title: Molecular characterization of IMP-type metallo-β-lactamases among multidrug-resistant Achromobacter xylosoxidans.
abstract: OBJECTIVE
  To analyse the metallo-β-lactamase (MBL) genes and their genetic environments in clinical isolates of Achromobacter xylosoxidans.
  
  
  METHODS
  From January 2004 to December 2010, four MBL-producing, multidrug-resistant (MDR) A. xylosoxidans strains were isolated and analysed at a tertiary care university hospital in Japan. Species-level identification was confirmed by 16S ribosomal RNA gene sequencing. Molecular typing was performed using PFGE, the presence of MBL genes was detected using PCR, and integron gene cassettes were examined by cloning and sequence analysis of integron PCR products. The plasmids obtained from individual isolates were analysed based on EcoRI restriction patterns, Southern hybridizations using digoxigenin-labelled probes for bla(IMP-1) and bla(IMP-19) as well as conjugation and transformation experiments.
  
  
  RESULTS
  No clonal relationship was found among the four A. xylosoxidans isolates. Three isolates harboured bla(IMP-1) and one isolate harboured bla(IMP-19). These MBL genes were carried on class 1 integrons. Four different class 1 integron gene cassette arrays were found, including orf1-bla(IMP-1)-aacA4, bla(IMP-1)-bla(IMP-1)-nit1/nit2-catB3-bla(IMP-1)-bla(IMP-1), aacA4-bla(IMP-1) and bla(IMP-19). The restriction pattern of the plasmid DNA obtained from each isolate was unique and the hybridization analyses revealed the presence of each MBL gene within a plasmid. Moreover, all of the plasmids carrying an MBL gene could be transformed into Escherichia coli HST08.
  
  
  CONCLUSIONS
  This study provides genetic evidence for the existence of IMP-type MBL genes in MDR A. xylosoxidans isolates. Moreover, the present findings provide evidence that A. xylosoxidans can receive IMP-type MBL genes via plasmid-mediated transfer, which contributes to their carbapenem resistance.
authors.affiliation: Department of Clinical Laboratory Medicine,  Kyoto University Graduate School of Medicine, Kyoto,  Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Yamamoto
authors.forename: Miki
authors.initials: M
authors.lastname: Nagao
authors.forename: Gou
authors.initials: G
authors.lastname: Hotta
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Matsumura
authors.forename: Aki
authors.initials: A
authors.lastname: Matsushima
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Ito
authors.forename: Shunji
authors.initials: S
authors.lastname: Takakura
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ichiyama
mesh_terms: D042441:Achromobacter denitrificans / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015139:Blotting, Southern; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism; D024901:Drug Resistance, Multiple, Bacterial; D016521:Electrophoresis, Gel, Pulsed-Field; D016905:Gram-Negative Bacterial Infections / Q000382:microbiology*; D006785:Hospitals, University; D006801:Humans; D007564:Japan; D058889:Molecular Typing; D010957:Plasmids / Q000032:analysis; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA; D062606:Tertiary Care Centers; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D015246:Deoxyribonuclease EcoRI; D001618:beta-Lactamases