scholarly journals PhoQ Mutations Promote Lipid A Modification and Polymyxin Resistance of Pseudomonas aeruginosa Found in Colistin-Treated Cystic Fibrosis Patients

2011 ◽  
Vol 55 (12) ◽  
pp. 5761-5769 ◽  
Author(s):  
Amanda K. Miller ◽  
Mark K. Brannon ◽  
Laurel Stevens ◽  
Helle Krogh Johansen ◽  
Sara E. Selgrade ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Lipid A ◽  
Loss Of Function ◽  
Content Type ◽  
Clinical Strains ◽  
Regulatory Systems ◽  
Polymyxin E ◽  
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 polymyxin resistance (MICs of 8 to 64 mg/liter) in laboratory and clinical strains of this organism. To explore the role of PhoPQ in high-level clinical polymyxin resistance,P. aeruginosastrains with colistin MICs > 512 mg/liter that had been isolated from cystic fibrosis patients treated with inhaled colistin (polymyxin E) were analyzed. Probable loss-of-functionphoQalleles found in these cystic fibrosis strains conferred resistance to polymyxin. Partial and complete suppressor mutations inphoPwere identified in some cystic fibrosis strains with resistance-conferringphoQmutations, suggesting that additional loci can be involved in polymyxin resistance inP. aeruginosa. Disruption of chromosomalphoQin the presence of an intactphoPallele stimulated 4-amino-l-arabinose addition to lipid A and induced transcription from the promoter of thepmrH(arnB) operon, consistent with the known role of this lipid A modification in polymyxin resistance. These results indicate thatphoQloss-of-function mutations can contribute to high-level polymyxin resistance in clinical strains ofP. aeruginosa.

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

2011 ◽  
Vol 56 (2) ◽  
pp. 1019-1030 ◽  
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.

scholarly journals Polymyxin Resistance ofPseudomonas aeruginosa phoQMutants Is Dependent on Additional Two-Component Regulatory Systems

2013 ◽  
Vol 57 (5) ◽  
pp. 2204-2215 ◽  
Author(s):  
Alina D. Gutu ◽  
Nicole Sgambati ◽  
Pnina Strasbourger ◽  
Mark K. Brannon ◽  
Michael A. Jacobs ◽  
...  
Keyword(s):  
Lipid A ◽  
Regulatory System ◽  
Content Type ◽  
Regulatory Systems ◽  
Clinical Resistance ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems ◽  
High Level

ABSTRACTPseudomonas aeruginosacan develop resistance to polymyxin as a consequence of mutations in the PhoPQ regulatory system, mediated by covalent lipid A modification. Transposon mutagenesis of a polymyxin-resistantphoQmutant defined 41 novel loci required for resistance, including two regulatory systems, ColRS and CprRS. Deletion of thecolRSgenes, individually or in tandem, abrogated the polymyxin resistance of a ΔphoQmutant, as did individual or tandem deletion ofcprRS. Individual deletion ofcolRorcolSin a ΔphoQmutant also suppressed 4-amino-l-arabinose addition to lipid A, consistent with the known role of this modification in polymyxin resistance. Surprisingly, tandem deletion ofcolRSorcprRSin the ΔphoQmutant or individual deletion ofcprRorcprSfailed to suppress 4-amino-l-arabinose addition to lipid A, indicating that this modification alone is not sufficient for PhoPQ-mediated polymyxin resistance inP. aeruginosa. Episomal expression ofcolRSorcprRSin tandem or ofcprRindividually complemented the Pm resistance phenotype in the ΔphoQmutant, while episomal expression ofcolR,colS, orcprSindividually did not. Highly polymyxin-resistantphoQmutants ofP. aeruginosaisolated from polymyxin-treated cystic fibrosis patients harbored mutant alleles ofcolRSandcprS; when expressed in a ΔphoQbackground, these mutant alleles enhanced polymyxin resistance. These results define ColRS and CprRS as two-component systems regulating polymyxin resistance inP. aeruginosa, indicate that addition of 4-amino-l-arabinose to lipid A is not the only PhoPQ-regulated biochemical mechanism required for resistance, and demonstrate thatcolRSandcprSmutations can contribute to high-level clinical resistance.

scholarly journals Draft Genome Sequences of Four Pseudomonas aeruginosa Clinical Strains with Various Biofilm Phenotypes

2020 ◽  
Vol 9 (1) ◽  
Author(s):  
Amine M. Boukerb ◽  
Marjolaine Simon ◽  
Erwan Pernet ◽  
Albane Jouault ◽  
Emilie Portier ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Genome Size ◽  
Draft Genome ◽  
Genome Sequences ◽  
Content Type ◽  
Clinical Strains

Biofilms produced by Pseudomonas aeruginosa present a serious threat to cystic fibrosis patients. Here, we report the draft genome sequences of four cystic fibrosis isolates displaying various mucoid and biofilm phenotypes. The estimated average genome size was about 6,255,986 ± 50,202 bp with a mean G+C content of 66.52 ± 0.06%.

scholarly journals The Efflux Pump MexXY/OprM Contributes to the Tolerance and Acquired Resistance of Pseudomonas aeruginosa to Colistin

2020 ◽  
Vol 64 (4) ◽  
Author(s):  
Hélène Puja ◽  
Arnaud Bolard ◽  
Aurélie Noguès ◽  
Patrick Plésiat ◽  
Katy Jeannot
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Lipid A ◽  
Efflux Pump ◽  
Acquired Resistance ◽  
Intrinsic Resistance ◽  
Drug Induced ◽  
Functional Link ◽  
Content Type ◽  
Regulatory Systems ◽  
Early Drug

ABSTRACT The intrinsic resistance of Pseudomonas aeruginosa to polymyxins in part relies on the addition of 4-amino-4-deoxy-l-arabinose (Ara4N) molecules to the lipid A of lipopolysaccharide (LPS), through induction of operon arnBCADTEF-ugd (arn) expression. As demonstrated previously, at least three two-component regulatory systems (PmrAB, ParRS, and CprRS) are able to upregulate this operon when bacteria are exposed to colistin. In the present study, gene deletion experiments with the bioluminescent strain PAO1::lux showed that ParRS is a key element in the tolerance of P. aeruginosa to this last-resort antibiotic (i.e., resistance to early drug killing). Other loci of the ParR regulon, such as those encoding the efflux proteins MexXY (mexXY), the polyamine biosynthetic pathway PA4773-PA4774-PA4775, and Ara4N LPS modification process (arnBCADTEF-ugd), also contribute to the bacterial tolerance in an intricate way with ParRS. Furthermore, we found that both stable upregulation of the arn operon and drug-induced ParRS-dependent overexpression of the mexXY genes accounted for the elevated resistance of pmrB mutants to colistin. Deletion of the mexXY genes in a constitutively activated ParR mutant of PAO1 was associated with significantly increased expression of the genes arnA, PA4773, and pmrA in the absence of colistin exposure, thereby highlighting a functional link between the MexXY/OprM pump, the PA4773-PA4774-PA4775 pathway, and Ara4N-based modification of LPS. The role played by MexXY/OprM in the adaptation of P. aeruginosa to polymyxins opens new perspectives for restoring the susceptibility of resistant mutants through the use of efflux inhibitors.

scholarly journals Adaptive and Mutational Responses to Peptide Dendrimer Antimicrobials in Pseudomonas aeruginosa

2020 ◽  
Vol 64 (4) ◽  
Author(s):  
Fatma Ben Jeddou ◽  
Léna Falconnet ◽  
Alexandre Luscher ◽  
Thissa Siriwardena ◽  
Jean-Louis Reymond ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Lipid A ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Sensor Kinase ◽  
Loss Of Function ◽  
Adaptive Responses ◽  
Kinase Gene ◽  
Polyamine Biosynthesis ◽  
Content Type

ABSTRACT Colistin (polymyxin E) is a last-resort antibiotic against multidrug-resistant isolates of Pseudomonas aeruginosa. However, the nephro-toxicity of colistin limits its use, spurring the interest in novel antimicrobial peptides (AMP). Here, we show that the synthetic AMP-dendrimer G3KL (MW 4,531.38 Da, 15 positive charges, MIC = 8 mg/liter) showed faster killing than polymyxin B (Pmx-B) with no detectable resistance selection in P. aeruginosa strain PA14. Spontaneous mutants selected on Pmx-B, harboring loss of function mutations in the PhoQ sensor kinase gene, showed increased Pmx-B MICs and arnB operon expression (4-amino-l-arabinose addition to lipid A), but remained susceptible to dendrimers. Two mutants carrying a missense mutation in the periplasmic loop of the PmrB sensor kinase showed increased MICs for Pmx-B (8-fold) and G3KL (4-fold) but not for the dendrimer T7 (MW 4,885.64 Da, 16 positive charges, MIC = 8 mg/liter). The pmrB mutants showed increased expression of the arnB operon as well as of the speD2-speE2-PA4775 operon, located upstream of pmrAB, and involved in polyamine biosynthesis. Exogenous supplementation with the polyamines spermine and norspermine increased G3KL and T7 MICs in a phoQ mutant background but not in the PA14 wild type. This suggests that both addition of 4-amino-l-arabinose and secretion of polyamines are required to reduce susceptibility to dendrimers, probably neutralizing the negative charges present on the lipid A and the 2-keto-3-deoxyoctulosonic acid (KDO) sugars of the lipopolysaccharide (LPS), respectively. We further show by transcriptome analysis that the dendrimers G3KL and T7 induce adaptive responses through the CprRS two-component system in PA14.

scholarly journals Role of the MexEF-OprN Efflux System in Low-Level Resistance of Pseudomonas aeruginosa to Ciprofloxacin

2011 ◽  
Vol 55 (12) ◽  
pp. 5676-5684 ◽  
Author(s):  
Catherine Llanes ◽  
Thilo Köhler ◽  
Isabelle Patry ◽  
Barbara Dehecq ◽  
Christian van Delden ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Reference Strain ◽  
Efflux Pump ◽  
Resistance Mechanisms ◽  
Efflux System ◽  
Content Type ◽  
Clinical Strains ◽  
Major Mechanism ◽  
Level Resistance

ABSTRACTIn this study, we investigated the resistance mechanisms to fluoroquinolones of 85 non-cystic fibrosis strains ofPseudomonas aeruginosaexhibiting a reduced susceptibility to ciprofloxacin (MICs from 0.25 to 2 μg/ml). In addition to MexAB-OprM (31 of 85 isolates) and MexXY/OprM (39 of 85), the MexEF-OprN efflux pump (10 of 85) was found to be commonly upregulated in this population that is considered susceptible or of intermediate susceptibility to ciprofloxacin, according to current breakpoints. Analysis of the 10 MexEF-OprN overproducers (nfxCmutants) revealed the presence of various mutations in themexT(2 isolates),mexS(5 isolates), and/ormvaT(2 isolates) genes, the inactivation of which is known to increase the expression of themexEF-oprNoperon in reference strain PAO1-UW. However, these genes were intact in 3 of 10 of the clinical strains. Interestingly, ciprofloxacin at 2 μg/ml or 4 μg/ml preferentially selectednfxCmutants from wild-type clinical strains (n= 10 isolates) and from first-step mutants (n= 10) overexpressing Mex pumps, thus indicating that MexEF-OprN represents a major mechanism by whichP. aeruginosamay acquire higher resistance levels to fluoroquinolones. These data support the notion that thenfxCmutants may be more prevalent in the clinical setting than anticipated and strongly suggest the involvement of still unknown genes in the regulation of this efflux system.

scholarly journals Draft Genome Sequence of a Pseudomonas aeruginosa Sequence Type 3351 Strain Exhibiting High-Level Resistance to Polymyxins in a Pediatric Patient with Cystic Fibrosis in Mexico

2020 ◽  
Vol 9 (2) ◽  
Author(s):  
Roberto Rosales-Reyes ◽  
Fernanda Esposito ◽  
Bruna Fuga ◽  
Louise Cerdeira ◽  
Catalina Gayosso-Vázquez ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Pediatric Patient ◽  
Genome Sequence ◽  
Draft Genome ◽  
Sequence Type ◽  
Draft Genome Sequence ◽  
Content Type ◽  
High Level ◽  
Level Resistance

Here, we present the draft genome sequence of a Pseudomonas aeruginosa isolate (strain CF16053) belonging to a novel sequence type (ST), ST3351, isolated from a pediatric patient with cystic fibrosis (CF). CF16053 shows high-level resistance to polymyxins associated with mutations in the pmrB gene. Biofilm, pyoverdine, exotoxin A, and type III secretion system (T3SS) genes were identified.

scholarly journals Role of CFTR, Pseudomonas aeruginosa and Toll-like receptors in cystic fibrosis lung inflammation

2009 ◽  
Vol 37 (4) ◽  
pp. 863-867 ◽  
Author(s):  
Paul J. Buchanan ◽  
Robert K. Ernst ◽  
J. Stuart Elborn ◽  
Bettina Schock
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Lipid A ◽  
Treatment Strategies ◽  
Host Interaction ◽  
Genetic Modifications ◽  
Lipid A Structure ◽  
Antimicrobial Peptide Resistance ◽  
Cf Transmembrane Conductance Regulator

CF (cystic fibrosis) is a severe autosomal recessive disease most common in Northwest European populations. Underlying mutations in the CFTR (CF transmembrane conductance regulator) gene cause deregulation of ion transport and subsequent dehydration of the airway surface liquid, producing a viscous mucus layer on the airway surface of CF patients. This layer is readily colonized by bacteria such as Pseudomonas aeruginosa. Owing to the resulting environment and treatment strategies, the bacteria acquire genetic modifications such as antibiotic resistance, biofilm formation, antimicrobial peptide resistance and pro-inflammatory lipid A structures. Lipid A is a component of the lipopolysaccharide cell wall present on bacteria and is recognized by TLR4 (Toll-like receptor 4). Its detection elicits a pro-inflammatory response that is heightened over time due to the addition of fatty acids to the lipid A structure. Eradication of bacteria from the lungs of CF patients becomes increasingly difficult and eventually leads to mortality. In the present review, we describe the role of lipid A as a virulent factor of Ps. aeruginosa; however, it appears that further work is needed to investigate the role of CFTR in the innate immune response and in modifying the pathogen–host interaction.

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