Mutational and acquired carbapenem resistance mechanisms in multidrug resistant Pseudomonas aeruginosa clinical isolates from Recife, Brazil

A total of 203 clinical isolates of Pseudomonas aeruginosa was collected during 2001–2006 from five university hospitals in Sofia, Bulgaria, to assess the current levels of antimicrobial susceptibility and to evaluate resistance mechanisms to antipseudomonal antimicrobial agents. The antibiotic resistance rates against the following antimicrobials were: carbenicillin 93.1 %, azlocillin 91.6 %, piperacillin 86.2 %, piperacillin/tazobactam 56.8 %, ceftazidime 45.8 %, cefepime 48.9 %, cefpirome 58.2 %, aztreonam 49.8 %, imipenem 42.3 %, meropenem 45.5 %, amikacin 59.1 %, gentamicin 79.7 %, tobramycin 89.6 %, netilmicin 69.6 % and ciprofloxacin 80.3 %. A total of 101 of the studied P. aeruginosa isolates (49.8 %) were multidrug resistant. Structural genes encoding class A and class D β-lactamases showed the following frequencies: bla VEB-1 33.1 %, bla PSE-1 22.5 %, bla PER-1 0 %, bla OXA-groupI 41.3 % and bla OXA-groupII 8.8 %. IMP- and VIM-type carbapenemases were not detected. In conclusion, the studied clinical strains of P. aeruginosa were problematic nosocomial pathogens. VEB-1 extended-spectrum β-lactamases appear to have a significant presence among clinical P. aeruginosa isolates from Sofia. Carbapenem resistance was related to non-enzymic mechanisms such as a deficiency of OprD proteins and active efflux.

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Distribution of carbapenem resistance mechanisms in clinical isolates of XDR Pseudomonas aeruginosa

European Journal of Clinical Microbiology & Infectious Diseases ◽

10.1007/s10096-019-03585-0 ◽

2019 ◽

Vol 38 (8) ◽

pp. 1547-1552 ◽

Cited By ~ 6

Author(s):

Annalisa De Rosa ◽

Nico T. Mutters ◽

Claudio M. Mastroianni ◽

Stefan J. Kaiser ◽

Frank Günther

Keyword(s):

Pseudomonas Aeruginosa ◽

Carbapenem Resistance ◽

Resistance Mechanisms ◽

Clinical Isolates

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P867 Carbapenem resistance mechanisms in Norwegian clinical isolates of Pseudomonas aeruginosa

International Journal of Antimicrobial Agents ◽

10.1016/s0924-8579(07)70708-2 ◽

2007 ◽

Vol 29 ◽

pp. S223-S224

Author(s):

Ø. Samuelsen ◽

L. Buarø ◽

B. Aasnæs ◽

C.G. Giske ◽

B. Haldorsen ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Carbapenem Resistance ◽

Resistance Mechanisms ◽

Clinical Isolates

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Pathogenicity Genomic Island-Associated CrpP-Like Fluoroquinolone-Modifying Enzymes among Pseudomonas aeruginosa Clinical Isolates in Europe

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00489-20 ◽

2020 ◽

Vol 64 (7) ◽

Cited By ~ 1

Author(s):

José Manuel Ortiz de la Rosa ◽

Patrice Nordmann ◽

Laurent Poirel

Keyword(s):

Pseudomonas Aeruginosa ◽

Genomic Island ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Genomic Islands ◽

Clinical Isolates ◽

Gram Negative Bacteria ◽

Gram Negative ◽

Content Type ◽

Genes Encoding

ABSTRACT Many transferable quinolone resistance mechanisms have been identified in Gram-negative bacteria. The plasmid-encoded 65-amino-acid-long ciprofloxacin-modifying enzyme CrpP was recently identified in Pseudomonas aeruginosa isolates. We analyzed a collection of 100 clonally unrelated and multidrug-resistant P. aeruginosa clinical isolates, among which 46 were positive for crpP-like genes, encoding five CrpP variants conferring variable levels of reduced susceptibility to fluoroquinolones. These crpP-like genes were chromosomally located as part of pathogenicity genomic islands.

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Carbapenem Resistance Mechanisms in Pseudomonas aeruginosa Clinical Isolates

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.2.480-484.2001 ◽

2001 ◽

Vol 45 (2) ◽

pp. 480-484 ◽

Cited By ~ 123

Author(s):

Hyunjoo Pai ◽

Jong-Won Kim ◽

Jungmin Kim ◽

Ji Hyang Lee ◽

Kang Won Choe ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Amino Acid ◽

Amino Acid Substitution ◽

Carbapenem Resistance ◽

Resistance Mechanisms ◽

Clinical Isolates ◽

Efflux System ◽

Clinical Strains ◽

Carbapenem Resistant

ABSTRACT In order to define the contributions of the mechanisms for carbapenem resistance in clinical strains of Pseudomonas aeruginosa, we investigated the presence of OprD, the expressions of the MexAB-OprM and MexEF-OprN systems, and the production of the β-lactamases for 44 clinical strains. All of the carbapenem-resistant isolates showed the loss of or decreased levels of OprD. Three strains overexpressed the MexAB-OprM efflux system by carrying mutations inmexR. These three strains had the amino acid substitution in MexR protein, Arg (CGG) → Gln (CAG), at the position of amino acid 70. None of the isolates, however, expressed the MexEF-OprN efflux system. For the characterization of β-lactamases, at least 13 isolates were the depressed mutants, and 12 strains produced secondary β-lactamases. Based on the above resistance mechanisms, the MICs of carbapenem for the isolates were analyzed. The MICs of carbapenem were mostly determined by the expression of OprD. The MICs of meropenem were two- to four-fold increased for the isolates which overexpressed MexAB-OprM in the background of OprD loss. However, the elevated MICs of meropenem for some individual isolates could not be explained. These findings suggested that other resistance mechanisms would play a role in meropenem resistance in clinical isolates of P. aeruginosa.

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Carbapenem-resistance mechanisms of multidrug-resistant Pseudomonas aeruginosa

Journal of Medical Microbiology ◽

10.1099/jmm.0.058354-0 ◽

2013 ◽

Vol 62 (9) ◽

pp. 1317-1325 ◽

Cited By ~ 23

Author(s):

Ester Fusté ◽

Lídia López-Jiménez ◽

Concha Segura ◽

Eusebio Gainza ◽

Teresa Vinuesa ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Outer Membrane ◽

Antimicrobial Agents ◽

Efflux Pump ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Class 1 Integron ◽

Carbonyl Cyanide ◽

Class 1

Clonal dissemination of multidrug-resistant Pseudomonas aeruginosa (MDRPA) is a major concern worldwide. The aim of this study was to explore the mechanisms leading to the carbapenem resistance of an MDRPA clone. Isolates were obtained from a surgical wound, sputum, urine and a blood culture. Pulsed-field gel electrophoresis (PFGE) showed high genomic homogeneity of these isolates and confirmed the circulation of an endemic clone belonging to serotype O4. Outer membrane protein (OMP) bands were visualized by SDS-PAGE, meropenem accumulation was measured in a bioassay and integrons were detected by PCR. Efflux pumps were studied for several antimicrobial agents and synergic combinations thereof in the presence or absence of both carbonyl cyanide m-chlorophenylhydrazone (CCCP) and Phe-Arg-β-naphthylamide (PAβN) at final concentrations of 10 and 40 mg l−1, respectively. On OMP electrophoretic profiles, MDRPA showed a reduction of outer membrane porin D (OprD) and PCR demonstrated the presence of a class 1 integron with a cassette encoding aminoglycoside adenyltransferase B (aadB). Meropenem accumulation was slightly higher in bacilli than in the filamentous cells that formed in the presence of antibiotics. Overexpression of the efflux pump MexAB-OprM and a functional MexXY-OprM were detected in all isolates.

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Comparative genome analysis of multidrug-resistant Pseudomonas aeruginosa JNQH-PA57, a clinically isolated mucoid strain with comprehensive carbapenem resistance mechanisms

BMC Microbiology ◽

10.1186/s12866-021-02203-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Mingju Hao ◽

Wanshan Ma ◽

Xiutao Dong ◽

Xiaofeng Li ◽

Fang Cheng ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Genome Analysis ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Genomic Islands ◽

Genome Database ◽

Genomic Features ◽

Wide Range ◽

Complete Genomes

Abstract Background The prevalence of clinical multidrug-resistant (MDR) Pseudomonas aeruginosa has been increasing rapidly worldwide over the years and responsible for a wide range of acute and chronic infections with high mortalities. Although hundreds of complete genomes of clinical P. aeruginosa isolates have been sequenced, only a few complete genomes of mucoid strains are available, limiting a comprehensive understanding of this important group of opportunistic pathogens. Herein, the complete genome of a clinically isolated mucoid strain P. aeruginosa JNQH-PA57 was sequenced and assembled using Illumina and Oxford nanopore sequencing technologies. Genomic features, phylogenetic relationships, and comparative genomics of this pathogen were comprehensively analyzed using various bioinformatics tools. A series of phenotypic and molecular-genetic tests were conducted to investigate the mechanisms of carbapenem resistance in this strain. Results Several genomic features of MDR P. aeruginosa JNQH-PA57 were identified based on the whole-genome sequencing. We found that the accessory genome of JNQH-PA57 including several prophages, genomic islands, as well as a PAPI-1 family integrative and conjugative element (ICE), mainly contributed to the larger genome of this strain (6,747,067 bp) compared to other popular P. aeruginosa strains (with an average genome size of 6,445,223 bp) listed in Pseudomonas Genome Database. Colony morphology analysis and biofilm crystal staining assay respectively demonstrated an enhanced alginate production and a thicker biofilm formation capability of JNQH-PA57. A deleted mutation at nt 424 presented in mucA gene, resulted in the upregulated expression of a sigma-factor AlgU and a GDP mannose dehydrogenase AlgD, which might explain the mucoid phenotype of this strain. As for the carbapenem resistance mechanisms, our results revealed that the interplay between impaired OprD porin, chromosomal β-lactamase OXA-488 expression, MexAB-OprM and MexXY-OprM efflux pumps overexpression, synergistically with the alginates-overproducing protective biofilm, conferred the high carbapenem resistance to P. aeruginosa JNQH-PA57. Conclusion Based on the genome analysis, we could demonstrate that the upregulated expression of algU and algD, which due to the truncation variant of MucA, might account for the mucoid phenotype of JNQH-PA57. Moreover, the resistance to carbapenem in P. aeruginosa JNQH-PA57 is multifactorial. The dataset presented in this study provided an essential genetic basis for the comprehensive cognition of the physiology, pathogenicity, and carbapenem resistance mechanisms of this clinical mucoid strain.

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Relevance of resistance levels to carbapenems and integron-borne bla IMP-1, bla IMP-7, bla IMP-10 and bla VIM-2 in clinical isolates of Pseudomonas aeruginosa

Journal of Medical Microbiology ◽

10.1099/jmm.0.010017-0 ◽

2009 ◽

Vol 58 (8) ◽

pp. 1080-1085 ◽

Cited By ~ 31

Author(s):

Wei-Hua Zhao ◽

Gelin Chen ◽

Ribu Ito ◽

Zhi-Qing Hu

Keyword(s):

Pseudomonas Aeruginosa ◽

Resistance Genes ◽

Gene Cassette ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Clinical Isolates ◽

Gene Cassettes ◽

Class 1 ◽

High Level ◽

Encoding Genes

Molecular detection and surveillance of the resistance genes harboured by Pseudomonas aeruginosa are becoming increasingly important in assessing and controlling spread and colonization in hospitals, and in guiding the treatment of infections. This study analysed the resistance mechanisms of carbapenem-resistant clinical isolates of P. aeruginosa and identified the associated integron-borne metallo-β-lactamase (MBL)-encoding genes. Twenty-seven imipenem (IPM)-resistant clinical isolates of P. aeruginosa were divided into three groups according to their resistance levels to carbapenems. Strains bearing bla IMP-10 showed extremely high-level resistance to IPM, with MICs of 512–2048 μg ml−1. By comparison, strains bearing bla IMP-1, bla IMP-7 and bla VIM-2 showed an intermediate level of resistance, with MICs of 32–256 μg ml−1. The non-MBL-producing strains showed a low level of resistance, with MICs of 8–32 μg ml−1. The same trend in resistance levels was also observed when resistance to other carbapenems, such as meropenem and panipenem, was determined. DNA sequencing showed that the MBL-encoding gene cassettes were carried by class 1 integrons. The bla IMP-1, bla IMP-7 and bla IMP-10 gene cassettes were preceded by a hybrid P ant promoter, TGGACA-N17-TAAACT, and the bla VIM-2 gene cassette was preceded by a weak promoter, TGGACA-N17-TAAGCT. Most of the MBL-encoding genes were linked to one or two resistance genes encoding aminoglycoside-modifying enzymes, such as aac(6′)Iae, aac(6′)II, aacA7, aacC4, aadA1, aadA2 and aadA6, highlighting the multidrug-resistant properties of these clinical isolates.

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Genetyping of Carbapenem-resistant Organisms Isolated from Clinical Isolates Received from Tertiary Care Hospitals of Ahmedabad, Gujarat

Journal of Pure and Applied Microbiology ◽

10.22207/jpam.15.3.65 ◽

2021 ◽

Vol 15 (3) ◽

pp. 1689-1696

Author(s):

Anurag D. Zaveri ◽

Dilip N. Zaveri ◽

Lakshmi Bhaskaran

Keyword(s):

Drug Resistance ◽

Molecular Characterization ◽

Multiple Drug Resistance ◽

Tertiary Care ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Clinical Isolates ◽

Multiple Drug ◽

Tertiary Care Hospitals

The world is seeing a continuous rise in the levels of antibiotic resistance1. Organisms develop new resistance mechanisms, emerge, and spread the resistance worldwide, making it challenging to treat common infectious diseases. In the current study, clinical isolates received between the years 2017 to 2020 were cultured and the isolated organisms were screened for antibiotic resistance; isolates with multiple drug resistance were further subjected to confirmatory screening through Combined Disc Test (CDT) and Modified Hodge Test (M.H.T.), and molecular characterization to be finally tested for gene expression analysis. Molecular characterization involved screening of genes blaVIM-2, blaKPC-3, blaNDM-1, and blaIMP-11 responsible for imparting carbapenem drug resistance2. From the laboratories of tertiary care hospitals, a total of 1452 clinical isolates were collected and identified. The organisms were subjected to antibiotic susceptibility screening and carbapenem resistance screening. The isolates found positive in the screenings were subjected to molecular characterization for genes, blaVIM-2, blaKPC-3, blaNDM-1, and blaIMP-11, responsible for imparting carbapenem drug resistance. Most of the isolates were resistant variably to aminoglycosides but were found to be resistant to fluoroquinolones and β-lactams group of antibiotics. Carbapenem activity was detected in twelve percent of total isolates and 27 percent among multidrug-resistant isolates. blaNDM-1 gene was found present in 77% isolates, and five organisms among the total number of organisms showed pan drug resistance.

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Interplay of Efflux System, ampC, and oprD Expression in Carbapenem Resistance of Pseudomonas aeruginosa Clinical Isolates

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.50.5.1633-1641.2006 ◽

2006 ◽

Vol 50 (5) ◽

pp. 1633-1641 ◽

Cited By ~ 230

Author(s):

John Quale ◽

Simona Bratu ◽

Jyoti Gupta ◽

David Landman

Keyword(s):

Pseudomonas Aeruginosa ◽

Target Genes ◽

Stop Codon ◽

Premature Stop Codon ◽

Carbapenem Resistance ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Carbapenem Resistant ◽

High Level ◽

Increased Activity

ABSTRACT Carbapenems are important agents for the therapy of infections due to multidrug-resistant Pseudomonas aeruginosa; the development of carbapenem resistance hampers effective therapeutic options. To assess the mechanisms leading to resistance, 33 clinical isolates with differing degrees of carbapenem susceptibility were analyzed for the expression of the chromosomal β-lactamase (ampC), the porin that is important for the entry of carbapenems (oprD), and the proteins involved in four efflux systems (mexA, mexC, mexE, and mexX). Real-time reverse transcriptase PCR was performed using primers and fluorescent probes for each of the target genes. The sequencing of regulatory genes (ampR, mexR, nalC, nalD, mexT, and mexZ) was also performed. Diminished expression of oprD was present in all imipenem- and meropenem-resistant isolates but was not required for ertapenem resistance. Increased expression of ampC was not observed in several isolates that were overtly resistant to carbapenems. Increased expression of several efflux systems was observed in many of the carbapenem-resistant isolates. Increased efflux activity correlated with high-level ertapenem resistance and reduced susceptibility to meropenem and aztreonam. Most isolates with increased expression of mexA had mutations affecting nalC and/or nalD. Two isolates with mutations leading to a premature stop codon in mexZ had markedly elevated mexX expressions, although mutations in mexZ were not a prerequisite for overexpression. β-Lactam resistance in clinical isolates of P. aeruginosa is a result of the interplay between diminished production of oprD, increased activity of ampC, and several efflux systems.

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