Liu Zeshi, Shen Siquan, Lei Jing, Zhao Shuai, Zhang Xue, Tang Chengkang, Wu Shi, Lei Ke, Yin Jian, Zhang Yanping, Guo Yan, Geng Yan, Hu Fupin
Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China; Department of Clinical Laboratory, The Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, China.
Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China; Key Laboratory of Clinical Pharmacology of Antibiotics, Ministry of Health, Shanghai, China.
Int J Antimicrob Agents. 2025 Sep;66(3):107536. doi: 10.1016/j.ijantimicag.2025.107536. Epub 2025 May 13.
This study investigates the transfer mechanism and functional properties of OXA-1224, the novel d-class β-lactase in the clinical isolate Acinetobacter pittii (A. pittii) PT-01.
Bacterial identification was performed using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and resistance genes were identified via polymerase chain reaction (PCR) and DNA sequencing. Antimicrobial susceptibility of A. pittii PT-01 and its recombinant transformants carrying bla was determined by broth microdilution method. Purified OXA-213 and OXA-1224 proteins were evaluated by the enzymatic reaction kinetics test.
A. pittii PT-01 was susceptible to amikacin, tigecycline, polymyxin B, ceftazidime, cefepime, cefoperazone-sulbactam, and piperacillin-tazobactam, but resistant to imipenem and meropenem. The strain harboured bla and a novel bla variant: bla. Recombinant strains expressing bla showed an 8-fold and 16-fold increase in minimum inhibitory concentrations (MICs) for imipenem and meropenem, respectively, compared to the control strain Acinetobacter baumannii (A. baumannii) ATCC 17978. Enzymatic assays indicated that OXA-1224 had a higher Km value for nitrocefin and stronger affinity for imipenem (Km = 496.7) and meropenem (Km = 126.1) compared to OXA-213. Catalytic efficiency for imipenem and meropenem was also increased with OXA-1224 compared to OXA-213. OXA-1224 has a higher affinity for imipenem (Km/Kcat = 3.362) and meropenem (Km/Kcat = 4.837) compared to OXA-213 (Km/Kcat = 2.469, Km /Kcat = 2.547, respectively). However, OXA-1224 showed minimal activity against the third-generation cephalosporin ceftazidime (Km > 1000).
The novel β-lactamase bla from a clinical A. pittii isolate confers resistance to carbapenems only in vitro, demonstrating increased MICs and greater catalytic efficiency for carbapenems compared to bla. However, its activity against cephalosporins remains limited.
本研究调查临床分离株皮氏不动杆菌（A. pittii）PT-01中新型D类β-内酰胺酶OXA-1224的转移机制和功能特性。
使用基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）进行细菌鉴定，并通过聚合酶链反应（PCR）和DNA测序鉴定耐药基因。采用肉汤微量稀释法测定皮氏不动杆菌PT-01及其携带bla的重组转化体的抗菌药敏性。通过酶促反应动力学试验评估纯化的OXA-213和OXA-1224蛋白。
皮氏不动杆菌PT-01对阿米卡星、替加环素、多粘菌素B、头孢他啶、头孢吡肟、头孢哌酮-舒巴坦和哌拉西林-他唑巴坦敏感，但对亚胺培南和美罗培南耐药。该菌株携带bla和一个新型bla变体：bla。与对照菌株鲍曼不动杆菌（A. baumannii）ATCC 17978相比，表达bla的重组菌株对亚胺培南和美罗培南的最低抑菌浓度（MICs）分别增加了8倍和16倍。酶促试验表明，与OXA-213相比，OXA-1224对硝基头孢菌素的Km值更高，对亚胺培南（Km = 496.7）和美罗培南（Km = 126.1）的亲和力更强。与OXA-213相比，OXA-1224对亚胺培南和美罗培南的催化效率也有所提高。与OXA-213（Km/Kcat分别为2.469和2.547）相比，OXA-1224对亚胺培南（Km/Kcat = 3.362）和美罗培南（Km/Kcat = 4.837）的亲和力更高。然而，OXA-1224对第三代头孢菌素头孢他啶的活性极小（Km > 1000）。
临床分离的皮氏不动杆菌中的新型β-内酰胺酶bla仅在体外赋予对碳青霉烯类的耐药性，与bla相比，其对碳青霉烯类的MICs增加且催化效率更高。然而，其对头孢菌素的活性仍然有限。
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