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Identification and characterization of a novel aac(6')-Iag associated with the blaIMP-1-integron in a multidrug-resistant Pseudomonas aeruginosa.

ABSTRACT: In a continuing study from Dec 2006 to Apr 2008, we characterized nine multi-drug resistant Pseudomonas aeruginosa strains isolated from four patients in a ward at the Hiroshima University Hospital, Japan. Pulsed-field gel electrophoresis of SpeI-digested genomic DNAs from the isolates suggested the clonal expansion of a single strain; however, only one strain, NK0009, was found to produce metallo-?-lactamase. PCR and subsequent sequencing analysis indicated NK0009 possessed a novel class 1 integron, designated as In124, that carries an array of four gene cassettes: a novel aminoglycoside (AG) resistance gene, aac(6')-Iag, blaIMP-1, a truncated form of blaIMP-1, and a truncated form of aac(6')-Iag. The aac(6')-Iag encoded a 167-amino-acid protein that shows 40% identity with AAC(6')-Iz. Recombinant AAC(6')-Iag protein showed aminoglycoside 6'-N-acetyltransferase activity using thin-layer chromatography (TLC) and MS spectrometric analysis. Escherichia coli carrying aac(6')-Iag showed resistance to amikacin, arbekacin, dibekacin, isepamicin, kanamycin, sisomicin, and tobramycin; but not to gentamicin. A conjugation experiment and subsequent Southern hybridization with the gene probes for blaIMP-1 and aac(6')-Ig strongly suggested In124 is on a conjugal plasmid. Transconjugants acquired resistance to gentamicin and were resistant to virtually all AGs, suggesting that the In124 conjugal plasmid also possesses a gene conferring resistance to gentamicin.

SUBMITTER: Kobayashi K

PROVIDER: S-EPMC3741272 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Identification and characterization of a novel aac(6')-Iag associated with the blaIMP-1-integron in a multidrug-resistant Pseudomonas aeruginosa.

Kobayashi Kanao K Hayashi Ikue I Kouda Syuntaro S Kato Fuminori F Fujiwara Tamaki T Kayama Shizuo S Hirakawa Hideki H Itaha Hideyuki H Ohge Hiroki H Gotoh Naomasa N Usui Tsuguru T Matsubara Akio A Sugai Motoyuki M

PloS one 20130812 8

In a continuing study from Dec 2006 to Apr 2008, we characterized nine multi-drug resistant Pseudomonas aeruginosa strains isolated from four patients in a ward at the Hiroshima University Hospital, Japan. Pulsed-field gel electrophoresis of SpeI-digested genomic DNAs from the isolates suggested the clonal expansion of a single strain; however, only one strain, NK0009, was found to produce metallo-β-lactamase. PCR and subsequent sequencing analysis indicated NK0009 possessed a novel class 1 inte ...[more]

PMID: 23950962

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Project description:BACKGROUND:We aimed to identify the main spreading clones, describe the resistance mechanisms associated with carbapenem- and/or multidrug-resistant P. aeruginosa and characterize patients at risk of acquiring these strains in Estonian hospitals. METHODS:Ninety-two non-duplicated carbapenem- and/or multidrug-resistant P. aeruginosa strains were collected between 27th March 2012 and 30th April 2013. Clinical data of the patients was obtained retrospectively from the medical charts. Clonal relationships of the strains were determined by whole genome sequencing and analyzed by multi-locus sequence typing. The presence of resistance genes and beta-lactamases and their origin was determined. Combined-disk method and PCR was used to evaluate carbapenemase and metallo-beta-lactamase production. RESULTS:Forty-three strains were carbapenem-resistant, 11 were multidrug-resistant and 38 were both carbapenem- and multidrug-resistant. Most strains (54%) were isolated from respiratory secretions and caused an infection (74%). Over half of the patients (57%) were???65 years old and 85% had ?1 co-morbidity; 96% had contacts with healthcare and/or had received antimicrobial treatment in the previous 90 days. Clinically relevant beta-lactamases (OXA-101, OXA-2 and GES-5) were found in 12% of strains, 27% of which were located in plasmids. No Ambler class B beta-lactamases were detected. Aminoglycoside modifying enzymes were found in 15% of the strains. OprD was defective in 13% of the strains (all with CR phenotype); carbapenem resistance triggering mutations (F170 L, W277X, S403P) were present in 29% of the strains. Ciprofloxacin resistance correlated well with mutations in topoisomerase genes gyrA (T83I, D87N) and parC (S87 L). Almost all strains (97%) with these mutations showed ciprofloxacin-resistant phenotype. Multi-locus sequence type analysis indicated high diversity at the strain level - 36 different sequence types being detected. Two sequence types (ST108 (n =?23) and ST260 (n =?18)) predominated. Whereas ST108 was associated with localized spread in one hospital and mostly carbapenem-resistant phenotype, ST260 strains occurred in all hospitals, mostly with multi-resistant phenotype and carried different resistance genotype/machinery. CONCLUSIONS:Diverse spread of local rather than international P. aeruginosa strains harboring multiple chromosomal mutations, but not plasmid-mediated Ambler class B beta-lactamases, were found in Estonian hospitals. TRIAL REGISTRATION:This trial was registered retrospectively in ClinicalTrials.gov ( NCT03343119 ).

Dataset Information

Identification and characterization of a novel aac(6')-Iag associated with the blaIMP-1-integron in a multidrug-resistant Pseudomonas aeruginosa.

Publications

Identification and characterization of a novel aac(6')-Iag associated with the blaIMP-1-integron in a multidrug-resistant Pseudomonas aeruginosa.

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