Emerging Antibiotic Resistance Unit, Medical and Molecular Microbiology, Faculty of Science and Medicine, University of Fribourggrid.8534.a, Fribourg, Switzerland.
Swiss National Reference Center for Emerging Antibiotic Resistance (NARA), University of Fribourggrid.8534.a, Fribourg, Switzerland.
Antimicrob Agents Chemother. 2021 Aug 17;65(9):e0089021. doi: 10.1128/AAC.00890-21.
Carbapenem-resistant , such as Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae, represent a major threat to public health due to their rapid spread. Novel drug combinations such as ceftazidime-avibactam (CZA), combining a broad-spectrum cephalosporin along with a broad-spectrum β-lactamase inhibitor, have recently been introduced and have been shown to exhibit excellent activity toward multidrug-resistant KPC-producing strains. However, CZA-resistant K. pneumoniae isolates are now being increasingly reported, mostly corresponding to producers of KPC variants. In this study, we evaluated the nature of the mutations in the KPC-2 and KPC-3 β-lactamase sequences (the most frequent KPC-type enzymes) that lead to CZA resistance and the subsequent effects of these mutations on susceptibility to other β-lactam antibiotics. Single-step selection assays were conducted, resulting in the identification of a series of mutations in the KPC sequence which conferred the ability of those mutated enzymes to confer resistance to CZA. Hence, 16 KPC-2 variants and 10 KPC-3 variants were obtained. Production of the KPC variants in an Escherichia coli recombinant strain resulted in a concomitant increased susceptibility to broad-spectrum cephalosporins and carbapenems, with the exceptions of ceftazidime and piperacillin-tazobactam, compared to wild-type KPC enzymes. Enzymatic assays showed that all of the KPC variants identified exhibited an increased affinity toward ceftazidime and a slightly decreased sensitivity to avibactam, sustaining their impact on CZA resistance. However, their respective carbapenemase activities were concurrently negatively impacted.
碳青霉烯类耐药菌，如产生 碳青霉烯酶的肺炎克雷伯菌（KPC），由于其快速传播，对公共健康构成重大威胁。新型药物组合，如头孢他啶-阿维巴坦（CZA），结合了一种广谱头孢菌素和一种广谱β-内酰胺酶抑制剂，最近已被引入，并已被证明对多药耐药的 KPC 产生菌具有极好的活性。然而，现在越来越多地报道了对 CZA 耐药的肺炎克雷伯菌分离株，这些分离株主要对应于 KPC 变体的产生者。在这项研究中，我们评估了导致 CZA 耐药的 KPC-2 和 KPC-3 β-内酰胺酶序列（最常见的 KPC 型酶）突变的性质，以及这些突变对其他β-内酰胺类抗生素敏感性的后续影响。进行了一步选择试验，导致 KPC 序列中的一系列突变被鉴定出来，这些突变使那些突变酶能够对 CZA 产生耐药性。因此，获得了 16 种 KPC-2 变体和 10 种 KPC-3 变体。在大肠杆菌重组菌株中产生 KPC 变体导致对广谱头孢菌素和碳青霉烯类药物的敏感性同时增加，除了头孢他啶和哌拉西林他唑巴坦，与野生型 KPC 酶相比。酶学分析表明，所有鉴定出的 KPC 变体对头孢他啶的亲和力均增加，对阿维巴坦的敏感性略有降低，维持了它们对 CZA 耐药性的影响。然而，它们各自的碳青霉烯酶活性同时受到负面影响。
