Beijing Key Laboratory of Antimicrobial Agents, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
National Institutes for Food and Drug Control, Beijing, China.
Microbiol Spectr. 2022 Oct 26;10(5):e0174822. doi: 10.1128/spectrum.01748-22. Epub 2022 Aug 18.
To study the effect of different tolerable levels of constitutive expression on Escherichia coli, and to provide direct evidence for moderate resistance mediated by , construction of E. coli strains carrying on the chromosome with promoters of different strengths was conducted using λ-red recombination. Our results demonstrated that over-high expression of cannot be tolerated, and seven constructs with more than 200-fold transcriptional expression differences were obtained. The colistin MICs of the seven strains increased with the increase of MCR-1 levels, and the highest MIC was 8 μg/mL. Lower expression of didn't demonstrate many effects on bacteria, while higher tolerable expression of tended to show fitness costs in growth rate, competitive ability, and cell structures, but no obvious change of virulence was observed in mice. Bacteria demonstrated colistin MICs of 4-8 μg/mL at expression levels similar to clinical isolates, which were the expression levels with relatively lower fitness costs. The effects of relatively lower tolerable levels of were not evaluated thoroughly, and direct evidence for moderate resistance mediated by was lacking. In the present study, we made constructs carrying on the E. coli K12 chromosome under the control of serial constitutive promoters of different strengths and studied the effects of different tolerable levels of expression and . The results demonstrated that generally, except QH0007 (the construct with the highest expression that showed some extent of cell death), the fitness costs of tolerable expression on bacteria were not apparent or low. Bacteria demonstrated colistin MICs of 4-8 μg/mL at expression levels similar to clinical isolates, which corresponded to the lower levels of expression that can lead to colistin resistance, indicating the cleverness of bacteria to balance the benefit and cost of MCR-1-mediated colistin resistance.
为了研究不同可耐受水平的组成型表达对大肠杆菌的影响，并为中效耐药提供直接证据，利用 λ-red 重组技术构建了携带不同强度启动子的染色体上 mcr-1 的大肠杆菌菌株。结果表明，过高的 mcr-1 表达水平不能被耐受，获得了 200 倍以上转录表达差异的 7 个构建体。7 株菌的黏菌素 MIC 随 mcr-1 水平的增加而增加，最高 MIC 为 8 μg/ml。mcr-1 低表达对细菌没有太多影响，而 mcr-1 高耐受表达则倾向于表现出生长速度、竞争能力和细胞结构的适应性代价，但在小鼠中没有观察到明显的毒力变化。细菌在与临床分离株相似的表达水平（相对较低的适应性代价水平）下表现出 4-8 μg/ml 的黏菌素 MIC，这一表达水平介导的中度耐药。相对较低耐受水平的 mcr-1 的作用没有被充分评估，也缺乏 mcr-1 介导的中度耐药的直接证据。在本研究中，我们构建了携带 mcr-1 的大肠杆菌 K12 染色体，并在不同强度的连续组成型启动子的控制下进行了研究，研究了不同可耐受水平的 mcr-1 表达和功能的影响。结果表明，除了 QH0007（表达最高的构建体显示出一定程度的细胞死亡）外，mcr-1 表达对细菌的适应性代价通常不明显或较低。细菌在与临床分离株相似的表达水平（与导致黏菌素耐药的较低 mcr-1 表达水平相对应）下表现出 4-8 μg/ml 的黏菌素 MIC，这表明细菌在 mcr-1 介导的黏菌素耐药中巧妙地平衡了效益和成本。
