Institute of Medical Microbiology and Infection Control, Hospital of the Goethe University, Frankfurt am Main, Germany.
Faculty of Biological Sciences of the Goethe University, Frankfurt am Main, Germany.
J Antimicrob Chemother. 2019 Jun 1;74(6):1494-1502. doi: 10.1093/jac/dkz080.
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.
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.
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.
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.
本研究旨在对具有新型碳青霉烯水解酶（CHDL）OXA-679 的临床分离株 Acinetobacter calcoaceticus AC_2117 进行表型和分子特征分析。
通过全基因组测序（PacBio）和系统发育分析对该菌的种属和β-内酰胺酶进行鉴定。采用抗生素梯度条带法和微量肉汤稀释法评估 AC_2117 及其携带克隆 blaOXA-679 的转化子的药敏性。通过异源表达纯化 OXA-679，采用光谱法或等温滴定量热法测定其动力学参数。利用大蜡螟（Galleria mellonella）感染模型评估 OXA-679 在亚胺培南治疗期间产生的影响。
对临床分离的 A. calcoaceticus 菌株 AC_2117 的全基因组测序鉴定出一种新型 CHDL，命名为 OXA-679。该酶与 OXA-143 和 OXA-24/40 家族的相似度分别为 71%。系统发育分析表明，OXA-679 是一个新的 OXA 家族的成员。blaOXA-679 的克隆和表达以及动力学参数的测定表明，该酶能有效水解碳青霉烯类药物，导致大肠埃希菌和 A. calcoaceticus 对碳青霉烯类药物的敏感性降低，并使鲍曼不动杆菌对碳青霉烯类药物产生高度耐药性。用表达 blaOXA-679 的鲍曼不动杆菌亚致死剂量感染大蜡螟幼虫，即使采用高剂量亚胺培南治疗也无法治愈，表明该菌在体内具有碳青霉烯类耐药性。
我们在临床分离的 A. calcoaceticus 菌株中发现了 blaOXA-679，它属于新型 OXA-679 家族，可导致体外和体内高水平碳青霉烯类耐药性。
