GPC-1, a novel class A carbapenemase detected in a clinical Pseudomonas aeruginosa isolate

2020 ◽  
Vol 75 (4) ◽  
pp. 911-916 ◽  
Author(s):  
Jennifer Schauer ◽  
Sören G Gatermann ◽  
Daniel Hoffmann ◽  
Lars Hupfeld ◽  
Niels Pfennigwerth
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Heterologous Expression ◽  
Biochemical Characterization ◽  
Resistance Mechanism ◽  
Carbapenem Resistance ◽  
Amino Acid Identity ◽  
The Novel ◽  
Bacterial Strains ◽  
Class A ◽  
Carbapenem Resistant

Abstract Objectives To investigate the carbapenem resistance mechanism of a carbapenem-resistant clinical Pseudomonas aeruginosa isolate. Methods A carbapenem-resistant P. aeruginosa isolate was recovered from a tracheal swab from a patient of a general ward in central Germany. Various phenotypic tests confirmed production of a carbapenemase that could not be identified further by PCR. A novel bla gene was identified by WGS and its carbapenemase activity was verified by heterologous expression in an Escherichia coli cloning strain. Kinetic parameters of the novel β-lactamase were determined by spectrophotometric measurements using purified enzyme. Results WGS confirmed the presence of a novel class A carbapenemase. The novel bla gene was named GPC-1 (GPC standing for German Pseudomonas Carbapenemase) and exhibited 77% amino acid identity to BKC-1. WGS also showed that blaGPC-1 was located on the chromosome surrounded by multiple ISs as part of a 26 kb genetic island. Heterologous expression of GPC-1 in E. coli TOP10 led to increased MICs of penicillins, oxyimino-cephalosporins, aztreonam and imipenem, but not of meropenem or ertapenem. Spectrophotometric measurements supported the MIC studies, but detected a slight hydrolysis of ertapenem and meropenem when using high concentrations of purified enzyme. Conclusions The biochemical characterization of GPC-1 emphasizes the ongoing emergence of novel carbapenemases. Strains expressing a weak carbapenemase like GPC-1 might go unrecognized by routine diagnostics due to low MICs for the bacterial strains producing such enzymes.

scholarly journals Investigation of Metallo-Beta-Lactamases in Carbapenem Resistant Pseudomonas aeruginosa Strains by Phenotypic and Genotypic Methods

2020 ◽  
Vol 25 (3) ◽  
pp. 301-307
Author(s):  
M. Duygu Aksoy ◽  
H. Murat Tuğrul
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Ethylenediaminetetraacetic Acid ◽  
Carbapenem Resistance ◽  
Beta Lactamase ◽  
Bacterial Strains ◽  
Beta Lactamases ◽  
Carbapenem Resistant ◽  
Synergy Test ◽  
Phenotypic Tests ◽  
Positive Controls

Introduction: Carbapenem resistant Pseudomonas aeruginosa strains cause serious problems in treatment. A large number of identified metallo-beta-lactamase (MBL) enzymes produced by P. aeruginosa are one of the most important mechanisms in resistance to carbapenems. MBL genes are located on the chromosome or plasmid, and they can easily spread between different bacterial strains. The activities of these enzymes are zinc-dependent, and they are inhibited by ethylenediaminetetraacetic acid (EDTA). Therefore, this advantage is used in MBL identification tests. In this study, it was aimed to determine MBL among P. aeruginosa strains. Materials and Methods: MBL existence was investigated in 35 P. aeruginosa strains accepted to be mildly susceptible/resistant to any of the carbapenem group of antibiotics through phenotypic and genotypic methods. Phenotypic tests were performed as double disk synergy test (DDST), combined disk diffusion tests (CDDT) by using 0.1 M and 0.5 M EDTA, MBL E-test, and modified Hodge test (MHT). blaIMP, blaVIM, blaGIM, blaSIM, blaSPM genes and blaNDM gene were investigated by multiplex polimerase chain reaction (PCR) and PCR, respectively. Escherichia coli ATCC 25922 and P. aeruginosa ATCC 27853 standard bacteria were used in tests. VIM-1, VIM-2, IMP-13, SPM-1, NDM-1 type MBL-producing P. aeruginosa strains were used as positive controls. Results: Among the carbapenems resistant P. aeruginosa isolates, positivity of MBL was found as 54.2% by MBL E-test, 42.8% by DDST, 94.2% and 37.1% by CDDT method using 0.5 M and 0.1 M EDTA, respectively. Modified Hodge test and genotypic method did not detect MBL. Conclusion: In order to correctly evaluate the results of the phenotypic method, the investigation of resistance genes by molecular methods is also required. The most common metallo-beta-lactamase enzymes responsible for resistance to carbapenem in Pseudomonas were not observed. It was thought that different mechanisms might be responsible for the identified carbapenem resistance.

scholarly journals PME-1, an Extended-Spectrum β-Lactamase Identified in Pseudomonas aeruginosa

2011 ◽  
Vol 55 (6) ◽  
pp. 2710-2713 ◽  
Author(s):  
Guo-Bao Tian ◽  
Jennifer M. Adams-Haduch ◽  
Tatiana Bogdanovich ◽  
Hong-Ning Wang ◽  
Yohei Doi
Keyword(s):  
Pseudomonas Aeruginosa ◽  
United Arab Emirates ◽  
Stenotrophomonas Maltophilia ◽  
Hydrolytic Activity ◽  
The United States ◽  
Amino Acid Identity ◽  
The Novel ◽  
Content Type ◽  
Class A ◽  
Extended Spectrum

ABSTRACTA novel extended-spectrum β-lactamase (ESBL) was identified in aPseudomonas aeruginosaclinical isolate obtained from a patient admitted to a hospital in Pennsylvania in 2008. The patient had a prolonged hospitalization in a hospital in Dubai, United Arab Emirates, before being transferred to the United States. The novel ESBL, designated PME-1 (Pseudomonas aeruginosaESBL 1), is a molecular class A, Bush-Jacoby-Medeiros group 2be enzyme and shared 50, 43, and 41% amino acid identity with the L2 β-lactamase ofStenotrophomonas maltophilia, CTX-M-9, and KPC-2, respectively. PME-1 conferred clinically relevant resistance to ceftazidime, cefotaxime, cefepime, and aztreonam inP. aeruginosaPAO1 but not to carbapenems. Purified PME-1 showed good hydrolytic activity against ceftazidime, cefotaxime, and aztreonam, while activity against carbapenems and cefepime could not be measured. PME-1 was inhibited well by β-lactamase inhibitors, including clavulanic acid, sulbactam, and tazobactam. TheblaPME-1gene was carried by an approximately 9-kb plasmid and flanked by tandem ISCR24elements.

Genetic and biochemical characterization of FRI-3, a novel variant of the Ambler class A carbapenemase FRI-1

2019 ◽  
Vol 74 (10) ◽  
pp. 2891-2894 ◽  
Author(s):  
Jennifer Schauer ◽  
Sören G Gatermann ◽  
Matthias Marschal ◽  
Niels Pfennigwerth
Keyword(s):  
Amino Acid ◽  
Biochemical Characterization ◽  
Amino Acid Identity ◽  
Class A ◽  
Carbapenem Resistant ◽  
New Variant ◽  
Novel Variant ◽  
Almost All ◽  
Phenotypic Tests

Abstract Objectives To characterize a new variant of the FRI class A carbapenemase isolated from an MDR clinical Enterobacter cloacae isolate. Methods A carbapenem-resistant E. cloacae was isolated from a rectal swab from a patient in an ICU in Southern Germany. Various phenotypic tests confirmed production of a putative class A carbapenemase. The new bla gene variant, blaFRI-3, and its genetic environment were characterized by WGS. Biochemical characterization was performed by heterologous expression in Escherichia coli TOP10 and by purification of the enzyme with subsequent determination of its kinetic parameters. Results PCR and sequencing carried out for different class A carbapenemase genes confirmed the presence of a novel variant of blaFRI-1. The novel variant was named FRI-3 and exhibited 91%, 96% and 92% amino acid identity to FRI-1, FRI-2 and FRI-4, respectively. E. coli TOP10 expressing blaFRI-3 showed increased resistance to almost all β-lactams. Comparing the catalytic behaviour of FRI-3 and FRI-1, it was shown that FRI-3 had the same substrate spectrum, but basically hydrolysed β-lactams less efficiently than FRI-1. WGS data revealed that blaFRI-3 was located on a 111 kb plasmid. Conclusions The biochemical characterization of FRI-3 illustrates that even a few differences in the amino acid sequence can lead to altered catalytic activities of β-lactamases belonging to the same family.

scholarly journals OXA-198, an Acquired Carbapenem-Hydrolyzing Class D β-Lactamase from Pseudomonas aeruginosa

2011 ◽  
Vol 55 (10) ◽  
pp. 4828-4833 ◽  
Author(s):  
Farid El Garch ◽  
Pierre Bogaerts ◽  
Carine Bebrone ◽  
Moreno Galleni ◽  
Youri Glupczynski
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Carbapenem Resistance ◽  
Amino Acid Identity ◽  
Ventilator Associated Pneumonia ◽  
Class 1 Integron ◽  
Content Type ◽  
New Class ◽  
Carbapenem Resistant ◽  
Class D ◽  
Class 1

ABSTRACTA carbapenem-resistantPseudomonas aeruginosastrain (PA41437) susceptible to expanded-spectrum cephalosporins was recovered from several consecutive lower-respiratory-tract specimens of a patient who developed a ventilator-associated pneumonia while hospitalized in an intensive care unit. Cloning experiments identified OXA-198, a new class D β-lactamase which was weakly related (less than 45% amino acid identity) to other class D β-lactamases. Expression inEscherichia coliTOP10 and inP. aeruginosaPAO1 led to transformants that were resistant to ticarcillin and showed reduced susceptibility to carbapenems and cefepime. TheblaOXA-198gene was harbored by a class 1 integron carried by a ca. 46-kb nontypeable plasmid. This study describes a novel class D β-lactamase involved in carbapenem resistance inP. aeruginosa.

scholarly journals Prevalence of metallo-β-lactamase genes among Pseudomonas aeruginosa isolated from various clinical samples in China

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Wei Wang ◽  
Xiaoya Wang
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Susceptibility ◽  
Ethylenediaminetetraacetic Acid ◽  
Carbapenem Resistance ◽  
Opportunistic Pathogen ◽  
Detection Methods ◽  
Clinical Samples ◽  
Routine Practice ◽  
Carbapenem Resistant ◽  
High Prevalence

AbstractBackgroundPseudomonas aeruginosa is an opportunistic pathogen which is associated with nosocomial infections and causes various diseases including urinary tract infection, pneumonia, soft-tissue infection and sepsis. The emergence of P. aeruginosa-acquired metallo-β-lactamase (MBL) is most worrisome and poses a serious threat during treatment and infection control. The objective of this study was to identify antibiotic susceptibility, phenotypic detection of MBL production and to determine the prevalence of MBL genes in carbapenem-resistant P. aeruginosa isolated from different clinical samples.MethodsA total of 329 non-duplicate P. aeruginosa isolated from various clinical samples from two hospitals in China between September 2017 and March 2019 were included in this study. Phenotypic detection of MBL was performed by the combined detection method using imipenem and imipenem-ethylenediaminetetraacetic acid (EDTA) discs. MBL-encoding genes including blaVIM-1, blaVIM-2, blaIMP-1, blaIMP-2, blaSPM-1, blaSIM, blaNDM-1 and blaGIM were detected by polymerase chain reaction (PCR).ResultsOf the 329 P. aeruginosa, majority of the isolates were resistant to imipenem (77.5%) followed by meropenem (64.7%). Of the 270 P. aeruginosa isolates tested, 149 (55.2%) isolates were found to be positive for MBL detection. Of the different samples, 57.8% (n = 26) of P. aeruginosa isolated from blood were found to be positive for MBL production. Of the various MBL genes, blaIMP-1 (28.2%) was the most predominant gene detected followed by blaVIM-2 (18.8%), blaVIM-1 (16.1%), blaNDM-1 (9.4%), blaIMP-2 (6.7%), blaSIM (6.0%), blaSPM-1 (4.0%) and blaGIM (1.3%) genes.ConclusionsThe high resistance of P. aeruginosa toward imipenem and meropenem and the high prevalence of blaIMP-1 and blaVIM-2 set the alarm on the increasing, perhaps the increased, carbapenem resistance. In addition to routine antibiotic susceptibility testings, our results emphasize the importance of both the phenotypic and genotypic MBL detection methods in routine practice for early detection of carbapenem resistance and to prevent further dissemination of this resistant pathogen.

scholarly journals Presence of Chromosomal crpP-like Genes Is Not Always Associated with Ciprofloxacin Resistance in Pseudomonas aeruginosa Clinical Isolates Recovered in ICU Patients from Portugal and Spain

2021 ◽  
Vol 9 (2) ◽  
pp. 388
Author(s):  
Marta Hernández-García ◽  
María García-Castillo ◽  
Sergio García-Fernández ◽  
Diego López-Mendoza ◽  
Jazmín Díaz-Regañón ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Protein Function ◽  
Resistance Mechanism ◽  
Amino Acid Position ◽  
Genomic Islands ◽  
Missense Mutations ◽  
The Novel ◽  
Surveillance Studies ◽  
Ciprofloxacin Resistance ◽  
Chromosomal Mutations

CrpP enzymes have been recently described as a novel ciprofloxacin-resistance mechanism. We investigated by whole genome sequencing the presence of crpP-genes and other mechanisms involved in quinolone resistance in MDR/XDR-Pseudomonas aeruginosa isolates (n = 55) with both ceftolozane-tazobactam susceptible or resistant profiles recovered from intensive care unit patients during the STEP (Portugal) and SUPERIOR (Spain) surveillance studies. Ciprofloxacin resistance was associated with mutations in the gyrA and parC genes. Additionally, plasmid-mediated genes (qnrS2 and aac(6′)-Ib-cr) were eventually detected. Ten chromosomal crpP-like genes contained in related pathogenicity genomic islands and 6 different CrpP (CrpP1-CrpP6) proteins were found in 65% (36/55) of the isolates. Dissemination of CrpP variants was observed among non-related clones of both countries, including the CC175 (Spain) high-risk clone and CC348 (Portugal) clone. Interestingly, 5 of 6 variants (CrpP1-CrpP5) carried missense mutations in an amino acid position (Gly7) previously defined as essential conferring ciprofloxacin resistance, and decreased ciprofloxacin susceptibility was only associated with the novel CrpP6 protein. In our collection, ciprofloxacin resistance was mainly due to chromosomal mutations in the gyrA and parC genes. However, crpP genes carrying mutations essential for protein function (G7, I26) and associated with a restored ciprofloxacin susceptibility were predominant. Despite the presence of crpP genes is not always associated with ciprofloxacin resistance, the risk of emergence of novel CrpP variants with a higher ability to affect quinolones is increasing. Furthermore, the spread of crpP genes in highly mobilizable genomic islands among related and non-related P. aeruginosa clones alert the dispersion of MDR pathogens in hospital settings.

scholarly journals Characterization of resistance mechanisms of Enterobacter cloacae Complex co-resistant to carbapenem and colistin

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Shixing Liu ◽  
Renchi Fang ◽  
Ying Zhang ◽  
Lijiang Chen ◽  
Na Huang ◽  
...  
Keyword(s):  
Lipid A ◽  
Efflux Pump ◽  
Resistance Mechanism ◽  
Carbapenem Resistance ◽  
Enterobacter Cloacae ◽  
Resistance Mechanisms ◽  
Colistin Resistance ◽  
Carbapenem Resistant ◽  
Horizontal Spread

Abstract Background The emergence of carbapenem-resistant and colistin-resistant ECC pose a huge challenge to infection control. The purpose of this study was to clarify the mechanism of the carbapenems and colistin co-resistance in Enterobacter cloacae Complex (ECC) strains. Results This study showed that the mechanisms of carbapenem resistance in this study are: 1. Generating carbapenemase (7 of 19); 2. The production of AmpC or ESBLs combined with decreased expression of out membrane protein (12 of 19). hsp60 sequence analysis suggested 10 of 19 the strains belong to colistin hetero-resistant clusters and the mechanism of colistin resistance is increasing expression of acrA in the efflux pump AcrAB-TolC alone (18 of 19) or accompanied by a decrease of affinity between colistin and outer membrane caused by the modification of lipid A (14 of 19). Moreover, an ECC strain co-harboring plasmid-mediated mcr-4.3 and blaNDM-1 has been found. Conclusions This study suggested that there is no overlap between the resistance mechanism of co-resistant ECC strains to carbapenem and colistin. However, the emergence of strain co-harboring plasmid-mediated resistance genes indicated that ECC is a potential carrier for the horizontal spread of carbapenems and colistin resistance.

scholarly journals Complete Sequence of pOZ176, a 500-Kilobase IncP-2 Plasmid Encoding IMP-9-Mediated Carbapenem Resistance, from Outbreak Isolate Pseudomonas aeruginosa 96

2013 ◽  
Vol 57 (8) ◽  
pp. 3775-3782 ◽  
Author(s):  
Jianhui Xiong ◽  
David C. Alexander ◽  
Jennifer H. Ma ◽  
Maxime Déraspe ◽  
Donald E. Low ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Carbapenem Resistance ◽  
Complete Sequence ◽  
Open Reading Frames ◽  
Content Type ◽  
Conserved Sequence ◽  
Carbapenem Resistant ◽  
Class 1 ◽  
Usage Analysis ◽  
Type 3

ABSTRACTPseudomonas aeruginosa96 (PA96) was isolated during a multicenter surveillance study in Guangzhou, China, in 2000. Whole-genome sequencing of this outbreak strain facilitated analysis of its IncP-2 carbapenem-resistant plasmid, pOZ176. The plasmid had a length of 500,839 bp and an average percent G+C content of 57%. Of the 618 predicted open reading frames, 65% encode hypothetical proteins. The pOZ176 backbone is not closely related to any plasmids thus far sequenced, but some similarity to pQBR103 ofPseudomonas fluorescensSBW25 was observed. Two multiresistant class 1 integrons and several insertion sequences were identified. TheblaIMP-9-carrying integron containedaacA4→blaIMP-9→aacA4, flanked upstream by Tn21 tnpMRAand downstream by a completetnioperon of Tn402and amermodule, named Tn6016. The second integron carriedaacA4→catB8a→blaOXA-10and was flanked by Tn1403-liketnpRAand asul1-type 3′ conserved sequence (3′-CS), named Tn6217. Other features include three resistance genes similar to those of Tn5, a tellurite resistance operon, and twopiloperons. The replication and maintenance systems exhibit similarity to a genomic island ofRalstonia solanacearumGM1000. Codon usage analysis suggests the recent acquisition ofblaIMP-9. The origins of the integrons on pOZ176 indicated separate horizontal gene transfer events driven by antibiotic selection. The novel mosaic structure of pOZ176 suggests that it is derived from environmental bacteria.

scholarly journals PFM-Like Enzymes Are a Novel Family of Subclass B2 Metallo-β-Lactamases from Pseudomonas synxantha Belonging to the Pseudomonas fluorescens Complex

2019 ◽  
Vol 64 (2) ◽  
Author(s):  
Laurent Poirel ◽  
Mattia Palmieri ◽  
Michael Brilhante ◽  
Amandine Masseron ◽  
Vincent Perreten ◽  
...  
Keyword(s):  
Amino Acid ◽  
Pseudomonas Fluorescens ◽  
Bacterial Species ◽  
Amino Acid Identity ◽  
Chicken Meat ◽  
The Novel ◽  
Content Type ◽  
Acid Identity ◽  
Carbapenem Resistant ◽  
Encoding Genes

ABSTRACT A carbapenem-resistant Pseudomonas synxantha isolate recovered from chicken meat produced the novel carbapenemase PFM-1. That subclass B2 metallo-β-lactamase shared 71% amino acid identity with β-lactamase Sfh-1 from Serratia fonticola. The blaPFM-1 gene was chromosomally located and likely acquired. Variants of PFM-1 sharing 90% to 92% amino acid identity were identified in bacterial species belonging to the Pseudomonas fluorescens complex, including Pseudomonas libanensis (PFM-2) and Pseudomonas fluorescens (PFM-3), highlighting that these species constitute reservoirs of PFM-like encoding genes.

scholarly journals The carbapenem resistance gene blaOXA-23 is disseminated by a conjugative plasmid containing the novel transposon Tn6681 in Acinetobacter johnsonii M19

2020 ◽  
Vol 9 (1) ◽  
Author(s):  
Gongli Zong ◽  
Chuanqing Zhong ◽  
Jiafang Fu ◽  
Yu Zhang ◽  
Peipei Zhang ◽  
...  
Keyword(s):  
Molecular Docking ◽  
Resistance Gene ◽  
Carbapenem Resistance ◽  
Conjugative Plasmid ◽  
The Novel ◽  
Illumina Hiseq ◽  
Discovery Studio ◽  
Acinetobacter Johnsonii ◽  
Type Iv ◽  
Carbapenem Resistant

Abstract Background Carbapenem resistant Acinetobacter species have caused great difficulties in clinical therapy in the worldwide. Here we describe an Acinetobacter johnsonii M19 with a novel blaOXA-23 containing transposon Tn6681 on the conjugative plasmid pFM-M19 and the ability to transferand carbapenem resistance. Methods A. johnsonii M19 was isolated under selection with 8 mg/L meropenem from hospital sewage, and the minimum inhibitory concentrations (MICs) for the representative carbapenems imipenem, meropenem and ertapenem were determined. The genome of A. johnsonii M19 was sequenced by PacBio RS II and Illumina HiSeq 4000 platforms. A homologous model of OXA-23 was generated, and molecular docking models with imipenem, meropenem and ertapenem were constructed by Discovery Studio 2.0. Type IV secretion system and conjugation elements were identified by the Pathosystems Resource Integration Center (PATRIC) server and the oriTfinder. Mating experiments were performed to evaluate transfer of OXA-23 to Escherichia coli 25DN. Results MICs of A. johnsonii M19 for imipenem, meropenem and ertapenem were 128 mg/L, 48 mg/L and 24 mg/L, respectively. Genome sequencing identified plasmid pFM-M19, which harbours the carbapenem resistance gene blaOXA-23 within the novel transposon Tn6681. Molecular docking analysis indicated that the elongated hydrophobic tunnel of OXA-23 provides a hydrophobic environment and that Lys-216, Thr-217, Met-221 and Arg-259 were the conserved amino acids bound to imipenem, meropenem and ertapenem. Furthermore, pFM-M19 could transfer blaOXA-23 to E. coli 25DN by conjugation, resulting in carbapenem-resistant transconjugants. Conclusions Our investigation showed that A. johnsonii M19 is a source and disseminator of blaOXA-23 and carbapenem resistance. The ability to transfer blaOXA-23 to other species by the conjugative plasmid pFM-M19 raises the risk of spread of carbapenem resistance. Graphic abstract The carbapenem resistance gene blaOXA-23 is disseminated by a conjugative plasmid containing the novel transposon Tn6681 in Acinetobacter johnsonii M19.

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