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Novel genetic environment of the plasmid-mediated KPC-3 gene detected in Escherichia coli and Citrobacter freundii isolates from China.

ABSTRACT: The imipenem and meropenem-resistant strains Citrobacter freundii HS70 and Escherichia coli HS510 were isolated from patients in Shanghai, China. By isoelectric focusing, PCR amplification and sequencing, these strains were each found to produce four ?-lactamases: TEM-1, KPC-3, SHV-7 and CTX-M-14. A conjugation experiment and plasmid restriction digestion revealed that the bla (KPC-3) gene was located on the same plasmid in both isolates. Bidirectional primer walking sequencing showed that the nucleotide sequence surrounding the 3.8 kb bla(KPC-3) contained a 671-bp insertion similar to that previously characterized in China. The insertion was located between the promoter and the coding region of the bla(KPC-3) gene. Susceptibility testing performed on recombinant strains carrying the bla(KPC-3) gene with or without the insertion revealed that minimum inhibitory concentrations of imipenem, meropenem, cefepime, and cefotaxime for E. coli EMU-KPC3 (without insertion) were four times higher than that of E. coli EKPC3 (with insertion). The 671 bp insertion reduced bla(KPC-3) expression significantly. Taken together, these results suggest that KPC-3-producing C. freundii and E. coli have begun to emerge in our hospital.

SUBMITTER: Li G

PROVIDER: S-EPMC3052496 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Novel genetic environment of the plasmid-mediated KPC-3 gene detected in Escherichia coli and Citrobacter freundii isolates from China.

Li G G Wei Q Q Wang Y Y Du X X Zhao Y Y Jiang X X

European journal of clinical microbiology & infectious diseases : official publication of the European Society of Clinical Microbiology 20101214 4

The imipenem and meropenem-resistant strains Citrobacter freundii HS70 and Escherichia coli HS510 were isolated from patients in Shanghai, China. By isoelectric focusing, PCR amplification and sequencing, these strains were each found to produce four β-lactamases: TEM-1, KPC-3, SHV-7 and CTX-M-14. A conjugation experiment and plasmid restriction digestion revealed that the bla (KPC-3) gene was located on the same plasmid in both isolates. Bidirectional primer walking sequencing showed that the n ...[more]

PMID: 21153909

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Project description:Although quinolone resistance usually results from chromosomal mutations, recent studies indicate that quinolone resistance can also be plasmid mediated. The gene responsible, qnr, is distinct from the known quinolone resistance genes and in previous studies seemed to be restricted to Klebsiella pneumoniae and Escherichia coli isolates from the University of Alabama in Birmingham, where this resistance was discovered. In Shanghai, the frequency of ciprofloxacin resistance in E. coli has exceeded 50% since 1993. Seventy-eight unique ciprofloxacin-resistant clinical isolates of E. coli from Shanghai hospitals were screened for the qnr gene by colony blotting and Southern hybridization of plasmid DNA. Conjugation experiments were done with azide-resistant E. coli J53 as a recipient with selection for plasmid-encoded antimicrobial resistance (chloramphenicol, gentamicin, or tetracycline) and azide counterselection. qnr genes were sequenced, and the structure of the plasmid DNA adjacent to qnr was analyzed by primer walking with a sequential series of outward-facing sequencing primers with plasmid DNA templates purified from transconjugants. Six (7.7%) of 78 strains gave a reproducible hybridization signal with a qnr gene probe on colony blots and yielded strong signals on plasmid DNA preparations. Quinolone resistance was transferred from all six probe-positive strains. Transconjugants had 16- to 250-fold increases in the MICs of ciprofloxacin relative to that of the recipient. All six strains contained qnr with a nucleotide sequence identical to that originally reported, except for a single nucleotide change (CTA-->CTG at position 537) encoding the same amino acid. qnr was located in complex In4 family class 1 integrons. Two completely sequenced integrons were designated In36 and In37. Transferable plasmid-mediated quinolone resistance associated with qnr is thus prevalent in quinolone-resistant clinical strains of E. coli from Shanghai and may contribute to the rapid increase in bacterial resistance to quinolones in China.

Dataset Information

Novel genetic environment of the plasmid-mediated KPC-3 gene detected in Escherichia coli and Citrobacter freundii isolates from China.

Publications

Novel genetic environment of the plasmid-mediated KPC-3 gene detected in Escherichia coli and Citrobacter freundii isolates from China.

Similar Datasets

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