scholarly journals Genetic and Functional Analysis of the Chromosome-Encoded Carbapenem-Hydrolyzing Oxacillinase OXA-40 of Acinetobacter baumannii

2003 ◽  
Vol 47 (1) ◽  
pp. 268-273 ◽  
Author(s):  
Claire Héritier ◽  
Laurent Poirel ◽  
Daniel Aubert ◽  
Patrice Nordmann
Keyword(s):  
Acinetobacter Baumannii ◽  
Nucleotide Substitution ◽  
Hydrolytic Activity ◽  
Cloning And Expression ◽  
Mutant Enzyme ◽  
Gene Encoding ◽  
Narrow Spectrum ◽  
Class D ◽  
Lactamase Gene ◽  
Dna Mutagenesis

ABSTRACT Clinical isolate Acinetobacter baumannii CLA-1 was resistant to a series of antibiotic molecules, including carbapenems. Cloning and expression of the β-lactamase gene content of this isolate in Escherichia coli DH10B identified a chromosome-encoded oxacillinase, OXA-40, that differed by one or two amino acid changes from OXA-24, -25, and -26 and an AmpC-type cephalosporinase. The OXA-40 β-lactamase had a mainly narrow-spectrum hydrolytic profile, but it included ceftazidime and imipenem. Its activity was resistant to inhibition by clavulanic acid, tazobactam, sulbactam, and, like most of the other carbapenem-hydrolyzing oxacillinases, NaCl. OXA-40 had an FGN triad replacing a YGN motif at class D β-lactamase (DBL) positions 144 to 146. Site-directed DNA mutagenesis leading to a Phe-to-Tyr change at DBL position 144 in OXA-40 gave a mutant enzyme with increased hydrolytic activity against most β-lactams, including imipenem. Conversely, with a gene encoding the narrow-spectrum oxacillinase OXA-1 as the template, a nucleotide substitution leading to a Tyr-to-Phe change in the YGN motif of OXA-1 gave a mutant enzyme with decreased hydrolytic activity without an increase in carbapenem-hydrolyzing activity. Thus, the Phe residue in the FGN motif was not associated with carbapenem-hydrolyzing activity by itself but instead was associated with weak overall hydrolytic activity. Finally, this Phe residue in OXA-40 explained resistance to inhibition by NaCl whereas a Tyr residue in motif YGN was related to susceptibility to NaCl.

scholarly journals Identification of the Novel Narrow-Spectrum β-Lactamase SCO-1 in Acinetobacter spp. from Argentina

2007 ◽  
Vol 51 (6) ◽  
pp. 2179-2184 ◽  
Author(s):  
Laurent Poirel ◽  
Stéphane Corvec ◽  
Melina Rapoport ◽  
Pauline Mugnier ◽  
Alejandro Petroni ◽  
...  
Keyword(s):  
Amino Acid Identity ◽  
The Novel ◽  
Gene Encoding ◽  
Class A ◽  
Narrow Spectrum ◽  
A Value ◽  
Acinetobacter Spp ◽  
Encoding Gene ◽  
High Level ◽  
Lactamase Gene

ABSTRACT By studying the β-lactamase content of several Acinetobacter spp. isolates from Argentina, producing the expanded-spectrum β-lactamases (ESBL) VEB-1a or PER-2, a novel Ambler class A β-lactamase gene was identified. It encoded the narrow-spectrum β-lactamase SCO-1, whose activity was inhibited by clavulanic acid. SCO-1 hydrolyzes penicillins at a high level and cephalosporins and carbapenems at a very low level. β-Lactamase SCO-1 was identified from unrelated VEB-1a-positive or PER-2-positive Acinetobacter spp. isolates recovered from three hospitals. The bla SCO-1 gene was apparently located on a plasmid of ca. 150 kb from all cases but was not associated with any ESBL-encoding gene. The G+C content of the bla SCO gene was 52%, a value that does not correspond to that of the A. baumannii genome (39%). β-Lactamase SCO-1 shares 47% amino acid identity with CARB-5 and ca. 40% with the enzymes TEM, SHV, and CTX-M. A gene encoding a putative resolvase was identified downstream of the bla SCO-1 gene, but its precise way of acquisition remains to be determined.

scholarly journals Genetics and Expression of the Carbapenem-Hydrolyzing Oxacillinase Gene blaOXA-23 in Acinetobacter baumannii

2007 ◽  
Vol 51 (4) ◽  
pp. 1530-1533 ◽  
Author(s):  
Stéphane Corvec ◽  
Laurent Poirel ◽  
Thierry Naas ◽  
Henri Drugeon ◽  
Patrice Nordmann
Keyword(s):  
Acinetobacter Baumannii ◽  
The Novel ◽  
Gene Encoding ◽  
Promoter Sequences ◽  
Insertion Elements ◽  
Genetic Structures ◽  
Lactamase Gene

ABSTRACT The genetic structures surrounding the plasmid-carried bla OXA-23 oxacillinase gene, encoding resistance to carbapenems, were studied in Acinetobacter baumannii. ISAba1 and the novel element ISAba4 were detected upstream of the bla OXA-23 gene, providing promoter sequences for its expression. These insertion elements were likely involved in transposition processes at the origin of acquisition of this β-lactamase gene.

scholarly journals Genetic and Biochemical Characterization of FUS-1 (OXA-85), a Narrow-Spectrum Class D β-Lactamase from Fusobacterium nucleatum subsp. polymorphum

2006 ◽  
Vol 50 (8) ◽  
pp. 2673-2679 ◽  
Author(s):  
Christine Voha ◽  
Jean-Denis Docquier ◽  
Gian Maria Rossolini ◽  
Thierry Fosse
Keyword(s):  
Biochemical Characterization ◽  
Fusobacterium Nucleatum ◽  
Cloning And Sequencing ◽  
Gene Encoding ◽  
Brachyspira Pilosicoli ◽  
Reading Frame ◽  
New Class ◽  
Class D ◽  
Lactamase Gene

ABSTRACT Previous studies have reported β-lactamase-mediated penicillin resistance in Fusobacterium nucleatum, but no β-lactamase gene has yet been identified in this species. An F. nucleatum subsp. polymorphum strain resistant to penicillin and amoxicillin was isolated from a human periodontitis sample. DNA cloning and sequencing revealed a 765-bp open reading frame encoding a new class D β-lactamase named FUS-1 (OXA-85). A recombinant Escherichia coli strain carrying the bla FUS-1 gene exhibited resistance to amoxicillin with a moderate decrease in the MICs with clavulanic acid. The bla FUS-1 gene was found in two additional clonally unrelated F. nucleatum subsp. polymorphum isolates. It was located on the chromosome in a peculiar genetic environment where a gene encoding a putative transposase-like protein is found, suggesting a possible acquisition of this class D β-lactamase gene. The FUS-1 enzyme showed the closest ancestral relationship with OXA-63 from Brachyspira pilosicoli (53% identity) and with putative chromosomal β-lactamases of Campylobacter spp. (40 to 42% identity). FUS-1 presents all of the conserved structural motifs of class D β-lactamases. Kinetic analysis revealed that FUS-1 exhibits a narrow substrate profile, efficiently hydrolyzing benzylpenicillin and oxacillin. FUS-1 was poorly inactivated by clavulanate and NaCl. FUS-1 is the first example of a class D β-lactamase produced by a gram-negative, anaerobic, rod-shaped bacterium to be characterized.

scholarly journals Genetic and Biochemical Characterization of a Chromosome-Encoded Carbapenem-Hydrolyzing Ambler Class D β-Lactamase from Shewanella algae

2004 ◽  
Vol 48 (5) ◽  
pp. 1670-1675 ◽  
Author(s):  
Claire Héritier ◽  
Laurent Poirel ◽  
Patrice Nordmann
Keyword(s):  
Escherichia Coli ◽  
Clinical Isolate ◽  
Biochemical Characterization ◽  
Resistance Phenotype ◽  
Narrow Spectrum ◽  
Class D ◽  
Shewanella Algae ◽  
Lactamase Gene

ABSTRACT A chromosome-encoded β-lactamase gene from Shewanella algae clinical isolate KB-1 was cloned and expressed in Escherichia coli. It encoded the Ambler class D enzyme OXA-55, sharing less than 55% identity with any other oxacillinases. Although conferring a narrow-spectrum β-lactam resistance phenotype, OXA-55 had carbapenem-hydrolyzing activity that mirrored the reduced susceptibility to imipenem observed in S. algae KB-1. Very similar oxacillinases were found in other S. algae isolates.

scholarly journals OXA-58, a Novel Class D β-Lactamase Involved in Resistance to Carbapenems in Acinetobacter baumannii

2005 ◽  
Vol 49 (1) ◽  
pp. 202-208 ◽  
Author(s):  
Laurent Poirel ◽  
Sophie Marqué ◽  
Claire Héritier ◽  
Christine Segonds ◽  
Gérard Chabanon ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acinetobacter Baumannii ◽  
Reference Strain ◽  
Carbapenem Resistance ◽  
Amino Acid Identity ◽  
Cloning And Expression ◽  
Acid Identity ◽  
Carbapenem Resistant ◽  
Class D ◽  
Insertion Elements

ABSTRACT A carbapenem-resistant Acinetobacter baumannii strain was isolated in Toulouse, France, in 2003. Cloning and expression in Escherichia coli identified the carbapenem-hydrolyzing β-lactamase OXA-58, which is weakly related (less than 50% amino acid identity) to other oxacillinases. It hydrolyzed penicillins, oxacillin, and imipenem but not expanded-spectrum cephalosporins. The bla OXA-58 gene was located on a ca. 30-kb non-self-transferable plasmid. After electrotransformation in the A. baumannii CIP7010T reference strain, it conferred reduced susceptibility to carbapenems. The bla OXA-58 gene was bracketed by two novel ISAba3-like insertion elements. This study describes a newly characterized β-lactamase that may contribute to carbapenem resistance in A. baumannii.

scholarly journals Clinical Variants of the Native Class D β-Lactamase of Acinetobacter baumannii Pose an Emerging Threat through Increased Hydrolytic Activity against Carbapenems

2016 ◽  
Vol 60 (10) ◽  
pp. 6155-6164 ◽  
Author(s):  
Emma C. Schroder ◽  
Zachary L. Klamer ◽  
Aysegul Saral ◽  
Kyle A. Sugg ◽  
Cynthia M. June ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Active Site ◽  
Hydrolytic Activity ◽  
Single Amino Acid ◽  
Kinetic Properties ◽  
Active Site Residues ◽  
Content Type ◽  
Class D ◽  
Clinical Variants ◽  
Dynamics Simulations

ABSTRACTThe threat posed by the chromosomally encoded class D β-lactamase ofAcinetobacter baumannii(OXA-51/66) has been unclear, in part because of its relatively low affinity and turnover rate for carbapenems. Several hundred clinical variants of OXA-51/66 have been reported, many with substitutions of active-site residues. We determined the kinetic properties of OXA-66 and five clinical variants with respect to a wide variety of β-lactam substrates. The five variants displayed enhanced activity against carbapenems and in some cases against penicillins, late-generation cephalosporins, and the monobactam aztreonam. Molecular dynamics simulations show that in OXA-66, P130 inhibits the side-chain rotation of I129 and thereby prevents doripenem binding because of steric clash. A single amino acid substitution at this position (P130Q) in the variant OXA-109 greatly enhances the mobility of both I129 and a key active-site tryptophan (W222), thereby facilitating carbapenem binding. This expansion of substrate specificity represents a very worrisome development for the efficacy of β-lactams against this troublesome pathogen.

scholarly journals Characterization of OXA-25, OXA-26, and OXA-27, Molecular Class D β-Lactamases Associated with Carbapenem Resistance in Clinical Isolates of Acinetobacter baumannii

2001 ◽  
Vol 45 (2) ◽  
pp. 583-588 ◽  
Author(s):  
Mariya Afzal-Shah ◽  
Neil Woodford ◽  
David M. Livermore
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Acinetobacter Baumannii ◽  
Sequence Data ◽  
Carbapenem Resistance ◽  
Hydrolytic Activity ◽  
Amino Acid Homology ◽  
Class D ◽  
Weak Activity

ABSTRACT Carbapenem resistance in Acinetobacter spp. is increasingly being associated with OXA-type β-lactamases with weak hydrolytic activity against imipenem and meropenem. Such enzymes were characterized from Acinetobacter isolates collected in Belgium, Kuwait, Singapore, and Spain. The isolates from Spain and Belgium had novel class D β-lactamases that were active against carbapenems. These were designated OXA-25 and OXA-26, respectively, and had >98% amino acid homology with each other and with the OXA-24 enzyme recently described by others from an Acinetobacterisolate collected elsewhere in Spain. The isolate from Singapore had OXA-27 β-lactamase, another novel class D type with only 60% homology to OXA-24, -25, and -26, but with 99% homology to OXA-23 (ARI-1), described previously from an Acinetobacter baumannii isolate collected in Scotland. Sequence data were not obtained for the carbapenem-hydrolyzing OXA enzyme from the isolate from Kuwait; nevertheless, the enzyme was phenotypically similar to OXA-25 and -26. The enzymes OXA-23, -24, -25, -26, and -27 retained the STFK and SXV motifs typical of class D β-lactamases, but the YGN motif was altered to FGN. The KTG motif was retained by OXA-27 and -23 but was replaced by KSG in OXA-24, -25, and -26. OXA-25 and -26 enzymes were strongly active against oxacillin, but unusually for an OXA-type β-lactamase, OXA-27 had apparently weak activity, although measurement was complicated by biphasic kinetics. None of the new enzymes was transmissible to Escherichia coli recipients. Many Acinetobacter isolates are multiresistant to other antibiotics, and the emergence of class D enzymes with carbapenem-hydrolyzing activity is a disturbing development for antimicrobial chemotherapy.

scholarly journals Genetic and Biochemical Characterization of the First Extended-Spectrum CARB-Type ß-Lactamase, RTG-4, from Acinetobacter baumannii

2009 ◽  
Vol 53 (7) ◽  
pp. 3010-3016 ◽  
Author(s):  
Anaïs Potron ◽  
Laurent Poirel ◽  
Jacques Croizé ◽  
Vanessa Chanteperdrix ◽  
Patrice Nordmann
Keyword(s):  
Acinetobacter Baumannii ◽  
Reference Strain ◽  
Biochemical Characterization ◽  
Repeat Sequence ◽  
Cloning And Expression ◽  
Class A ◽  
Extended Spectrum ◽  
Ca 50 ◽  
Lactamase Gene

ABSTRACT Acinetobacter baumannii isolate KAR was uncommonly more resistant to cefepime and cefpirome than to ceftazidime and cefotaxime. Cloning and expression of the β-lactamase gene content of this isolate into Escherichia coli TOP10 identified ß-lactamase RTG-4 (or CARB-10), which corresponds to the first reported extended-spectrum CARB-type enzyme. RTG-4 is a plasmid-encoded Ambler class A β-lactamase whose sequence differs by 4 amino acid substitutions from the narrow-spectrum β-lactamase RTG-3. RTG-4 hydrolyzes cefepime and cefpirome and weakly hydrolyzes ceftazidime due to the single Ser-to-Thr substitution at Ambler position 69. RTG-4 is less susceptible to inhibition by tazobactam and sulbactam than RTG-3. Expression of β-lactamase RTG-4 in a wild-type A. baumannii reference strain showed that it conferred resistance to cefepime and cefpirome. The genetic environment of the bla RTG-4 gene was made of a peculiar transposon located on a ca. 50-kb plasmid. ISAba9, located upstream of bla RTG-4, may be responsible for its acquisition by recognizing a secondary right inverted repeat sequence, thus acting by a one-ended transposition process.

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