Department of Diagnostics and Public Health, University of Verona, Verona, Italy.
Department of Diagnostics and Public Health, University of Verona, Verona, Italy.
Clin Microbiol Infect. 2020 Jul;26(7):946.e1-946.e3. doi: 10.1016/j.cmi.2020.02.007. Epub 2020 Feb 12.
Carbapenemase-producing strains of Klebsiella pneumoniae (KPC) are a great health concern, and therapy with ceftazidime-avibactam represents a choice for the treatment of infections involving these strains. We report a strain resistant to ceftazidime-avibactam due to a deletion of six nucleotides in the bla gene sequence.
Two strains, namely AMP920 and AMP2009, were isolated from the same patient a month apart. Antimicrobial susceptibility testing was performed both by broth microdilution and by Etest. Immunoenzymatic assay to detect carbapenemase was performed for both strains. The bla gene of both strains was amplified by PCR and sequenced. Enzyme activity towards carbapenems was tested by the CarbaNP test and hydrolysis spectrophotometer assay.
The two isolates differed in antimicrobial susceptibility. AMP920 showed meropenem and imipenem resistance (MIC 32 and 32 mg/mL). A month later the carbapenem MIC decreased to 8 and 1 mg/mL respectively, while the ceftazidime-avibactam MIC increased from 1 to 16 mg/mL. Both isolates showed a positive immunoenzymatic test for the KPC enzyme, but only AMP920 showed a positive CarbaNP test hydrolysing imipenem. The Bla gene was amplified in both strains. After sequencing, the two amplicons showed a KPC3 variant. The gene of the second isolate showed a deletion of six nucleotides at 498-503, resulting in a mutant variant with the deletion of glutamic acid and leucine residues at positions 167 and 168.
We detected a new deletion in the bla gene of a clinical strain of K. pneumoniae which resulted in resistance to ceftazidime-avibactam. The amino acids deleted are in the Ω loop (amino acids 165-179) of the KPC enzyme, enhancing ceftazidime affinity and preventing avibactam binding.
产碳青霉烯酶肺炎克雷伯菌（KPC）是一个严重的健康问题，头孢他啶-阿维巴坦的治疗为感染这些菌株的治疗提供了一种选择。我们报告了一株因 bla 基因序列中 6 个核苷酸缺失而对头孢他啶-阿维巴坦耐药的菌株。
两个月内从同一患者中分离出两株菌，分别命名为 AMP920 和 AMP2009。采用肉汤微量稀释法和 Etest 进行药敏试验。对两株菌均进行碳青霉烯酶免疫酶检测。采用 PCR 扩增和测序方法扩增两株菌的 bla 基因。采用 CarbaNP 试验和水解分光光度计法检测酶对碳青霉烯类的水解活性。
两株分离株的药敏结果不同。AMP920 表现出美罗培南和亚胺培南耐药（MIC 为 32 和 32 mg/mL）。一个月后，碳青霉烯类药物的 MIC 分别降至 8 和 1 mg/mL，而头孢他啶-阿维巴坦的 MIC 从 1 增加到 16 mg/mL。两株分离株的 KPC 酶免疫酶试验均为阳性，但只有 AMP920 的 CarbaNP 试验水解亚胺培南为阳性。两株菌均扩增出 bla 基因。测序后，两个扩增子均显示出 KPC3 变体。第二株菌的基因在 498-503 位缺失了 6 个核苷酸，导致第 167 和 168 位的谷氨酸和亮氨酸残基缺失的突变体变体。
我们在一株临床分离的肺炎克雷伯菌 bla 基因中检测到一个新的缺失，导致其对头孢他啶-阿维巴坦耐药。缺失的氨基酸位于 KPC 酶的 Ω 环（氨基酸 165-179），增强了头孢他啶的亲和力，阻止了阿维巴坦的结合。
