Dissemination of imipenem-resistant Acinetobacter baumannii with new plasmid-borne bla OXA-72 in Taiwan

Abstract

BackgroundThe systemic surveillance of imipenem-resistant Acinetobacter baumannii (IRAB) from multicenters in Taiwan revealed the emergence of isolates with blaOXA-72. This study described their genetic makeup, mechanism of spread, and contribution to carbapenem resistance.MethodsTwo hundred and ninety-one non-repetitive isolates of A. baumannii were collected from 10 teaching hospitals from different geographical regions in Taiwan from June 2007 to September 2007. Minimal inhibitory concentrations (MICs) were determined by agar dilution. Clonality was determined by pulsed-field gel electrophoresis. Plasmid was extracted and digested by restriction enzymes, and subsequently analyzed by electrophoresis and Southern blot for blaOXA-72. The flanking regions of blaOXA-72 were determined by inverse PCR. The contribution of blaOXA-72 to imipenem MIC was determined by transforming plasmids carrying blaOXA-72 into imipenem-susceptible A. baumannii.ResultsAmong 142 IRAB in Taiwan, 27 harbored blaOXA-72; 22 originated from Southern Taiwan, 5 from Central Taiwan, and none from Northern Taiwan. There were two major clones. The blaOXA-72 was identified in the plasmids of all isolates. Two genetic structures flanking plasmid-borne blaOXA-72 were identified and shared identical sequences in certain regions; the one described in previous literature was present in only one isolate, and the new one was present in the remaining isolates. Introduction of blaOXA-72 resulted in an increase of imipenem MIC in the transformants. The overexpression of blaOXA-72 mRNA in response to imipenem further supported the contribution of blaOXA-72.ConclusionsIn conclusion, isolates with new plasmid-borne blaOXA-72 were found to be disseminated successfully in Southern Taiwan. The spread of the resistance gene depended on clonal spread and dissemination of a new plasmid. BlaOXA-72 in these isolates directly led to their imipenem-resistance.

First Occurrence of OXA-72-Producing Acinetobacter baumannii in Serbia

“…OXA-72 is a single-amino-acid variant of OXA-24/40 (G224D). In contrast to OXA-23, which is the most common acquired carbapenemase in A. baumannii worldwide, OXA-72-producing isolates have been reported in only a few countries, including Brazil, Colombia, Taiwan, France, Croatia, Poland, Italy, Lithuania, and Sweden (19,21,(23)(24)(25)(26)(27)(28)(29)(30). The clonal spread of OXA-72-producing A. baumannii isolates was previously reported in the Balkan region, in a Croatian university hospital (24).…”

OXA β-Lactamases

“…Until recently, the bla OXA-40-like genes appeared to be restricted largely to the Iberian Peninsula, where they have been the predominant acquired OXA-type ␤-lactamase (133)(134)(135). However, reports have identified bla OXA-40-like genes in more distant locations, such as the United States (136) and Asia (31), and in particular, there are increasingly reports of these enzymes being identified on plasmids in Taiwan (137,138). It may be that a more recent transfer onto plasmids by this enzyme group has limited its spread to date.…”

“…Another commonly used method is repetitive sequence-based PCR (rep-PCR) using the DiversiLab system (52), and the discriminative power of this system is similar to that of PFGE. The use of these techniques and comparing them with bla OXA gene content illustrated the degree of mobility that the bla OXA genes have, where isolates with the same PFGE or rep-PCR profile vary in their bla OXA gene content (137,138,167) or isolates with different profiles can have the same bla OXA gene content (152,167,168). The use of such techniques with high discriminatory power is useful in outbreak situations, where, when used in conjunction with bla OXA gene screening, they can be used to detect small changes in local epidemiology, such as the acquisition of a resistance gene or conversion between one allele of a resistance gene and another (55,169).…”

Comparative Phosphoproteomics Reveals the Role of AmpC β-lactamase Phosphorylation in the Clinical Imipenem-resistant Strain Acinetobacter baumannii SK17

“…These recombinant pYMAb2 plasmids were transformed into the kanamycin-susceptible strain Ab290 by electroporation using a gene pulser electroporator (Bio-Rad) and 2-mm electrode gap cuvettes (7,28,39). Briefly, electrocompetent cells were prepared as follows: overnight cultures of strain Ab290 were diluted 1:100 and grown to the exponential phase (OD 600 nm ϭ 0.5-0.7).…”

“…The conserved forward primer sequence of ISAba1, located upstream of ampC to facilitate efficient production of the enzyme, and the conserved reverse primer of ampC were used in this study (32). The PCR product was cloned between the XbaI and XhoI sites of pYMAb2 and transformed into chemically competent Escherichia coli JM109 cells (39). The primers for ISAba1-ampC wildtype (WT) and the site-directed mutants (S81A, S81D, S88A, S88D, S90A, S90D, S88A/S90A, S88A/S90D, S88D/S90A, S88D/S90D, S81A/S90A, S81A/S90D, S81D/S90A, and S81D/S90D) were synthesized based on the criteria of site-directed ligation-independent mutagenesis (SLIM) (supplemental Table S7) (41).…”