of the host to controlP. aeruginosainfection. However, among various virulence strategies, including attack, this opportunistic bacterial pathogen is able to avoid host clearance by triggering its own internalization in nonphagocytic cells. We previously showed that a protein secretion/injection machinery, called the H2 type VI secretion system (H2-T6SS), promotesP. aeruginosauptake by epithelial cells. Here we investigate which H2-T6SS effector enablesP. aeruginosato enter nonphagocytic cells. We show that VgrG2b is delivered by the H2-T6SS machinery into epithelial cells, where it interacts with microtubules and, more particularly, with the γ-tubulin ring complex (γTuRC) known as the microtubule-nucleating center. This interaction precedes a microtubule- and actin-dependent internalization ofP. aeruginosa. We thus discovered an unprecedented target for a bacterial virulence factor since VgrG2b constitutes, to our knowledge, the first example of a bacterial protein interacting with the γTuRC.

scholarly journals The Type VI Secretion System Modulates Flagellar Gene Expression and Secretion in Citrobacter freundii and Contributes to Adhesion and Cytotoxicity to Host Cells

2015 ◽  
Vol 83 (7) ◽  
pp. 2596-2604 ◽  
Author(s):  
Liyun Liu ◽  
Shuai Hao ◽  
Ruiting Lan ◽  
Guangxia Wang ◽  
Di Xiao ◽  
...  
Keyword(s):  
Secretion System ◽  
Host Cells ◽  
Target Cells ◽  
Deletion Mutants ◽  
Citrobacter Freundii ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Mutant Strains

The type VI secretion system (T6SS) as a virulence factor-releasing system contributes to virulence development of various pathogens and is often activated upon contact with target cells.Citrobacter freundiistrain CF74 has a complete T6SS genomic island (GI) that containsclpV,hcp-2, andvgrT6SS genes. We constructedclpV,hcp-2,vgr, and T6SS GI deletion mutants in CF74 and analyzed their effects on the transcriptome overall and, specifically, on the flagellar system at the levels of transcription and translation. Deletion of the T6SS GI affected the transcription of 84 genes, with 15 and 69 genes exhibiting higher and lower levels of transcription, respectively. Members of the cell motility class of downregulated genes of the CF74ΔT6SS mutant were mainly flagellar genes, including effector proteins, chaperones, and regulators. Moreover, the production and secretion of FliC were also decreased inclpV,hcp-2,vgr, or T6SS GI deletion mutants in CF74 and were restored upon complementation. In swimming motility assays, the mutant strains were found to be less motile than the wild type, and motility was restored by complementation. The mutant strains were defective in adhesion to HEp-2 cells and were restored partially upon complementation. Further, the CF74ΔT6SS, CF74ΔclpV, and CF74Δhcp-2mutants induced lower cytotoxicity to HEp-2 cells than the wild type. These results suggested that the T6SS GI in CF74 regulates the flagellar system, enhances motility, is involved in adherence to host cells, and induces cytotoxicity to host cells. Thus, the T6SS plays a wide-ranging role inC. freundii.

scholarly journals Complete Genome Sequence of Pseudomonas aeruginosa Strain AA2 (LMG 27630), an Early Isolate Recovered from the Airway of a German Cystic Fibrosis Patient

2020 ◽  
Vol 9 (26) ◽  
Author(s):  
Andrea Sass ◽  
Tom Coenye
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Respiratory Tract ◽  
Cystic Fibrosis Patient ◽  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Opportunistic Pathogen ◽  
Content Type ◽  
Airway Infections ◽  
Early Isolate

ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen that is able to cause various infections, including airway infections in cystic fibrosis patients. Here, we present the complete closed and annotated genome sequence of P. aeruginosa AA2, an isolate obtained early during infection of the respiratory tract of a German cystic fibrosis patient.

scholarly journals Fur-Dam Regulatory Interplay at an Internal Promoter of the Enteroaggregative Escherichia coli Type VI Secretion sci1 Gene Cluster

2020 ◽  
Vol 202 (10) ◽  
Author(s):  
Yannick R. Brunet ◽  
Christophe S. Bernard ◽  
Eric Cascales
Keyword(s):  
Escherichia Coli ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Secretion System ◽  
Target Cells ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Internal Promoter ◽  
E Coli ◽  
Type Vi Secretion

ABSTRACT The type VI secretion system (T6SS) is a weapon for delivering effectors into target cells that is widespread in Gram-negative bacteria. The T6SS is a highly versatile machine, as it can target both eukaryotic and prokaryotic cells, and it has been proposed that T6SSs are adapted to the specific needs of each bacterium. The expression of T6SS gene clusters and the activation of the secretion apparatus are therefore tightly controlled. In enteroaggregative Escherichia coli (EAEC), the sci1 T6SS gene cluster is subject to a complex regulation involving both the ferric uptake regulator (Fur) and DNA adenine methylase (Dam)-dependent DNA methylation. In this study, an additional, internal, promoter was identified within the sci1 gene cluster using +1 transcriptional mapping. Further analyses demonstrated that this internal promoter is controlled by a mechanism strictly identical to that of the main promoter. The Fur binding box overlaps the −10 transcriptional element and a Dam methylation site, GATC-32. Hence, the expression of the distal sci1 genes is repressed and the GATC-32 site is protected from methylation in iron-rich conditions. The Fur-dependent protection of GATC-32 was confirmed by an in vitro methylation assay. In addition, the methylation of GATC-32 negatively impacted Fur binding. The expression of the sci1 internal promoter is therefore controlled by iron availability through Fur regulation, whereas Dam-dependent methylation maintains a stable ON expression in iron-limited conditions. IMPORTANCE Bacteria use weapons to deliver effectors into target cells. One of these weapons, the type VI secretion system (T6SS), assembles a contractile tail acting as a spring to propel a toxin-loaded needle. Its expression and activation therefore need to be tightly regulated. Here, we identified an internal promoter within the sci1 T6SS gene cluster in enteroaggregative E. coli. We show that this internal promoter is controlled by Fur and Dam-dependent methylation. We further demonstrate that Fur and Dam compete at the −10 transcriptional element to finely tune the expression of T6SS genes. We propose that this elegant regulatory mechanism allows the optimum production of the T6SS in conditions where enteroaggregative E. coli encounters competing species.

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