scholarly journals Changes in toxin production of environmental Pseudomonas aeruginosa isolates exposed to sub-inhibitory concentrations of three common antibiotics

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0248014
Author(s):  
Biljana Mojsoska ◽  
Melanie Ghoul ◽  
Gabriel G. Perron ◽  
Håvard Jenssen ◽  
Fatima AlZahra’a Alatraktchi
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Toxin Production ◽  
Biologically Relevant ◽  
A Value ◽  
Severe Infections ◽  
Bacterial Virulence Factor ◽  
And Diffusion

Pseudomonas aeruginosa is an environmental pathogen that can cause severe infections in immunocompromised patients. P. aeruginosa infections are typically treated with multiple antibiotics including tobramycin, ciprofloxacin, and meropenem. However, antibiotics do not always entirely clear the bacteria from the infection site, where they may remain virulent. This is because the effective antibiotic concentration and diffusion in vitro may differ from the in vivo environment in patients. Therefore, it is important to understand the effect of non-lethal sub-inhibitory antibiotic concentrations on bacterial phenotype. Here, we investigate if sub-inhibitory antimicrobial concentrations cause alterations in bacterial virulence factor production using pyocyanin as a model toxin. We tested this using the aforementioned antibiotics on 10 environmental P. aeruginosa strains. Using on-the-spot electrochemical screening, we were able to directly quantify changes in production of pyocyanin in a measurement time of 17 seconds. Upon selecting 3 representative strains to further test the effects of sub-minimum inhibitory concentration (MICs), we found that pyocyanin production changed significantly when the bacteria were exposed to 10-fold MIC of the 3 antibiotics tested, and this was strain specific. A series of biologically relevant measured pyocyanin concentrations were also used to assess the effects of increased virulence on a culture of epithelial cells. We found a decreased viability of the epithelial cells when incubated with biologically relevant pyocyanin concentrations. This suggests that the antibiotic-induced virulence also is a value worth being enclosed in regular testing of pathogens.

scholarly journals Human Tear Fluid Protects against Pseudomonas aeruginosa Keratitis in a Murine Experimental Model

2007 ◽  
Vol 75 (5) ◽  
pp. 2325-2332 ◽  
Author(s):  
Mary S. F. Kwong ◽  
David J. Evans ◽  
Minjian Ni ◽  
Brigitte A. Cowell ◽  
Suzanne M. J. Fleiszig
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Bacterial Colonization ◽  
Healing Process ◽  
Tear Fluid ◽  
Protective Mechanism ◽  
Healthy Human ◽  
Epithelial Barrier Function

ABSTRACT Pseudomonas aeruginosa keratitis is an acute sight-threatening infection. We previously reported that human tear fluid could protect individual human corneal epithelial cells in vitro against invasion by and cytotoxicity due to clinical and laboratory isolates of P. aeruginosa and that the protective mechanism was independent of bacteriostatic activity. In the present study, we examined the effects of human tear fluid in vivo. Tears were collected from healthy human volunteers and were studied in vivo in mice. The effects on the virulence of both invasive and cytotoxic clinical isolates of P. aeruginosa were examined. Tear fluid was found to reduce the severity of disease when corneas were challenged with cytotoxic bacteria immediately after scratch injury, and it completely protected against susceptibility to infection by a cytotoxic strain in a model in which corneas were infected during the healing process 6 h after scratching. Visible protection correlated with the inhibition of bacterial colonization 1, 4, and 48 h postinoculation. Tear fluid also significantly reduced the severity of infections caused by invasive P. aeruginosa in the 6-h-healing model. This result also coincided with significantly reduced bacterial colonization at 48 h. In vitro, human tear fluid significantly reduced the ability of invasive and cytotoxic bacteria to translocate across corneal epithelia and increased transepithelial resistance with or without bacterial inoculation. These data show that human tear fluid can protect against P. aeruginosa corneal infection in vivo and that the mechanism likely involves enhanced epithelial barrier function in addition to protection of individual epithelial cells against bacterial internalization and cytotoxicity.

scholarly journals Pseudomonas aeruginosa ExoT Acts In Vivo as a GTPase-Activating Protein for RhoA, Rac1, and Cdc42

2002 ◽  
Vol 70 (4) ◽  
pp. 2198-2205 ◽  
Author(s):  
B. I. Kazmierczak ◽  
J. N. Engel
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Morphological Changes ◽  
Stress Fiber ◽  
Secreted Proteins ◽  
In Vitro Activity ◽  
Gtpase Activating Protein ◽  
Infected Cells

ABSTRACT The Pseudomonas aeruginosa protein ExoT is a bacterial GTPase-activating protein (GAP) that has in vitro activity toward Rho, Rac, and Cdc42 GTPases. Expression of ExoT both inhibits the internalization of strain PA103 by macrophages and epithelial cells and is associated with morphological changes (cell rounding and detachment) of infected cells. We find that expression of ExoT leads to the loss of GTP-bound RhoA, Rac1, and Cdc42 in transfected HeLa cells, demonstrating that ExoT has GAP activity in vivo toward all three GTPases. GAP activity is absolutely dependent on the presence of arginine at position 149 but is not affected by whether ExoT is expressed in the absence or presence of other P. aeruginosa type III secreted proteins. We also demonstrate that expression of ExoT in epithelial cells is sufficient to cause stress fiber disassembly by means of ExoT's GAP activity toward RhoA.

Dimethyl sulfoxide decreases interleukin-8-mediated neutrophil recruitment in the airways

1996 ◽  
Vol 271 (5) ◽  
pp. L838-L843 ◽  
Author(s):  
P. P. Massion ◽  
A. Linden ◽  
H. Inoue ◽  
M. Mathy ◽  
K. M. Grattan ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Dimethyl Sulfoxide ◽  
Neutrophil Recruitment ◽  
Interleukin 8 ◽  
Bronchial Epithelial ◽  
Anti Inflammatory

In this study, we investigated the role of dimethyl sulfoxide (DMSO) in inhibiting interleukin-8 (IL-8)-mediated neutrophil recruitment induced by Pseudomonas aeruginosa (PA) bacterial supernatant. First, we tested whether DMSO could inhibit IL-8 production induced by PA in human bronchial epithelial (16-HBE) cells in vitro. In these cells, exposure to PA or H2O2 induced IL-8 production dose dependently, an effect that was inhibited by 1% DMSO at both the protein and RNA level. Second, we tested whether DMSO could block the recruitment of neutrophils induced by PA in a bypassed segment of dog trachea in vivo. PA supernatant was placed in the tracheal segment for 6 h in four dogs, and neutrophil recruitment and IL-8 concentrations were measured in the superfusate. DMSO prevented the recruitment of neutrophils and IL-8 production induced by PA time dependently. The results suggest that DMSO may play an anti-inflammatory role in the airway by inhibiting IL-8 production in epithelial cells.

scholarly journals Effect of Hypoxia on the Pathogenesis of Acinetobacter baumannii and Pseudomonas aeruginosa In Vitro and in Murine Experimental Models of Infection

2018 ◽  
Vol 86 (10) ◽  
Author(s):  
María Luisa Gil-Marqués ◽  
María Eugenía Pachón-Ibáñez ◽  
Jerónimo Pachón ◽  
Younes Smani
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Acinetobacter Baumannii ◽  
Experimental Models ◽  
Host Cells ◽  
Content Type ◽  
Bactericidal Activities ◽  
Bacterial Concentrations

ABSTRACT Hypoxia modulates bacterial virulence and the inflammation response through hypoxia-inducible factor 1α (HIF-1α). Here we study the influence of hypoxia on Acinetobacter baumannii and Pseudomonas aeruginosa infections. In vitro, hypoxia increases the bactericidal activities of epithelial cells against A. baumannii and P. aeruginosa, reducing extracellular bacterial concentrations to 50.5% ± 7.5% and 90.8% ± 13.9%, respectively, at 2 h postinfection. The same phenomenon occurs in macrophages (67.6% ± 18.2% for A. baumannii at 2 h and 50.3% ± 10.9% for P. aeruginosa at 24 h). Hypoxia decreases the adherence of A. baumannii to epithelial cells (42.87% ± 8.16% at 2 h) and macrophages (52.0% ± 18.7% at 24 h), as well as that of P. aeruginosa (24.9% ± 4.5% in epithelial cells and 65.7% ± 5.5% in macrophages at 2 h). Moreover, hypoxia decreases the invasion of epithelial cells (48.6% ± 3.8%) and macrophages (8.7% ± 6.9%) by A. baumannii at 24 h postinfection and by P. aeruginosa at 2 h postinfection (75.0% ± 16.3% and 63.4% ± 5.4%, respectively). In vivo, hypoxia diminishes bacterial loads in fluids and tissues in animal models of infection by both pathogens. In contrast, mouse survival time was shorter under hypoxia (23.92 versus 36.42 h) with A. baumannii infection. No differences in the production of cytokines or HIF-1α were found between hypoxia and normoxia in vitro or in vivo. We conclude that hypoxia increases the bactericidal activities of host cells against both pathogens and reduces the interaction of pathogens with host cells. Moreover, hypoxia accelerates the rate at which animals die despite the lower bacterial concentrations in vivo.

Three-dimensional Growth of Renal Epithelial Cells in Vitro: A Tool in Toxicity Testing

1993 ◽  
Vol 21 (2) ◽  
pp. 191-195 ◽  
Author(s):  
Knut-Jan Andersen ◽  
Erik Ilsø Christensen ◽  
Hogne Vik
Keyword(s):  
Epithelial Cells ◽  
Three Dimensional ◽  
Toxicity Testing ◽  
Renal Tissue ◽  
Epithelial Cell Line ◽  
Multicellular Spheroids ◽  
Renal Epithelial Cell ◽  
Renal Epithelial Cells

The tissue culture of multicellular spheroids from the renal epithelial cell line LLC-PK1 (proximal tubule) is described. This represents a biological system of intermediate complexity between renal tissue in vivo and simple monolayer cultures. The multicellular structures, which show many similarities to kidney tubules in vivo, including a vectorial water transport, should prove useful for studying the potential nephrotoxicity of drugs and chemicals in vitro. In addition, the propagation of renal epithelial cells as multicellular spheroids in serum-free culture may provide information on the release of specific biological parameters, which may be suppressed or masked in serum-supplemented media.

scholarly journals Characterization of the SARS-CoV-2 coronavirus X4-like accessory protein

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Olanrewaju Ayodeji Durojaye ◽  
Nkwachukwu Oziamara Okoro ◽  
Arome Solomon Odiba
Keyword(s):  
Rapid Development ◽  
Protein A ◽  
The Novel ◽  
Quality Model ◽  
A Value ◽  
Model Protein ◽  
Novel Coronavirus ◽  
Global Threat

Abstract Background The novel coronavirus SARS-CoV-2 is currently a global threat to health and economies. Therapeutics and vaccines are in rapid development; however, none of these therapeutics are considered as absolute cure, and the potential to mutate makes it necessary to find therapeutics that target a highly conserved regions of the viral structure. Results In this study, we characterized an essential but poorly understood coronavirus accessory X4 protein, a core and stable component of the SARS-CoV family. Sequence analysis shows a conserved ~ 90% identity between the SARS-CoV-2 and previously characterized X4 protein in the database. QMEAN Z score of the model protein shows a value of around 0.5, within the acceptable range 0–1. A MolProbity score of 2.96 was obtained for the model protein and indicates a good quality model. The model has Ramachandran values of φ = − 57o and ψ = − 47o for α-helices and values of φ = − 130o and ψ = + 140o for twisted sheets. Conclusions The protein data obtained from this study provides robust information for further in vitro and in vivo experiment, targeted at devising therapeutics against the virus. Phylogenetic analysis further supports previous evidence that the SARS-CoV-2 is positioned with the SL-CoVZC45, BtRs-BetaCoV/YN2018B and the RS4231 Bat SARS-like corona viruses.

scholarly journals A Novel Infection Protocol in Zebrafish Embryo to Assess Pseudomonas aeruginosa Virulence and Validate Efficacy of a Quorum Sensing Inhibitor In Vivo

Pathogens ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 401
Author(s):  
Pauline Nogaret ◽  
Fatima El El Garah ◽  
Anne-Béatrice Blanc-Potard
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Animal Model ◽  
Quorum Sensing ◽  
Drug Testing ◽  
Drug Efficacy ◽  
Embryo Viability ◽  
Immersion Method ◽  
Opportunistic Human Pathogen

The opportunistic human pathogen Pseudomonas aeruginosa is responsible for a variety of acute infections and is a major cause of mortality in chronically infected cystic fibrosis patients. Due to increased resistance to antibiotics, new therapeutic strategies against P. aeruginosa are urgently needed. In this context, we aimed to develop a simple vertebrate animal model to rapidly assess in vivo drug efficacy against P. aeruginosa. Zebrafish are increasingly considered for modeling human infections caused by bacterial pathogens, which are commonly microinjected in embryos. In the present study, we established a novel protocol for zebrafish infection by P. aeruginosa based on bath immersion in 96-well plates of tail-injured embryos. The immersion method, followed by a 48-hour survey of embryo viability, was first validated to assess the virulence of P. aeruginosa wild-type PAO1 and a known attenuated mutant. We then validated its relevance for antipseudomonal drug testing by first using a clinically used antibiotic, ciprofloxacin. Secondly, we used a novel quorum sensing (QS) inhibitory molecule, N-(2-pyrimidyl)butanamide (C11), the activity of which had been validated in vitro but not previously tested in any animal model. A significant protective effect of C11 was observed on infected embryos, supporting the ability of C11 to attenuate in vivo P. aeruginosa pathogenicity. In conclusion, we present here a new and reliable method to compare the virulence of P. aeruginosa strains in vivo and to rapidly assess the efficacy of clinically relevant drugs against P. aeruginosa, including new antivirulence compounds.

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