Nakamura Akihiro, Nakamura Tatsuya, Niki Makoto, Kuchibiro Tomokazu, Nishi Isao, Komatsu Masaru
Department of Clinical Laboratory Science, Faculty of Health Care, Tenri Health Care University, Tenri, Japan.
Faculty of Health Sciences, Kyoto Tachibana University, Kyoto, Japan.
Front Microbiol. 2021 Aug 24;12:665432. doi: 10.3389/fmicb.2021.665432. eCollection 2021.
Worldwide spread of Enterobacteriaceae resistant to colistin, a polypeptide antibacterial drug for last-resort treatment of carbapenemase-producing Enterobacteriaceae (CPE) infections, is concerning. This study aimed to elucidate colistin MICs and molecular characteristics of to of ESBL-producing (ESBL-Ec) and CPE in Japan and clarify the genomic structure of strains harboring genes (especially ). This study included 168 ESBL-Ec and 126 CPE strains isolated at Japanese medical facilities. Colistin susceptibility testing and multiplex PCR targeting to were performed for all strains with S1-nuclease pulsed-field gel electrophoresis, Southern blot hybridization, and whole-genome sequencing (WGS) with hybrid assembly performed for gene-carrying strains. Two CPE strains showed a MIC ≥ 4 μg/ml in colistin susceptibility testing, with no known resistance mechanism detected. However, PCR conducted on all target strains detected three -carrying strains showing colistin susceptibility. The -positive THUN648 strain simultaneously carried and on a 275-kbp plasmid. Besides, + -positive THUN262 and -positive THUN627 had encoded on the chromosome. Only THUN627 encoded , which is suggested to be a regulatory gene for , downstream of . However, this strain showed no increased expression of these genes in mRNA quantitative analysis under colistin exposure. Colistin MICs of ESBL-Ec and CPE in Japan were all below 2 μg/ml, which is below the epidemiological cutoff (ECOFF) value (https://eucast.org/) or clinical breakpoint (CB) (CLSI M100-S30) reported for colistin, indicating neither "microbiological" nor "clinical" resistance. Several colistin-susceptible Enterobacteriaceae carrying silent encoded on plasmids and chromosomes have already spread worldwide along with other antimicrobial resistance genes. However, the mechanism of colistin resistance by remains unclear.
作为治疗产碳青霉烯酶肠杆菌科细菌（CPE）感染的最后手段的多肽抗菌药物，对黏菌素耐药的肠杆菌科细菌在全球范围内的传播令人担忧。本研究旨在阐明日本产超广谱β-内酰胺酶（ESBL）的大肠埃希菌（ESBL-Ec）和CPE对黏菌素的最低抑菌浓度（MIC）及分子特征，并阐明携带mcr基因（尤其是mcr-1）菌株的基因组结构。本研究纳入了在日本医疗机构分离出的168株ESBL-Ec和126株CPE菌株。对所有菌株进行了黏菌素敏感性试验以及针对mcr-1至mcr-5的多重聚合酶链反应（PCR），并采用S1核酸酶脉冲场凝胶电泳、Southern印迹杂交以及对携带mcr基因的菌株进行杂交组装的全基因组测序（WGS）。在黏菌素敏感性试验中，有两株CPE菌株的MIC≥4μg/ml，未检测到已知的耐药机制。然而，对所有目标菌株进行的PCR检测到三株携带mcr基因且对黏菌素敏感的菌株。mcr-1阳性的THUN648菌株在一个275kbp的质粒上同时携带mcr-1和mcr-2。此外，mcr-1+mcr-2阳性的THUN262和mcr-3阳性的THUN627在染色体上编码mcr。只有THUN627在mcr-3下游编码mcr-4，mcr-4被认为是mcr-3的调控基因。然而，在黏菌素暴露下的mRNA定量分析中，该菌株这些基因的表达并未增加。日本ESBL-Ec和CPE对黏菌素的MIC均低于2μg/ml，低于欧洲抗菌药物敏感性试验委员会（EUCAST）（https://eucast.org/）或美国临床和实验室标准协会（CLSI）M100-S30报告的黏菌素的流行病学临界值（ECOFF）或临床断点（CB），表明既无“微生物学”耐药也无“临床”耐药。几种携带沉默mcr基因的对黏菌素敏感的肠杆菌科细菌已与其他抗菌药物耐药基因一起在全球范围内传播。然而，mcr介导的黏菌素耐药机制仍不清楚。
