Non-coding small (micro) RNAs of Pseudomonas aeruginosa isolated from clinical isolates from adult patients with cystic fibrosis

Abstract Objectives The cephalosporin nitric oxide (NO)-donor prodrug DEA-C3D (‘DiEthylAmin-Cephalosporin-3′-Diazeniumdiolate’) has been shown to initiate the dispersal of biofilms formed by the Pseudomonas aeruginosa laboratory strain PAO1. In this study, we investigated whether DEA-C3D disperses biofilms formed by clinical cystic fibrosis (CF) isolates of P. aeruginosa and its effect in combination with two antipseudomonal antibiotics, tobramycin and colistin, in vitro. Methods β-Lactamase-triggered release of NO from DEA-C3D was confirmed using a gas-phase chemiluminescence detector. MICs for P. aeruginosa clinical isolates were determined using the broth microdilution method. A crystal violet staining technique and confocal laser scanning microscopy were used to evaluate the effects of DEA-C3D on P. aeruginosa biofilms alone and in combination with tobramycin and colistin. Results DEA-C3D was confirmed to selectively release NO in response to contact with bacterial β-lactamase. Despite lacking direct, cephalosporin/β-lactam-based antibacterial activity, DEA-C3D was able to disperse biofilms formed by three P. aeruginosa clinical isolates. Confocal microscopy revealed that DEA-C3D in combination with tobramycin produces similar reductions in biofilm to DEA-C3D alone, whereas the combination with colistin causes near complete eradication of P. aeruginosa biofilms in vitro. Conclusions DEA-C3D is effective in dispersing biofilms formed by multiple clinical isolates of P. aeruginosa and could hold promise as a new adjunctive therapy to patients with CF.

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Persistency of Pseudomonas aeruginosa in sputum cultures and clinical outcomes in adult patients with cystic fibrosis

European Journal of Clinical Microbiology & Infectious Diseases ◽

10.1007/s10096-011-1483-8 ◽

2011 ◽

Vol 31 (7) ◽

pp. 1603-1610 ◽

Cited By ~ 12

Author(s):

A. Burkett ◽

K. L. Vandemheen ◽

T. Giesbrecht-Lewis ◽

K. Ramotar ◽

W. Ferris ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Clinical Outcomes ◽

Adult Patients

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Efficacy and Safety of Liposomal Clarithromycin and Its Effect on Pseudomonas aeruginosa Virulence Factors

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00235-13 ◽

2013 ◽

Vol 57 (6) ◽

pp. 2694-2704 ◽

Cited By ~ 61

Author(s):

Mai Alhajlan ◽

Moayad Alhariri ◽

Abdelwahab Omri

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Virulence Factors ◽

Biological Fluid ◽

Free Drug ◽

Clinical Isolates ◽

Epithelial Cell Line ◽

Efficacy And Safety ◽

Surface Charges ◽

Content Type

ABSTRACTWe investigated the efficacy and safety of liposomal clarithromycin formulations with different surface charges against clinical isolates ofPseudomonas aeruginosafrom the lungs of cystic fibrosis (CF) patients. The liposomal clarithromycin formulations were prepared by the dehydration-rehydration method, and their sizes were measured using the dynamic-light-scattering technique. Encapsulation efficiency was determined by microbiological assay, and the stabilities of the formulations in biological fluid were evaluated for a period of 48 h. The MICs and minimum bactericidal concentrations (MBCs) of free and liposomal formulations were determined withP. aeruginosastrains isolated from CF patients. Liposomal clarithromycin activity against biofilm-formingP. aeruginosawas compared to that of free antibiotic using the Calgary Biofilm Device (CBD). The effects of subinhibitory concentrations of free and liposomal clarithromycin on bacterial virulence factors and motility on agar were investigated on clinical isolates ofP. aeruginosa. The cytotoxicities of the liposome preparations and free drug were evaluated on a pulmonary epithelial cell line (A549). The average diameter of the formulations was >222 nm, with encapsulation efficiencies ranging from 5.7% to 30.4%. The liposomes retained more than 70% of their drug content during the 48-h time period. The highly resistant strains ofP. aeruginosabecame susceptible to liposome-encapsulated clarithromycin (MIC, 256 mg/liter versus 8 mg/liter;P< 0.001). Liposomal clarithromycin reduced the bacterial growth within the biofilm by 3 to 4 log units (P< 0.001), significantly attenuated virulence factor production, and reduced bacterial twitching, swarming, and swimming motilities. The clarithromycin-entrapped liposomes were less cytotoxic than the free drug (P< 0.001). These data indicate that our novel formulations could be a useful strategy to enhance the efficacy of clarithromycin against resistantP. aeruginosastrains that commonly affect individuals with cystic fibrosis.

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Pseudomonas aeruginosa cystic fibrosis clinical isolates produce exotoxin A with altered ADP-ribosyltransferase activity and cytotoxicity The GenBank accession numbers for the toxA sequences are: strain 4384, AF227419; strain 5154, AF227420; strain 5166, AF227421; strain 5552, AF227422; strain 5585, AF227423; strain 5588, AF227424.

Microbiology ◽

10.1099/00221287-146-8-1891 ◽

2000 ◽

Vol 146 (8) ◽

pp. 1891-1899 ◽

Cited By ~ 17

Author(s):

Claude V. Gallant ◽

Tracy L. Raivio ◽

Joan C. Olson ◽

Donald E. Woods ◽

Douglas G. Storey

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Clinical Isolates ◽

Exotoxin A ◽

Adp Ribosyltransferase

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Sera from Adult Patients with Cystic Fibrosis Contain Antibodies to Pseudomonas aeruginosa Type III Apparatus

Infection and Immunity ◽

10.1128/iai.69.2.1185-1188.2001 ◽

2001 ◽

Vol 69 (2) ◽

pp. 1185-1188 ◽

Cited By ~ 32

Author(s):

Joel Moss ◽

Mary E. Ehrmantraut ◽

Bruce D. Banwart ◽

Dara W. Frank ◽

Joseph T. Barbieri

Keyword(s):

Cystic Fibrosis ◽

Immune Response ◽

Pseudomonas Aeruginosa ◽

Adult Patients ◽

Eukaryotic Cells ◽

Type Iii

ABSTRACT Expression of type III proteins of Pseudomonas aeruginosa in patients with cystic fibrosis (CF) was investigated by measuring the immune response against components of the type III pathway. Twenty-three of the 33 sera contained antibodies against PcrV, a protein involved in translocation of type III cytotoxins into eukaryotic cells, and 11 of 33 had antibodies against ExoS, while most CF sera contained antibodies against PopB and PopD, components of the type III apparatus. These data indicate that P. aeruginosacommonly expresses components of the type III translocation apparatus in adult CF patients.

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PmrB Mutations Promote Polymyxin Resistance of Pseudomonas aeruginosa Isolated from Colistin-Treated Cystic Fibrosis Patients

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05829-11 ◽

2011 ◽

Vol 56 (2) ◽

pp. 1019-1030 ◽

Cited By ~ 106

Author(s):

Samuel M. Moskowitz ◽

Mark K. Brannon ◽

Nandini Dasgupta ◽

Miyuki Pier ◽

Nicole Sgambati ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Lipid A ◽

Clinical Isolates ◽

Gain Of Function ◽

Content Type ◽

Regulatory Systems ◽

Repeated Passage ◽

High Level

ABSTRACTPseudomonas aeruginosacan develop resistance to polymyxin and other cationic antimicrobial peptides. Previous work has shown that mutations in the PmrAB and PhoPQ regulatory systems can confer low to moderate levels of colistin (polymyxin E) resistance in laboratory strains and clinical isolates of this organism (MICs of 8 to 64 mg/liter). To explore the role of PmrAB in high-level clinical polymyxin resistance,P. aeruginosaisolates from chronically colistin-treated cystic fibrosis patients, most with colistin MICs of >512 mg/liter, were analyzed. These cystic fibrosis isolates contained probable gain-of-functionpmrBalleles that conferred polymyxin resistance to strains with a wild-type orpmrABdeletion background. Double mutantpmrBalleles that contained mutations in both the periplasmic and dimerization-phosphotransferase domains markedly augmented polymyxin resistance. Expression of mutantpmrBalleles induced transcription from the promoter of thearnBoperon and stimulated addition of 4-amino-l-arabinose to lipid A, consistent with the known role of this lipid A modification in polymyxin resistance. For some highly polymyxin-resistant clinical isolates, repeated passage without antibiotic selection pressure resulted in loss of resistance, suggesting that secondary suppressors occur at a relatively high frequency and account for the instability of this phenotype. These results indicate thatpmrBgain-of-function mutations can contribute to high-level polymyxin resistance in clinical strains ofP. aeruginosa.

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