期刊
JOURNAL OF ANTIMICROBIAL CHEMOTHERAPY
卷 74, 期 8, 页码 2239-2246出版社
OXFORD UNIV PRESS
DOI: 10.1093/jac/dkz209
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资金
- Assistance Publique-Hopitaux de Paris
- Universite Paris-Sud [EA 7361]
- LabEx LERMIT
- French National Research Agency [ANR-10-LABX-33]
- Joint Programming Initiative on Antimicrobial Resistance [ANR-14-JAMR-0002]
- Agence Nationale de la Recherche (ANR) [ANR-14-JAMR-0002] Funding Source: Agence Nationale de la Recherche (ANR)
Background: KPC-like carbapenemases have spread worldwide with more than 30 variants identified that differ by single or double amino-acid substitutions. Objectives: To describe the steady-state kinetic parameters of KPC-28, which differs from KPC-2 by a H274Y substitution and the deletion of two amino acids (Delta 242-GT-243). Methods: The bla(KPC-2), bla(KPC-3), bla(KPC-14) and bla(KPC-28) genes were cloned into a pTOPO vector for susceptibility testing or into pET41b for overexpression, purification and subsequent kinetic parameter (K-m, k(cat)) determination. Molecular docking experiments were performed to explore the role of the amino-acid changes in the carbapenemase activity. Results: Susceptibility testing revealed that Escherichia coli producing KPC-28 displayed MICs that were lower for carbapenems and higher for ceftazidime and ceftazidime/avibactam as compared with KPC-2. The catalytic efficiencies of KPC-28 and KPC-14 for imipenem were 700-fold and 200-fold lower, respectively, than those of KPC-2, suggesting that Delta 242-GT-243 in KPC-28 and KPC-14 is responsible for reduced carbapenem hydrolysis. Similarly, the H274Y substitution resulted in KPC-28 in a 50-fold increase in ceftazidime hydrolysis that was strongly reversed by clavulanate. Conclusions: We have shown that KPC-28 lacks carbapenemase activity, has increased ceftazidime hydrolytic activity and is strongly inhibited by clavulanate. KPC-28-producing E. coli isolates display an avibactam-resistant ESBL profile, which may be wrongly identified by molecular and immunochromatographic assays as the presence of a carbapenemase. Accordingly, confirmation of carbapenem hydrolysis will be mandatory with assays based solely on bla(KPC) gene or gene product detection.
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