Development of Tigecycline Resistance in Carbapenemase-Producing Klebsiella pneumoniae Sequence Type 147 via AcrAB Overproduction Mediated by Replacement of the ramA Promoter

• Published in: Annals of laboratory medicine Vol. 40; no. 1; pp. 15 - 20
• Main Authors: Yoon, Eun Jeong, Oh, Yena, Jeong, Seok Hoon
• Format: Journal Article
• Language: English
• Published: Korea (South) The Korean Society for Laboratory Medicine 01.01.2020; 대한진단검사의학회
• Subjects: Anti-Bacterial Agents - pharmacology; Anti-Bacterial Agents - therapeutic use; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; beta-Lactamases - genetics; beta-Lactamases - metabolism; Drug Resistance, Bacterial - genetics; Genome, Bacterial; Humans; Klebsiella Infections - diagnosis; Klebsiella Infections - drug therapy; Klebsiella Infections - microbiology; Klebsiella pneumoniae - drug effects; Klebsiella pneumoniae - genetics; Klebsiella pneumoniae - isolation & purification; Membrane Transport Proteins - genetics; Membrane Transport Proteins - metabolism; Microbial Sensitivity Tests; Multilocus Sequence Typing; Original; Promoter Regions, Genetic; Tigecycline - pharmacology; Tigecycline - therapeutic use; Whole Genome Sequencing; 병리학; Resistance; AcrAB; Tigecycline; ramA; Klebsiella pneumoniae
• ISSN: 2234-3806; 2234-3814
• DOI: 10.3343/alm.2020.40.1.15

Carbapenem-resistant 2297, isolated from a patient treated with tigecycline for pneumonia, developed tigecycline resistance, in contrast to carbapenem-resistant isolate 1215, which was collected four months prior to the 2297 isolate. Mechanisms underlying tigecycline resistance were elucidated for the clinical isolates. The tigecycline minimum inhibitory concentration (MIC) was determined using the broth microdilution method, with or without phenylalanine-arginine β-naphthylamide (PABN), and whole-genome sequencing was carried out by single-molecule real-time sequencing. The expression levels of the genes , , , , and were determined by reverse-transcription quantitative PCR. Both isolates presented identical antibiograms, except for tigecycline, which showed an MIC of 0.5 mg/L in 1215 and 2 mg/L in 2297. The addition of PABN to tigecycline-resistant 2297 caused a four-fold decrease in the tigecycline MIC to 0.5 mg/L, although expression (encoding the AcrAB efflux pump) was upregulated by 2.5 fold and expression (encoding the pump activator RamA) was upregulated by 1.4 fold. We identified a 6,096-bp fragment insertion flanking direct TATAT repeats that disrupted the gene located upstream of in the chromosome of 2297; the insertion led the gene promoter replacement resulting in stronger activation of the gene. The isolate developed tigecycline resistance during tigecycline treatment. It was related to the overexpression of the AcrAB resistance-nodulation-cell division efflux system due to promoter replacement.