Unravelling ceftazidime/avibactam resistance of KPC-28, a KPC-2 variant lacking carbapenemase activity. (24th May 2019)

• Record Type: Journal Article
• Authors: Oueslati, Saoussen; Iorga, Bogdan I; Tlili, Linda; Exilie, Cynthia; Zavala, Agustin; Dortet, Laurent; Jousset, Agnès B; Bernabeu, Sandrine; Bonnin, Rémy A; Naas, Thierry
• Is Part Of: Journal of antimicrobial chemotherapy. Volume 74:Number 8(2019)
• Page Start: 2239
• Page End: 2246
• Publication Date: 2019-05-24
• Subjects: Anti-infective agents -- Periodicals; Chemotherapy -- Periodicals; 615.58
• DOI: 10.1093/jac/dkz209
• Languages: English
• ISSNs: 0305-7453

Abstract

Abstract: 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 (Δ242-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 Δ242-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.