title: High-level carbapenem resistance in a Citrobacter freundii clinical isolate is due to a combination of KPC-2 production and decreased porin expression.
abstract: An imipenem-resistant isolate of Citrobacter freundii ZJ163 (MIC 256 microg ml(-1)) isolated from a Chinese hospital was investigated. The C. freundii ZJ163 isolate exhibited high-level resistance to carbapenems, penicillins, cephalosporins, cefoxitin, aztreonam, quinolones and aminoglycosides. Isoelectric focusing (IEF) demonstrated three beta-lactamases with pIs of 5.4 (TEM-1), 6.7 (KPC-2) and 7.9 (CTX-M-14). Two different transconjugants (types A and B) were obtained by conjugation studies. The type A transconjugant exhibited reduced susceptibility or resistance to penicillins, cephalosporins and aztreonam, but was susceptible to carbapenems, quinolones and aminoglycosides. The antimicrobial susceptibility patterns of the type B transconjugant were similar to that of type A, except for its significantly reduced carbapenem susceptibility (imipenem MIC 2 microg ml(-1)). IEF, specific PCRs and DNA sequence analysis indicated that the type A transconjugant produced CTX-M-14 beta-lactamase with a pI of 7.9, that the type B transconjugant produced KPC-2 beta-lactamase with a pI of 6.7 and that the beta-lactamase with a pI of 5.4 was TEM-1. PCR analysis and sequencing confirmed the presence of the ampC gene in the chromosomal DNA from C. freundii ZJ163, although no activity of AmpC beta-lactamase was detected by IEF. Urea/SDS-PAGE analysis of outer-membrane proteins revealed that the levels of the 41 and 38 kDa porins were decreased in C. freundii ZJ163. It was concluded that production of KPC-2 combined with decreased expression of porins contributes to high-level resistance to carbapenems in C. freundii ZJ163.
authors.affiliation: Second Affiliated Hospital of Zhejiang University, Zhejiang University, 88 JieFang Road, Hangzhou 310009, PR China.
authors.forename: Rong
authors.initials: R
authors.lastname: Zhang
authors.affiliation: Huazhong University of Science & Technology, Wuhan, Hubei 430074, PR China.
authors.forename: Lijiang
authors.initials: L
authors.lastname: Yang
authors.affiliation: Second Affiliated Hospital of Zhejiang University, Zhejiang University, 88 JieFang Road, Hangzhou 310009, PR China.
authors.forename: Jia Chang
authors.initials: JC
authors.lastname: Cai
authors.affiliation: Second Affiliated Hospital of Zhejiang University, Zhejiang University, 88 JieFang Road, Hangzhou 310009, PR China.
authors.forename: Hong Wei
authors.initials: HW
authors.lastname: Zhou
authors.affiliation: Second Affiliated Hospital of Zhejiang University, Zhejiang University, 88 JieFang Road, Hangzhou 310009, PR China.
authors.forename: Gong-Xiang
authors.initials: GX
authors.lastname: Chen
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015780:Carbapenems / Q000494:pharmacology*; D016971:Citrobacter freundii / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification; D003227:Conjugation, Genetic; D024881:Drug Resistance, Bacterial; D006801:Humans; D015378:Imipenem / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D018272:Porins / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D001618:beta-Lactamases / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D015780:Carbapenems; D018272:Porins; D015378:Imipenem; C491478:beta-lactamase KPC-2; D001618:beta-Lactamases