A mathematical model for quorum sensing in Pseudomonas aeruginosa

The bacteria Pseudomonas aeruginosa use the size and density of their colonies to regulate the production of a large variety of substances, including toxins. This phenomenon, called quorum sensing, apparently enables colonies to grow to sufficient size undetected by the immune system of the host organism. In this paper, we present a mathematical model of quorum sensing in P. aeruginosa that is based on the known biochemistry of regulation of the autoinducer that is crucial to this signalling mechanism. Using this model we show that quorum sensing works because of a biochemical switch between two stable steady solutions, one with low levels of autoinducer and one with high levels of autoinducer.     

The Influence of Fluid Flow on Modeling Quorum Sensing in Bacterial Biofilms

In this paper, we study quorum sensing in Pseudomonas aeruginosa biofilms. Quorum sensing is a process where bacteria monitor their population density through the release of extra-cellular signalling molecules. The presence of these molecules affects gene modulation leading to changes in behaviour such as the release of virulence factors. Here, we use numerical methods to approximate a 2-D model of quorum sensing. It is observed that the shape of the biofilm can have a profound effect on the onset of quorum sensing. This has serious repercussions for experimental observations since biofilms of the same biomass but different shapes can produce quite different results.     

A Quadruple Knockout of lasIR and rhlIR of Pseudomonas aeruginosa PAO1 That Retains Wild-Type Twitching Motility Has Equivalent Infectivity and Persistence to PAO1 in a Mouse Model of Lung Infection

It has been widely reported that quorum-sensing incapable strains of Pseudomonas aeruginosa are less virulent than wild type strains. However, quorum sensing mutants of P. aeruginosa have been shown to develop other spontaneous mutations under prolonged culture conditions, and one of the phenotypes of P. aeruginosa that is frequently affected by this phenomenon is type IV pili-dependent motility, referred to as twitching motility. As twitching motility has been reported to be important for adhesion and colonisation, we aimed to generate a quorum-sensing knockout for which the heritage was recorded and the virulence factor production in areas unrelated to quorum sensing was known to be intact. We created a lasIRrhlIR quadruple knockout in PAO1 using a published technique that allows for the deletion of antibiotic resistance cartridges following mutagenesis, to create an unmarked QS knockout of PAO1, thereby avoiding the need for use of antibiotics in culturing, which can have subtle effects on bacterial phenotype. We phenotyped this mutant demonstrating that it produced reduced levels of protease and elastase, barely detectable levels of pyoverdin and undetectable levels of the quorum sensing signal molecules N-3-oxododecanoly-L-homoserine lactone and N-butyryl homoserine lactone, but retained full twitching motility. We then used a mouse model of acute lung infection with P. aeruginosa to demonstrate that the lasIRrhlIR knockout strain showed equal persistence to wild type parental PAO1, induced equal or greater neutrophil infiltration to the lungs, and induced similar levels of expression of inflammatory cytokines in the lungs and similar antibody responses, both in terms of magnitude and isotype. Our results suggest, in contrast to previous reports, that lack of quorum sensing alone does not significantly affect the immunogenicity, infectiveness and persistence of P. aeruginosa in a mouse model of acute lung infection.     

The dependence of quorum sensing on the depth of a growing biofilm

In a process called quorum sensing, bacteria monitor their population density via extracellular signaling molecules and modulate gene expression accordingly. In this paper, a one-dimensional model of a growing Pseudomonas aeruginosa biofilm is examined. Quorum sensing has been included in the model through equations describing the production, degradation, and diffusion of the signaling molecules, acyl-homoserine lactones, in the biofilm. From this model, we are able to make some important observations about quorum sensing. First, in order for quorum sensing to initiate near the substratum, in accordance with experimental observations, the model suggests that cells in oxygen-deficient regions of the biofilm must still be synthesizing the signal compound. Second, the induction of quorum sensing is related to a critical biofilm depth; once the biofilm grows to the critical depth, quorum sensing is induced. Third, the critical biofilm depth varies with the pH of the surrounding fluid. Of particular interest is the prediction of a critical pH threshold, above which quorum sensing is not possible at any depth. These results highlight the importance of careful study of the relationship among metabolic activity of the bacterium, signal synthesis, and the chemistry of the surrounding environment.     

A mathematical model of quorum sensing in a growing bacterial biofilm

In a process called quorum sensing, bacteria monitor their population density via extracellular signaling molecules and modulate gene expression accordingly. This paper describes a one-dimensional model of a growing Pseudomonas aeruginosa biofilm. Quorum sensing has been included in the model by the addition of equations describing the production, degradation, and diffusion of acyl-homoserine lactones in the biofilm. In order for quorum sensing to initiate near the substratum, in accordance with experimental observations, model results suggest that cells in oxygen-deficient regions of the biofilm must still be synthesizing the signal compound. This result highlights the importance of careful study of the relationship between metabolic activity of the bacterium and signal synthesis. Received 11 March 2002/ Accepted in revised form 01 August 2002     

Bacterial quorum‐sensing signal IQS induces host cell apoptosis by targeting POT1–p53 signalling pathway

Pseudomonas aeruginosa, an opportunistic life‐threatening human bacterial pathogen, employs quorum‐sensing (QS) signal molecules to modulate virulence gene expression. 2‐(2‐hydroxyphenyl)‐thiazole‐4‐carbaldehyde (IQS) is a recently identified QS signal that integrates the canonical lasR‐type QS of P. aeruginosa and host phosphate stress response to fine‐tune its virulence production for a successful infection. To address the role of IQS in pathogen–host interaction, we here present that IQS inhibits host cell growth and stimulates apoptosis in a dosage‐dependent manner. By downregulating the telomere‐protecting protein POT1 in host cells, IQS activates CHK1, CHK2, and p53 in an Ataxia telangiectasia mutated (ATM)/ATM and RAD3‐related (ATR)‐dependent manner and induces DNA damage response. Overexpression of POT1 in host cells presents a resistance to IQS treatment. These results suggest a pivotal role of IQS in host apoptosis, highlighting the complexity of pathogenesis mechanisms developed by P. aeruginosa during infection.     

The effect of Quorum sensing inhibitors on the evolution of CRISPR-based phage immunity in Pseudomonas aeruginosa

Quorum sensing controls the expression of a wide range of important traits in the opportunistic pathogen Pseudomonas aeruginosa, including the expression of virulence genes and its CRISPR-cas immune system, which protects from bacteriophage (phage) infection. This finding has led to the speculation that synthetic quorum sensing inhibitors could be used to limit the evolution of CRISPR immunity during phage therapy. Here we use experimental evolution to explore if and how a quorum sensing inhibitor influences the population and evolutionary dynamics of P. aeruginosa upon phage DMS3vir infection. We find that chemical inhibition of quorum sensing decreases phage adsorption rates due to downregulation of the Type IV pilus, which causes delayed lysis of bacterial cultures and favours the evolution of CRISPR immunity. Our data therefore suggest that inhibiting quorum sensing may reduce rather than improve the therapeutic efficacy of pilus-specific phage, and this is likely a general feature when phage receptors are positively regulated by quorum sensing.Subject terms: Microbiology, Antimicrobials     

Alteration of protein homeostasis mediates the interaction of Pseudomonas aeruginosa with Staphylococcus aureus

Intracellular protein degradation is essential for the survival of all organisms, but its role in interspecies interaction is unknown. Here, we show that the ClpXP protease of Pseudomonas aeruginosa suppresses its antimicrobial activity against Staphylococcus aureus, a common pathogen co-isolated with P. aeruginosa from polymicrobial human infections. Using proteomic, biochemical, and molecular genetic approaches, we found that this effect is due to the inhibitory effects of ClpXP on the quorum sensing (QS) of P. aeruginosa, mainly by degrading proteins (e.g., PhnA, PhnB, PqsR, and RhlI) which are critical for the production of QS signal molecules PQS and C4-HSL. We provide evidence that co-culturing with S. aureus induces a decrease in the activity of ClpXP in P. aeruginosa, an effect which was also achieved by the treatment of P. aeruginosa with N-acetylglucosamine (GlcNAc), a widespread chemical present on the surface of diverse cell types from bacteria to humans. These findings extend the range of biological events governed by proteolytic machinery to microbial community structure, thus also suggesting that a chemical-induced alteration of protein homeostasis is a mechanism for interspecies interactions.     

Mechanistic understanding of Phenyllactic acid mediated inhibition of quorum sensing and biofilm development in Pseudomonas aeruginosa

Pseudomonas aeruginosa depends on its quorum sensing (QS) system for its virulence factors’ production and biofilm formation. Biofilms of P. aeruginosa on the surface of indwelling catheters are often resistant to antibiotic therapy. Alternative approaches that employ QS inhibitors alone or in combination with antibiotics are being developed to tackle P. aeruginosa infections. Here, we have studied the mechanism of action of 3-Phenyllactic acid (PLA), a QS inhibitory compound produced by Lactobacillus species, against P. aeruginosa PAO1. Our study revealed that PLA inhibited the expression of virulence factors such as pyocyanin, protease, and rhamnolipids that are involved in the biofilm formation of P. aeruginosa PAO1. Swarming motility, another important criterion for biofilm formation of P. aeruginosa PAO1, was also inhibited by PLA. Gene expression, mass spectrometric, functional complementation assays, and in silico data indicated that the quorum quenching and biofilm inhibitory activities of PLA are attributed to its ability to interact with P. aeruginosa QS receptors. PLA antagonistically binds to QS receptors RhlR and PqsR with a higher affinity than its cognate ligands N-butyryl-l-homoserine lactone (C₄–HSL) and 2-heptyl-3,4-dihydroxyquinoline (PQS; Pseudomonas quinolone signal). Using an in vivo intraperitoneal catheter-associated medaka fish infection model, we proved that PLA inhibited the initial attachment of P. aeruginosa PAO1 on implanted catheter tubes. Our in vitro and in vivo results revealed the potential of PLA as anti-biofilm compound against P. aeruginosa.

     

Pseudomonas aeruginosa possesses three distinct systems for sensing and using the host molecule haem

Pathogens have developed several strategies to obtain iron during infection, including the use of iron-containing molecules from the host. Haem accounts for the vast majority of the iron pool in vertebrates and thus represents an important source of iron for pathogens. Using a proteomic approach, we have identified in this work a previously uncharacterized system, which we name Hxu, that together with the known Has and Phu systems, is used by the human pathogen Pseudomonas aeruginosa to respond to haem. We show that the Has and Hxu systems are functional signal transduction pathways of the cell-surface signalling class and report the mechanism triggering the activation of these signalling systems. Both signalling cascades involve an outer membrane receptor (HasR and HxuA respectively) that upon sensing haem in the extracellular medium produces the activation of an σ^ECF factor in the cytosol. HxuA has a major role in signalling and a minor role in haem acquisition in conditions in which the HasR and PhuR receptors or other sources of iron are present. Remarkably, P. aeruginosa compensates the lack of the HasR receptor by increasing the production of HxuA, which underscores the importance of haem signalling for this pathogen.     

Heterogeneous virulence potential and high antibiotic resistance of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> strains isolated from Korean pneumonia patients

Pseudomonas aeruginosa is an opportunistic human pathogen of clinical importance that causes airway infections in immunocompromised patients. Here, we report the virulence-associated characteristics of strains of P. aeruginosa, isolated from the sputa of 25 Korean pneumonia patients. A high degree of genomic plasticity was observed by random amplified polymorphic DNA genotype analysis, suggesting that the infections were caused by strains with diverse genomic backgrounds. Biofilm formation of each isolate was heterogeneous in terms of their relative motilities. In addition, 48% of isolates were defective in the production of 3-oxo-C₁₂-HSL (PAI-1), a quorum sensing signal molecule. In these strains, PAI-1-dependent elastase production was correspondingly decreased, suggesting that a large number of strains were presumed to be quorum sensing deficient. Multidrug resistance (MDR) was seen in 56% of the isolates tested, and 44% of the MDR strains were resistant to five or more antibiotics. Taken together, our results provide additional insights into the virulence traits of P. aeruginosa clinical isolates, which will aid in treating P. aeruginosa infections in pneumonia patients.     

Revisiting the virulence hallmarks of Pseudomonas aeruginosa: a chronicle through the perspective of quorum sensing

Pseudomonas aeruginosa is an opportunistic pathogen and the leading cause of mortality among immunocompromised patients in clinical setups. The hallmarks of virulence in P. aeruginosa encompass six biologically competent attributes that cumulatively drive disease progression in a multistep manner. These multifaceted hallmarks lay the principal foundation for rationalizing the complexities of pseudomonal infections. They include factors for host colonization and bacterial motility, biofilm formation, production of destructive enzymes, toxic secondary metabolites, iron-chelating siderophores and toxins. This arsenal of virulence hallmarks is fostered and stringently regulated by the bacterial signalling system called quorum sensing (QS). The central regulatory functions of QS in controlling the timely expression of these virulence hallmarks for adaptation and survival drive the disease outcome. This review describes the intricate mechanisms of QS in P. aeruginosa and its role in shaping bacterial responses, boosting bacterial fitness. We summarize the virulence hallmarks of P. aeruginosa, relating them with the QS circuitry in clinical infections. We also examine the role of QS in the development of drug resistance and propose a novel antivirulence therapy to combat P. aeruginosa infections. This can prove to be a next-generation therapy that may eventually become refractory to the use of conventional antimicrobial treatments.     

Monitoring of Vibrio harveyi quorum sensing activity in real time during infection of brine shrimp larvae

Quorum sensing, bacterial cell-to-cell communication, has been linked to the virulence of pathogenic bacteria. Indeed, in vitro experiments have shown that many bacterial pathogens regulate the expression of virulence genes by this cell-to-cell communication process. Moreover, signal molecules have been detected in samples retrieved from infected hosts and quorum sensing disruption has been reported to result in reduced virulence in different host–pathogen systems. However, data on in vivo quorum sensing activity of pathogens during infection of a host are currently lacking. We previously reported that quorum sensing regulates the virulence of Vibrio harveyi in a standardised model system with gnotobiotic brine shrimp (Artemia franciscana) larvae. Here, we monitored quorum sensing activity in Vibrio harveyi during infection of the shrimp, using bioluminescence as a read-out. We found that wild-type Vibrio harveyi shows a strong increase in quorum sensing activity early during infection. In this respect, the bacteria behave remarkably similar in different larvae, despite the fact that only half of them survive the infection. Interestingly, when expressed per bacterial cell, Vibrio harveyi showed around 200-fold higher maximal quorum sensing-regulated bioluminescence when associated with larvae than in the culture water. Finally, the in vivo quorum sensing activity of mutants defective in the production of one of the three signal molecules is consistent with their virulence, with no detectable in vivo quorum sensing activity in AI-2- and CAI-1-deficient mutants. These results indicate that AI-2 and CAI-1 are the dominant signals during infection of brine shrimp.     

Aspergillus ochraceopetaliformis SSP13 modulates quorum sensing regulated virulence and biofilm formation in Pseudomonas aeruginosa PAO1

Pseudomonas aeruginosa is an opportunistic nosocomial pathogen causing the majority of acute and persistent infections in human beings. The ability to form biofilm adds a new dimension to its resistance to conventional therapeutic agents. In the present study, down-regulation of quorum sensing regulated virulence and biofilm development resulting from exposure to Aspergillus ochraceopetaliformis SSP13 extract was investigated. The in vitro results inferred impairment in the production of LasA protease, LasB elastase, chitinase, pyocyanin, exopolysaccharides and rhamnolipids. In addition, motility and biofilm formation by P. aeruginosa PAO1 was significantly altered. The in vitro results were further supported by molecular docking studies of the metabolites obtained from GC-MS analysis depicting the quorum sensing attenuation by targeting the receptor proteins LasR and RhlR. The in vitro and in silico studies suggested new avenues for the development of bioactive metabolites from A. ochraceopetaliformis SSP13 extract as potential anti-infective agents.     

Detection of quorum-sensing-related molecules in <Emphasis Type="Italic">Vibrio scophthalmi</Emphasis>

Background  

Cell-to-cell communication (also referred to as quorum sensing) based on N-acyl-homoserine lactones (AHLs) is a widespread response to environmental change in Gram-negative bacteria. AHLs seem to be highly variable, both in terms of the acyl chain length and in the chemical structure of the radicals. Another quorum sensing pathway, the autoinducer-2-based system, is present both in Gram-positive and Gram-negative bacteria. In this study the presence of signal molecules belonging to both quorum sensing signalling pathways was analysed in the marine symbiotic species Vibrio scophthalmi.