Department of Medical Microbiology, Ruhr-University Bochum, Universitätsstraße 150, 44801 Bochum, Germany
Institute for Medical Laboratory Diagnostics Bochum GmbH, Castroper Straße 45, 44791 Bochum, Germany.
J Antimicrob Chemother. 2015 Jul;70(7):1973-80. doi: 10.1093/jac/dkv079. Epub 2015 Apr 2.
The objective of this study was to characterize a novel IMP-type metallo-β-lactamase (MBL) found in an MDR clinical isolate of Pseudomonas aeruginosa.
The P. aeruginosa isolate NRZ-00156 was recovered from an inguinal swab from a patient hospitalized in Western Germany and showed high MICs of carbapenems. MBL production was analysed by Etest for MBLs, an EDTA combined disc test and an EDTA bioassay. Typing of the isolate was performed by MLST. Genetic characterization of the new blaIMP gene was performed by sequencing the PCR products. A phylogenetic tree was constructed. The novel blaIMP gene was expressed in Escherichia coli TOP10 and the enzyme was subjected to biochemical characterization.
The P. aeruginosa isolate NRZ-00156 expressed the ST235 allelic profile and was resistant to all the β-lactams tested except aztreonam. The isolate was positive for MBL production and harboured a new IMP allele, blaIMP-31, located on a disrupted class I integron [also carrying the blaOXA-35, aac(6')-Ib, aac(3)-Ic and aphA15 genes]. Its closest relative was IMP-35, with 96.7% amino acid identity. Expression of blaIMP-31 demonstrated that E. coli TOP10 producing IMP-31 had elevated resistance to all the β-lactams tested except aztreonam. Kinetic data were obtained for both IMP-31 and IMP-1. In comparison with IMP-1, IMP-31 showed weaker hydrolytic activity against all the β-lactams tested, which resulted from lower kcat values.
The characterization of the new IMP-type gene blaIMP-31 from an ST235 P. aeruginosa isolate indicates an ongoing spread of highly divergent IMP-type carbapenemases in clinical P. aeruginosa strains and highlights the continuous need for the prevention of nosocomial infections caused by MDR Gram-negative bacteria.
本研究旨在对一株耐多药铜绿假单胞菌临床分离株中发现的新型 IMP 型金属β-内酰胺酶（MBL）进行特征分析。
从德国西部住院患者腹股沟拭子中分离出铜绿假单胞菌 NRZ-00156 分离株，该分离株对碳青霉烯类药物的 MIC 较高。采用 MBLs Etest、EDTA 联合药敏纸片试验和 EDTA 生物测定法分析 MBL 的产生情况。采用 MLST 对分离株进行分型。通过测序 PCR 产物对新型 blaIMP 基因进行遗传特征分析。构建系统发育树。将新型 blaIMP 基因在大肠杆菌 TOP10 中表达，并对酶进行生化特性分析。
铜绿假单胞菌 NRZ-00156 分离株表达 ST235 等位基因谱，对除氨曲南外所有测试的β-内酰胺类药物均耐药。该分离株产 MBL 阳性，携带一种新型 IMP 等位基因 blaIMP-31，位于破坏的 I 类整合子上[还携带 blaOXA-35、aac(6')-Ib、aac(3)-Ic 和 aphA15 基因]。其最接近的亲缘关系是 IMP-35，氨基酸序列同源性为 96.7%。blaIMP-31 的表达表明，产 IMP-31 的大肠杆菌 TOP10 对除氨曲南外所有测试的β-内酰胺类药物均有较高的耐药性。获得了 IMP-31 和 IMP-1 的动力学数据。与 IMP-1 相比，IMP-31 对所有测试的β-内酰胺类药物的水解活性较弱，这是由于较低的 kcat 值所致。
从一株 ST235 铜绿假单胞菌分离株中鉴定出新型 IMP 型基因 blaIMP-31，表明具有高度差异的 IMP 型碳青霉烯酶在临床铜绿假单胞菌菌株中的传播不断，突显了持续需要预防耐多药革兰氏阴性菌引起的医院感染。
