Table_1_Characterization of IncHI1B Plasmids Encoding Efflux Pump TmexCD2-ToprJ2 in Carbapenem-Resistant Klebsiella variicola, Klebsiella quasipneumoniae, and Klebsiella michiganensis Strains.DOCX

Tigecycline serves as one of the last-resort antibiotics to treat severe infections caused by carbapenem-resistant Enterobacterales. Recently, a novel plasmid-mediated resistance-nodulation-division (RND)-type efflux pump gene cluster, TmexCD1-ToprJ1, and its variants, TmexCD2-ToprJ2 and TmexCD3-ToprJ3, encoding tetracyclines and tigecycline resistance, were revealed. In this study, we reported three TmexCD2-ToprJ2-harboring Klebsiella species strains, collected from two teaching tertiary hospitals in China, including one K. quasipneumoniae, one K. variicola, and one K. michiganensis. The three strains were characterized by antimicrobial susceptibility testing (AST), conjugation assay, WGS, and bioinformatics analysis. AST showed that K. variicola and K. quasipneumoniae strains were resistant to tigecycline with MIC values of 4μg/ml, whereas the K. michiganensis was susceptible to tigecycline with an MIC value of 1μg/ml. The TmexCD2-ToprJ2 clusters were located on three similar IncHI1B plasmids, of which two co-harbored the metallo-β-lactamase gene blaNDM-1. Conjugation experiments showed that all three plasmids were capable of self-transfer via conjugation. Our results showed, for the first time, that this novel plasmid-mediated tigecycline resistance mechanism TmexCD2-ToprJ2 has spread into different Klebsiella species, and clinical susceptibility testing may fail to detect. The co-occurrence of blaNDM-1 and TmexCD2-ToprJ2 in the same plasmid is of particular public health concern as the convergence of “mosaic” plasmids can confer both tigecycline and carbapenem resistance. Its further spread into other clinical high-risk Klebsiella clones will likely exacerbate the antimicrobial resistance crisis. A close monitoring of the dissemination of TmexCD-ToprJ encoding resistance should be considered.

替加环素作为治疗由碳青霉烯类抗生素耐药肠杆菌科细菌引起的严重感染的最后手段抗生素之一。近期，一种新型的质粒介导的抗性-聚集-分裂（RND）型外排泵基因簇，TmexCD1-ToprJ1及其变体TmexCD2-ToprJ2和TmexCD3-ToprJ3，编码四环素和替加环素耐药性，已被揭示。在本研究中，我们报道了三个携带TmexCD2-ToprJ2的克雷伯菌属菌株，这些菌株分别来自中国两所教学三级医院，包括一株K. quasipneumoniae、一株K. variicola和一株K. michiganensis。这三个菌株通过抗菌敏感性测试（AST）、接合试验、全基因组测序（WGS）和生物信息学分析进行了表征。AST结果显示，K. variicola和K. quasipneumoniae菌株对替加环素具有耐药性，最小抑菌浓度（MIC）值为4μg/ml，而K. michiganensis对替加环素敏感，MIC值为1μg/ml。TmexCD2-ToprJ2基因簇位于三个相似的IncHI1B质粒上，其中两个质粒共携带金属β-内酰胺酶基因blaNDM-1。接合实验表明，所有三个质粒均可通过接合实现自我转移。我们的研究首次表明，这种新型的质粒介导的替加环素耐药机制TmexCD2-ToprJ2已扩散至不同的克雷伯菌属，而临床敏感性测试可能无法检测到。blaNDM-1与TmexCD2-ToprJ2在同一质粒中的共现，尤其是“镶嵌”型质粒的汇聚可以赋予替加环素和碳青霉烯类抗生素的耐药性，对公共卫生构成了特别的关注。这种耐药基因向其他临床高风险克雷伯菌克隆的进一步扩散，可能会加剧抗菌药物耐药性危机。因此，对TmexCD-ToprJ编码的耐药基因的传播进行密切监测应予以考虑。