Manuela Tietgen1,2, Jan S Kramer3, Steffen Brunst3, Bardya Djahanschiri4, Sonali Wohra1, Paul G Higgins5,6, Marko Weidensdorfer1, Sara Riedel-Christ1, Klaas M Pos7, Aitor Gonzaga8, Matthias Steglich8,9, Ulrich Nübel8,9, Ingo Ebersberger4,10, Ewgenij Proschak3, Stephan Göttig1. 1. Institute of Medical Microbiology and Infection Control, Hospital of the Goethe University, Frankfurt am Main, Germany. 2. Faculty of Biological Sciences of the Goethe University, Frankfurt am Main, Germany. 3. Institute of Pharmaceutical Chemistry, Goethe University, Frankfurt am Main, Germany. 4. Applied Bioinformatics Group, Institute of Cell Biology and Neuroscience, Goethe University, Frankfurt am Main, Germany. 5. Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Cologne, Germany. 6. German Center for Infection Research (DZIF), partner site Bonn-Cologne, Germany. 7. Institute of Biochemistry, Goethe University, Frankfurt am Main, Germany. 8. Leibniz Institute DSMZ - Deutsche Sammlung von Mikroorganismen und Zellkulturen, Braunschweig, Germany. 9. German Center for Infection Research (DZIF), partner site Hannover-Braunschweig, Germany. 10. Senckenberg Biodiversity and Climate Research Centre Frankfurt (BIK-F), Frankfurt am Main, Germany.
Abstract
OBJECTIVES: The aim of this study was to characterize the Acinetobacter calcoaceticus clinical isolate AC_2117 with the novel carbapenem-hydrolysing class D β-lactamase (CHDL) OXA-679. METHODS: Identification of the species and β-lactamases was verified by genome sequencing (PacBio) and phylogenetic analyses. Antibiotic susceptibility of AC_2117 and transformants harbouring cloned blaOXA-679 was evaluated using antibiotic gradient strips and microbroth dilution. OXA-679 was purified heterologously and kinetic parameters were determined using spectrometry or isothermal titration calorimetry. The impact of OXA-679 production during imipenem therapy was evaluated in the Galleria mellonella infection model. RESULTS: Sequencing of the complete genome of the clinical A. calcoaceticus isolate AC_2117 identified a novel CHDL, termed OXA-679. This enzyme shared sequence similarity of 71% to each of the families OXA-143 and OXA-24/40. Phylogenetic analyses revealed that OXA-679 represents a member of a new OXA family. Cloning and expression of blaOXA-679 as well as measurement of kinetic parameters revealed the effective hydrolysis of carbapenems which resulted in reduced susceptibility to carbapenems in Escherichia coli and A. calcoaceticus, and high-level carbapenem resistance in Acinetobacter baumannii. Infection of larvae of G. mellonella with a sublethal dose of blaOXA-679-expressing A. baumannii could not be cured by high-dose imipenem therapy, indicating carbapenem resistance in vivo. CONCLUSIONS: We identified blaOXA-679 in a clinical A. calcoaceticus isolate that represents a member of the new OXA-679 family and that conferred high-level carbapenem resistance in vitro and in vivo.
OBJECTIVES: The aim of this study was to characterize the Acinetobacter calcoaceticus clinical isolate AC_2117 with the novel carbapenem-hydrolysing class D β-lactamase (CHDL) OXA-679. METHODS: Identification of the species and β-lactamases was verified by genome sequencing (PacBio) and phylogenetic analyses. Antibiotic susceptibility of AC_2117 and transformants harbouring cloned blaOXA-679 was evaluated using antibiotic gradient strips and microbroth dilution. OXA-679 was purified heterologously and kinetic parameters were determined using spectrometry or isothermal titration calorimetry. The impact of OXA-679 production during imipenem therapy was evaluated in the Galleria mellonella infection model. RESULTS: Sequencing of the complete genome of the clinical A. calcoaceticus isolate AC_2117 identified a novel CHDL, termed OXA-679. This enzyme shared sequence similarity of 71% to each of the families OXA-143 and OXA-24/40. Phylogenetic analyses revealed that OXA-679 represents a member of a new OXA family. Cloning and expression of blaOXA-679 as well as measurement of kinetic parameters revealed the effective hydrolysis of carbapenems which resulted in reduced susceptibility to carbapenems in Escherichia coli and A. calcoaceticus, and high-level carbapenem resistance in Acinetobacter baumannii. Infection of larvae of G. mellonella with a sublethal dose of blaOXA-679-expressing A. baumannii could not be cured by high-dose imipenem therapy, indicating carbapenem resistance in vivo. CONCLUSIONS: We identified blaOXA-679 in a clinical A. calcoaceticus isolate that represents a member of the new OXA-679 family and that conferred high-level carbapenem resistance in vitro and in vivo.
Authors: Julia I Ries; Marie Heß; Noura Nouri; Thomas A Wichelhaus; Stephan Göttig; Franco H Falcone; Peter Kraiczy Journal: Front Immunol Date: 2022-07-26 Impact factor: 8.786