Aminoglycoside resistance of<i>Pseudomonas aeruginosa</i>in cystic fibrosis results from convergent evolution in the<i>mexZ</i>gene

Prickett, Michelle; Hauser, Alan R.; McColley, Susanna A.; Cullina, Joanne; Potter, Eileen; Powers, Catherine A.; Jain, Manu

doi:10.1136/thoraxjnl-2015-208027

Cited by 53 publications

(46 citation statements)

References 34 publications

(23 reference statements)

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“…There was high variation in the expression of both ampC (over 20,000-fold) and mexX (2,000-fold) among isolates of P. aeruginosa grown on laboratory agar. Other researchers have also reported extensive variation in the expression of these genes among isolates of P. aeruginosa from a number of clinical settings (25,27,30,(39)(40)(41), reflecting the frequent occurrence of mutations that result in upregulation of ampC and mexX in clinical isolates of P. aeruginosa (22)(23)(24)(25)(26)(27)(28)(29)(30)(31). Meropenem increased the expression of ampC, and tobramycin increased expression of mexX in the isolates in our study.…”

Section: Discussionsupporting

confidence: 71%

“…␤-Lactam antibiotics increase expression of the ampC gene, and aminoglycoside substrates increase expression of the mexXY genes, which are expressed as an operon (21). P. aeruginosa isolates cultured from individuals with CF commonly possess genetic mutations that give rise to increased ampC and mexXY expression and are associated with increased ␤-lactam and aminoglycoside antibiotic resistance (22)(23)(24)(25)(26)(27)(28)(29)(30)(31).…”

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confidence: 99%

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Expression of Pseudomonas aeruginosa Antibiotic Resistance Genes Varies Greatly during Infections in Cystic Fibrosis Patients

Martin

Robson

Watts

et al. 2018

Antimicrob Agents Chemother

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The lungs of individuals with cystic fibrosis (CF) become chronically infected with that is difficult to eradicate by antibiotic treatment. Two key antibiotic resistance mechanisms are the AmpC β-lactamase that degrades β-lactam antibiotics and MexXYOprM, a three-protein efflux pump that expels aminoglycoside antibiotics from the bacterial cells. Levels of antibiotic resistance gene expression are likely to be a key factor in antibiotic resistance but have not been determined during infection. The aims of this research were to investigate the expression of the and genes during infection in patients with CF and in bacteria isolated from the same patients and grown under laboratory conditions. isolates from 36 CF patients were grown in laboratory culture and gene expression measured by reverse transcription-quantitative PCR (RT-qPCR). The expression of varied over 20,000-fold and that of over 2,000-fold between isolates. The median expression levels of both genes were increased by the presence of subinhibitory concentrations of antibiotics. To measure gene expression during infection, we carried out RT-qPCR using RNA extracted from fresh sputum samples obtained from 31 patients. The expression of varied over 4,000-fold, while expression varied over 100-fold, between patients. Despite these wide variations, median levels of expression of in bacteria in sputum were similar to those in laboratory-grown bacteria. The expression of was higher in sputum than in laboratory-grown bacteria. Overall, our data demonstrate that genes that contribute to antibiotic resistance can be highly expressed in patients, but there is extensive isolate-to-isolate and patient-to-patient variation.

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Section: Discussionsupporting

confidence: 71%

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confidence: 99%

Expression of Pseudomonas aeruginosa Antibiotic Resistance Genes Varies Greatly during Infections in Cystic Fibrosis Patients

Martin

Robson

Watts

et al. 2018

Antimicrob Agents Chemother

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“…MIC determinations showed that all isolates, except for 1991 and 1995, became resistant to tobramycin. The main origin of high-level resistance to aminoglycosides is the overexpression of MexXY-OprM efflux system (50), which is primarily caused by mexZ mutations (10, 21, 51). In addition, mutations in the amgRS and parRS two-component systems genes have also been involved in the regulation of MexXY expression (52).…”

Section: Resultsmentioning

confidence: 99%

HypermutatorPseudomonas aeruginosaexploits multiple genetic pathways to develop multidrug resistance during long-term infections in the airways of cystic fibrosis patients

Colque

Orio

Feliziani

et al. 2019

Preprint

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21 cystic fibrosis 22 WORD COUNT ABSTRACT: Abstract, 249 words. Importance, 147 words 23 WORD COUNT TEXT: 4995 words.24 2 25 ABSTRACT 26 Pseudomonas aeruginosa exploits intrinsic and acquired resistance mechanisms to resist almost 27 every antibiotic used in chemotherapy. Antimicrobial resistance in P. aeruginosa isolated from 28 cystic fibrosis (CF) patients is further enhanced by the occurrence of hypermutator strains, a 29 hallmark of chronic CF infections. However, the within-patient genetic diversity of P. aeruginosa 30 populations related to antibiotic resistance remains unexplored. Here, we show the evolution of 31 the mutational resistome profile of a P. aeruginosa hypermutator lineage by performing 32 longitudinal and transversal analyses of isolates collected from a CF patient throughout 20 years 33 of chronic infection. Our results show the accumulation of thousands of mutations with an overall 34 evolutionary history characterized by purifying selection. However, mutations in antibiotic 35resistance genes appear to be positively selected, driven by antibiotic treatment. Antibiotic 36 resistance increased as infection progressed towards the establishment of a population constituted 37 by genotypically diversified coexisting sub-lineages, all of which converged to multi-drug 38 resistance. These sub-lineages emerged by parallel evolution through distinct evolutionary 39 pathways, which affected genes of the same functional categories. Interestingly, ampC and fstI, 40 encoding the β-lactamase and penicillin-binding protein 3, respectively, were found among the 41 most frequently mutated genes. In fact, both genes were targeted by multiple independent 42 mutational events, which led to a wide diversity of coexisting alleles underlying β-lactam 43 resistance. Our findings indicate that hypermutators, apart from boosting antibiotic resistance 44 evolution by simultaneously targeting several genes, favor the emergence of adaptive innovative 45 alleles by clustering beneficial/compensatory mutations in the same gene, hence expanding P. 46 aeruginosa strategies for persistence. 47 IMPORTANCE 483 By increasing mutation rates, hypermutators boost antibiotic resistance evolution by enabling 49 bacterial pathogens to fully exploit their genetic potential and achieve resistance mechanisms for 50 almost every known antimicrobial agent. Here, we show how co-existing clones from a P. 51 aeruginosa hypermutator lineage that evolved during 20 years of chronic infection and antibiotic 52 chemotherapy, converged to multidrug resistance by targeting genes from alternative genetic 53 pathways that are part of the broad P. aeruginosa resistome. Within this complex assembly of 54 combinatorial genetic changes, in some specific cases, multiple mutations are needed in the same 55 gene to reach a fine tuned resistance phenotype. Hypermutability enables this genetic edition 56 towards higher resistance profiles by recurrently targeting these genes, thus promoting new 57 epistatic relationships and the emergence of innovativ...

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“…Additional pumps known to impact susceptibility to antibiotics are MexXY (OprM), which is inducible by antibiotics that inhibit protein synthesis (13)(14)(15)(16), and MexCD-OprJ and MexEF-OprN, which are typically silent but which can be strongly upregulated in so-called nfxB (17)(18)(19) and nfxC (20)(21)(22) mutants, respectively. Regulatory mutations (e.g., in the repressor gene mexZ) or mutations that interfere with protein synthesis can also lead to the constitutive expression of MexXY and corresponding reductions in susceptibility to antibiotics (13,16,(23)(24)(25)(26)(27)(28). Overexpression of these and other pumps has been found in clinical isolates, presumably selected by therapeutic exposure to antibiotics (2).…”

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confidence: 99%

Target (MexB)- and Efflux-Based Mechanisms Decreasing the Effectiveness of the Efflux Pump Inhibitor D13-9001 in Pseudomonas aeruginosa PAO1: Uncovering a New Role for MexMN-OprM in Efflux of β-Lactams and a Novel Regulatory Circuit (MmnRS) Controlling MexMN Expression

Ranjitkar

Jones

Mostafavi

et al. 2019

Antimicrob Agents Chemother

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Efflux pumps contribute to antibiotic resistance in Gram-negative pathogens. Correspondingly, efflux pump inhibitors (EPIs) may reverse this resistance. D13-9001 specifically inhibits MexAB-OprM in Pseudomonas aeruginosa. Mutants with decreased susceptibility to MexAB-OprM inhibition by D13-9001 were identified, and these fell into two categories: those with alterations in the target MexB (F628L and ΔV177) and those with an alteration in a putative sensor kinase of unknown function, PA1438 (L172P). The alterations in MexB were consistent with reported structural studies of the D13-9001 interaction with MexB. The PA1438L172P alteration mediated a >150-fold upregulation of MexMN pump gene expression and a >50-fold upregulation of PA1438 and the neighboring response regulator gene, PA1437. We propose that these be renamed mmnR and mmnS for MexMN regulator and MexMN sensor, respectively. MexMN was shown to partner with the outer membrane channel protein OprM and to pump several β-lactams, monobactams, and tazobactam. Upregulated MexMN functionally replaced MexAB-OprM to efflux these compounds but was insusceptible to inhibition by D13-9001. MmnSL172P also mediated a decrease in susceptibility to imipenem and biapenem that was independent of MexMN-OprM. Expression of oprD, encoding the uptake channel for these compounds, was downregulated, suggesting that this channel is also part of the MmnSR regulon. Transcriptome sequencing (RNA-seq) of cells encoding MmnSL172P revealed, among other things, an interrelationship between the regulation of mexMN and genes involved in heavy metal resistance.

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Aminoglycoside resistance ofPseudomonas aeruginosain cystic fibrosis results from convergent evolution in themexZgene

Cited by 53 publications

References 34 publications

Expression of Pseudomonas aeruginosa Antibiotic Resistance Genes Varies Greatly during Infections in Cystic Fibrosis Patients

Expression of Pseudomonas aeruginosa Antibiotic Resistance Genes Varies Greatly during Infections in Cystic Fibrosis Patients

HypermutatorPseudomonas aeruginosaexploits multiple genetic pathways to develop multidrug resistance during long-term infections in the airways of cystic fibrosis patients

Target (MexB)- and Efflux-Based Mechanisms Decreasing the Effectiveness of the Efflux Pump Inhibitor D13-9001 in Pseudomonas aeruginosa PAO1: Uncovering a New Role for MexMN-OprM in Efflux of β-Lactams and a Novel Regulatory Circuit (MmnRS) Controlling MexMN Expression

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