Effect of Two Unique Nanoparticle Formulations on the Efficacy of a Broadly Protective Vaccine Against Pseudomonas Aeruginosa

Pseudomonas aeruginosa is an opportunistic pathogen responsible for a wide range of infections in humans. In addition to its innate antibiotic resistance, P. aeruginosa is very effective in acquiring resistance resulting in the emergence of multi-drug resistance strains and a licensed vaccine is not yet available. We have previously demonstrated the protective efficacy of a novel antigen PaF (Pa Fusion), a fusion of the type III secretion system (T3SS) needle tip protein, PcrV, and the first of two translocator proteins, PopB. PaF was modified to provide a self-adjuvanting activity by fusing the A1 subunit of the heat-labile enterotoxin from Enterotoxigenic E. coli to its N-terminus to give L-PaF. In addition to providing protection against 04 and 06 serotypes of P. aeruginosa, L-PaF elicited opsonophagocytic killing and stimulated IL-17A secretion, which have been predicted to be required for a successful vaccine. While monomeric recombinant subunit vaccines can be protective in mice, this protection often does not transfer to humans where multimeric formulations perform better. Here, we use two unique formulations, an oil-in-water (o/w) emulsion and a chitosan particle, as well as the addition of a unique TLR4 agonist, BECC438 (a detoxified lipid A analogue designated Bacterial Enzymatic Combinatorial Chemistry 438), as an initial step in optimizing L-PaF for use in humans. The o/w emulsion together with BECC438 provided the best protective efficacy, which correlated with high levels of opsonophagocytic killing and IL-17A secretion, thereby reducing the lung burden among all the vaccinated groups tested.

Download Full-text

Development of Novel Protective Polyvalent Irradiated Pseudomonas aeruginosa Vaccine for Immuno-Compromised Patients

International Journal of Pharmacology Phytochemistry and Ethnomedicine ◽

10.18052/www.scipress.com/ijppe.16.1 ◽

2021 ◽

Vol 16 ◽

pp. 1-10

Author(s):

Manal M.E. Ahmed ◽

Jakeen Eljakee ◽

Tarek Mahran

Keyword(s):

Pseudomonas Aeruginosa ◽

Protective Efficacy ◽

Challenge Test ◽

Opportunistic Pathogen ◽

Effective Vaccine ◽

The Novel ◽

Therapeutic Approaches ◽

Promising Strategy ◽

Antigenic Expression ◽

Compromised Patients

Pseudomonas aeruginosa is an opportunistic pathogen affecting immuno-compromised patients; however, no effective vaccine is currently available in the market. Here, we developed novel polyvalent irradiated P. aeruginosa vaccine using cobalt 60 that inhibited pathogen viability but retained antigenic expression functionally. Mice were vaccinated by the developed vaccine by intranasal, intramuscular and subcutaneous route of administration followed by challenge test. The protective efficacy of the novel vaccine reached up to 95%. This significant protection was mainly associated with measurable antiserum opsonic killing activity. In conclusion, the novel vaccine provides a promising strategy of both prophylactic and therapeutic approaches for immuno-compromised patients against MDR P. aeruginosa.

Download Full-text

The Role of Pseudomonas aeruginosa Lipopolysaccharide in Bacterial Pathogenesis and Physiology

Pathogens ◽

10.3390/pathogens9010006 ◽

2019 ◽

Vol 9 (1) ◽

pp. 6 ◽

Cited By ~ 3

Author(s):

Steven M. Huszczynski ◽

Joseph S. Lam ◽

Cezar M. Khursigara

Keyword(s):

Pseudomonas Aeruginosa ◽

Lipid A ◽

Opportunistic Pathogen ◽

Major Constituent ◽

Bacterial Survival ◽

Core Oligosaccharide ◽

Persistent Infections ◽

The Relationship ◽

Extracellular Environment

The major constituent of the outer membrane of Gram-negative bacteria is lipopolysaccharide (LPS), which is comprised of lipid A, core oligosaccharide, and O antigen, which is a long polysaccharide chain extending into the extracellular environment. Due to the localization of LPS, it is a key molecule on the bacterial cell wall that is recognized by the host to deploy an immune defence in order to neutralize invading pathogens. However, LPS also promotes bacterial survival in a host environment by protecting the bacteria from these threats. This review explores the relationship between the different LPS glycoforms of the opportunistic pathogen Pseudomonas aeruginosa and the ability of this organism to cause persistent infections, especially in the genetic disease cystic fibrosis. We also discuss the role of LPS in facilitating biofilm formation, antibiotic resistance, and how LPS may be targeted by new antimicrobial therapies.

Download Full-text

Transcriptomic Analysis of the Sulfate Starvation Response of Pseudomonas aeruginosa

Journal of Bacteriology ◽

10.1128/jb.00889-07 ◽

2007 ◽

Vol 189 (19) ◽

pp. 6743-6750 ◽

Cited By ~ 47

Author(s):

Tewes Tralau ◽

Stéphane Vuilleumier ◽

Christelle Thibault ◽

Barry J. Campbell ◽

C. Anthony Hart ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Stress Proteins ◽

Large Family ◽

Opportunistic Pathogen ◽

Transport Systems ◽

Sulfur Source ◽

Secretion Systems ◽

Starvation Response ◽

Wide Range

ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen that causes a number of infections in humans, but is best known for its association with cystic fibrosis. It is able to use a wide range of sulfur compounds as sources of sulfur for growth. Gene expression in response to changes in sulfur supply was studied in P. aeruginosa E601, a cystic fibrosis isolate that displays mucin sulfatase activity, and in P. aeruginosa PAO1. A large family of genes was found to be upregulated by sulfate limitation in both isolates, encoding sulfatases and sulfonatases, transport systems, oxidative stress proteins, and a sulfate-regulated TonB/ExbBD complex. These genes were localized in five distinct islands on the genome and encoded proteins with a significantly reduced content of cysteine and methionine. Growth of P. aeruginosa E601 with mucin as the sulfur source led not only to a sulfate starvation response but also to induction of genes involved with type III secretion systems.

Download Full-text

Mutational Analysis of RetS, an Unusual Sensor Kinase-Response Regulator Hybrid Required for Pseudomonas aeruginosa Virulence

Infection and Immunity ◽

10.1128/iai.00575-06 ◽

2006 ◽

Vol 74 (8) ◽

pp. 4462-4473 ◽

Cited By ~ 53

Author(s):

Michelle A. Laskowski ◽

Barbara I. Kazmierczak

Keyword(s):

Pseudomonas Aeruginosa ◽

Mutational Analysis ◽

Response Regulator ◽

Acute Infection ◽

Opportunistic Pathogen ◽

Sensor Kinase ◽

Chronic Infections ◽

Reciprocal Regulation ◽

Wide Range

ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen capable of causing both acute and chronic infections in a wide range of hosts. Expression of the type III secretion system (T3SS) proteins is correlated with virulence in models of acute infection, while downregulation of the T3SS and upregulation of genes important for biofilm formation are observed during chronic infections. RetS, a hybrid sensor kinase-response regulator protein of P. aeruginosa, plays a key role in the reciprocal regulation of virulence factors required for acute versus chronic infection and is postulated to act in concert with two other sensor kinase-response regulator hybrids, GacS and LadS. This work examines the roles of the putative sensing and signal transduction domains of RetS in induction of the T3SS in vitro and in a murine model of acute pneumonia. We identify distinct signaling roles for the tandem receiver domains of RetS and present evidence suggesting that RetS may serve as a substrate for another sensor kinase. Phenotypes associated with RetS alleles lacking periplasmic and/or transmembrane domains further indicate that the periplasmic domain of RetS may transmit a signal that inhibits RetS activity during acute infections.

Download Full-text

Cell envelope proteases and peptidases of Pseudomonas aeruginosa: multiple roles, multiple mechanisms

FEMS Microbiology Reviews ◽

10.1093/femsre/fuaa036 ◽

2020 ◽

Vol 44 (6) ◽

pp. 857-873

Author(s):

Astra Heywood ◽

Iain L Lamont

Keyword(s):

Pseudomonas Aeruginosa ◽

Outer Membrane ◽

Protein Quality ◽

Environmental Changes ◽

Cell Envelope ◽

Opportunistic Pathogen ◽

Multiple Roles ◽

Current State ◽

Wide Range ◽

Membrane Cell

ABSTRACT Pseudomonas aeruginosa is a Gram-negative bacterium that is commonly isolated from damp environments. It is also a major opportunistic pathogen, causing a wide range of problematic infections. The cell envelope of P. aeruginosa, comprising the cytoplasmic membrane, periplasmic space, peptidoglycan layer and outer membrane, is critical to the bacteria's ability to adapt and thrive in a wide range of environments. Over 40 proteases and peptidases are located in the P. aeruginosa cell envelope. These enzymes play many crucial roles. They are required for protein secretion out of the cytoplasm to the periplasm, outer membrane, cell surface or the environment; for protein quality control and removal of misfolded proteins; for controlling gene expression, allowing adaptation to environmental changes; for modification and remodelling of peptidoglycan; and for metabolism of small molecules. The key roles of cell envelope proteases in ensuring normal cell functioning have prompted the development of inhibitors targeting some of these enzymes as potential new anti-Pseudomonas therapies. In this review, we summarise the current state of knowledge across the breadth of P. aeruginosa cell envelope proteases and peptidases, with an emphasis on recent findings, and highlight likely future directions in their study.

Download Full-text

Discovery of an inhibitor of the production of the Pseudomonas aeruginosa virulence factor pyocyanin in wild-type cells

Beilstein Journal of Organic Chemistry ◽

10.3762/bjoc.12.137 ◽

2016 ◽

Vol 12 ◽

pp. 1428-1433 ◽

Cited By ~ 8

Author(s):

Bernardas Morkunas ◽

Balint Gal ◽

Warren R J D Galloway ◽

James T Hodgkinson ◽

Brett M Ibbeson ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Cell Signalling ◽

Opportunistic Pathogen ◽

Wild Type ◽

Wide Range ◽

Potential Applications ◽

Therapeutic Context ◽

Pyocyanin Production

Pyocyanin is a small molecule produced by Pseudomonas aeruginosa that plays a crucial role in the pathogenesis of infections by this notorious opportunistic pathogen. The inhibition of pyocyanin production has been identified as an attractive antivirulence strategy for the treatment of P. aeruginosa infections. Herein, we report the discovery of an inhibitor of pyocyanin production in cultures of wild-type P. aeruginosa which is based around a 4-alkylquinolin-2(1H)-one scaffold. To the best of our knowledge, this is the first reported example of pyocyanin inhibition by a compound based around this molecular framework. The compound may therefore be representative of a new structural sub-class of pyocyanin inhibitors, which could potentially be exploited in in a therapeutic context for the development of critically needed new antipseudomonal agents. In this context, the use of wild-type cells in this study is notable, since the data obtained are of direct relevance to native situations. The compound could also be of value in better elucidating the role of pyocyanin in P. aeruginosa infections. Evidence suggests that the active compound reduces the level of pyocyanin production by inhibiting the cell–cell signalling mechanism known as quorum sensing. This could have interesting implications; quorum sensing regulates a range of additional elements associated with the pathogenicity of P. aeruginosa and there is a wide range of other potential applications where the inhibition of quorum sensing is desirable.

Download Full-text

A Large-Scale Whole-Genome Comparison Shows that Experimental Evolution in Response to Antibiotics Predicts Changes in Naturally Evolved Clinical Pseudomonas aeruginosa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01619-19 ◽

2019 ◽

Vol 63 (12) ◽

Cited By ~ 6

Author(s):

Samuel J. T. Wardell ◽

Attika Rehman ◽

Lois W. Martin ◽

Craig Winstanley ◽

Wayne M. Patrick ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Experimental Evolution ◽

Large Scale ◽

Opportunistic Pathogen ◽

Clinical Isolates ◽

Genome Comparison ◽

Chronic Infections ◽

Content Type ◽

Large Deletions ◽

Wide Range

ABSTRACT Pseudomonas aeruginosa is an opportunistic pathogen that causes a wide range of acute and chronic infections. An increasing number of isolates have mutations that make them antibiotic resistant, making treatment difficult. To identify resistance-associated mutations, we experimentally evolved the antibiotic-sensitive strain P. aeruginosa PAO1 to become resistant to three widely used antipseudomonal antibiotics, namely, ciprofloxacin, meropenem, and tobramycin. Mutants could tolerate up to 2,048-fold higher concentrations of antibiotics than strain PAO1. Genome sequences were determined for 13 mutants for each antibiotic. Each mutant had between 2 and 8 mutations. For each antibiotic, at least 8 genes were mutated in multiple mutants, demonstrating the genetic complexity of resistance. For all three antibiotics, mutations arose in genes known to be associated with resistance but also in genes not previously associated with resistance. To determine the clinical relevance of mutations uncovered in this study, we analyzed the corresponding genes in 558 isolates of P. aeruginosa from patients with chronic lung disease and in 172 isolates from the general environment. Many genes identified through experimental evolution had predicted function-altering changes in clinical isolates but not in environmental isolates, showing that mutated genes in experimentally evolved bacteria can predict those that undergo mutation during infection. Additionally, large deletions of up to 479 kb arose in experimentally evolved meropenem-resistant mutants, and large deletions were present in 87 of the clinical isolates. These findings significantly advance understanding of antibiotic resistance in P. aeruginosa and demonstrate the validity of experimental evolution in identifying clinically relevant resistance-associated mutations.

Download Full-text

Roles of Two-Component Systems in Pseudomonas aeruginosa Virulence

International Journal of Molecular Sciences ◽

10.3390/ijms222212152 ◽

2021 ◽

Vol 22 (22) ◽

pp. 12152

Author(s):

Maria Sultan ◽

Rekha Arya ◽

Kyeong Kyu Kim

Keyword(s):

Pseudomonas Aeruginosa ◽

Virulence Factors ◽

Opportunistic Pathogen ◽

Secretion Systems ◽

Chronic Infections ◽

Potential Threat ◽

Wide Range ◽

Component Systems ◽

Two Component ◽

Two Component Systems

Pseudomonas aeruginosa is an opportunistic pathogen that synthesizes and secretes a wide range of virulence factors. P. aeruginosa poses a potential threat to human health worldwide due to its omnipresent nature, robust host accumulation, high virulence, and significant resistance to multiple antibiotics. The pathogenicity of P. aeruginosa, which is associated with acute and chronic infections, is linked with multiple virulence factors and associated secretion systems, such as the ability to form and utilize a biofilm, pili, flagella, alginate, pyocyanin, proteases, and toxins. Two-component systems (TCSs) of P. aeruginosa perform an essential role in controlling virulence factors in response to internal and external stimuli. Therefore, understanding the mechanism of TCSs to perceive and respond to signals from the environment and control the production of virulence factors during infection is essential to understanding the diseases caused by P. aeruginosa infection and further develop new antibiotics to treat this pathogen. This review discusses the important virulence factors of P. aeruginosa and the understanding of their regulation through TCSs by focusing on biofilm, motility, pyocyanin, and cytotoxins.

Download Full-text

β-lactam Resistance in Pseudomonas aeruginosa: Current Status, Future Prospects

Pathogens ◽

10.3390/pathogens10121638 ◽

2021 ◽

Vol 10 (12) ◽

pp. 1638

Author(s):

Karl A. Glen ◽

Iain L. Lamont

Keyword(s):

Pseudomonas Aeruginosa ◽

Cell Division ◽

Biofilm Formation ◽

Opportunistic Pathogen ◽

Current Status ◽

Future Prospects ◽

Penicillin Binding Protein ◽

Chronic Infections ◽

Penicillin Binding Proteins ◽

Wide Range

Pseudomonas aeruginosa is a major opportunistic pathogen, causing a wide range of acute and chronic infections. β-lactam antibiotics including penicillins, carbapenems, monobactams, and cephalosporins play a key role in the treatment of P. aeruginosa infections. However, a significant number of isolates of these bacteria are resistant to β-lactams, complicating treatment of infections and leading to worse outcomes for patients. In this review, we summarize studies demonstrating the health and economic impacts associated with β-lactam-resistant P. aeruginosa. We then describe how β-lactams bind to and inhibit P. aeruginosa penicillin-binding proteins that are required for synthesis and remodelling of peptidoglycan. Resistance to β-lactams is multifactorial and can involve changes to a key target protein, penicillin-binding protein 3, that is essential for cell division; reduced uptake or increased efflux of β-lactams; degradation of β-lactam antibiotics by increased expression or altered substrate specificity of an AmpC β-lactamase, or by the acquisition of β-lactamases through horizontal gene transfer; and changes to biofilm formation and metabolism. The current understanding of these mechanisms is discussed. Lastly, important knowledge gaps are identified, and possible strategies for enhancing the effectiveness of β-lactam antibiotics in treating P. aeruginosa infections are considered.

Download Full-text

Serratia marcescens RamA expression is under PhoP-dependent control and modulates lipid A-related genes transcription and antibiotic resistance phenotypes

Journal of Bacteriology ◽

10.1128/jb.00523-20 ◽

2021 ◽

Author(s):

Javier F. Mariscotti ◽

Eleonora García Véscovi

Keyword(s):

Antibiotic Resistance ◽

Serratia Marcescens ◽

Nalidixic Acid ◽

Lipid A ◽

Efflux Pump ◽

Transcriptional Regulator ◽

Opportunistic Pathogen ◽

Binding Motif ◽

Promoter Regions ◽

Wide Range

Serratia marcescens is an enteric bacterium that can function as an opportunistic pathogen with with increasing incidence in clinical settings. This is mainly due to the ability of express a wide range of virulence factors and the acquisition of antibiotic resistance mechanisms. For these reasons, S. marcescens has been declared by the WHO as a research priority to develop alternative antimicrobial strategies. In this work, we found a PhoP-binding motif in the promoter region of transcriptional regulator RamA of the S. marcescens RM66262. We demonstrated that the expression of ramA is autoregulated and that ramA is also part of the PhoP/PhoQ regulon. We have also shown that PhoP binds directly and specifically to ramA, mgtE1, mgtE2, lpxO1 and lpxO2 promoter regions and that RamA binds to ramA and lpxO1 but not to mgtE1 and lpxO2, suggesting an indirect control for these latter genes. Finally, we have demonstrated that, in S. marcescens, the RamA overexpression induces the AcrAB-TolC efflux pump required to reduce the susceptibility of the bacteria to tetracycline and nalidixic acid. In sum, we herein show the first report describing the regulation of ramA under the PhoP/PhoQ regulon, and the regulatory role of RamA in S. marcescens. Importance We demonstrate that, in S. marcescens, the transcriptional regulator RamA is autoregulated and also controlled by the PhoP/PhoQ signal transduction system. We have shown that PhoP is able to directly and specifically bind to ramA, mgtE1, mgtE2, lpxO1 and lpxO2 promoter regions. In addition, RamA is able to directly interact with the promoter regions of ramA, lpxO1 but indirectly regulates mgtE1 and lpxO2. Finally, we found that, in S. marcescens, RamA overexpression induces the AcrAB-TolC efflux pump required to reduce susceptibility to tetracycline and nalidixic acid. Collectively, these results further our understanding of PhoP/PhoQ regulon in S. marcescens and demonstrate the involvement of RamA in the protection against antibiotic challenges.

Download Full-text