摘要
      水平基因转移（HGT）在抗生素抗性基因（ARGs）的传播中起着重要作用。大多数报道的化合物通过增加细胞膜通透性来促进HGT。据报道，粘菌素在发挥其抗菌作用时会增加细胞膜的通透性。因此，本研究旨在通过建立体外交配模型，研究粘菌素在通过质粒偶联促进ARGs传播中的潜在作用。将携带质粒RP4-7的大肠杆菌DH5α、大肠杆菌L65和大肠杆菌LD67-1、blaNDM-5阳性IncX3质粒和mcr-1阳性IncI2质粒分别作为供体菌株，将大肠杆菌J53作为受体菌株。暴露于亚抑制浓度的粘菌素（1/4、1/8、1/16 MIC）显著刺激了RP-4质粒、宽型IncI2和IncX3质粒的结合频率。扫描电子显微镜显示粘菌素处理后细胞膜收缩，而碘化丙啶染料和1-N-苯基萘胺荧光探针显示细胞膜通透性增加。此外，外膜蛋白（ompF和ompC）的表达水平增加。这些结果表明膜屏障发生了断裂。交配对形成基因（trbBp）的表达得到促进，下调trbBp表达的全局调节基因（korA，trbA）的表达受到抑制。因此，在粘菌素暴露后，交配配对机的产量可以提高。这些发现有助于理解粘菌素在抗微生物耐药性传播中的潜在风险。
ABSTRACT
Horizontal gene transfer (HGT) plays a significant role in the spread of antibiotic resistance genes (ARGs). Most reported compounds promote HGT by increasing the cell membrane permeability. Colistin has been reported to increase the cell membrane permeability when exhibiting its antibacterial effect. Therefore, this study aimed to investigate the potential role of colistin in facilitating the dissemination of ARGs via plasmid conjugation by establishing an in vitro mating model. Three strains Escherichia coli (E. coli) DH5α, E. coli L65, and E. coli LD67-1 carrying plasmid RP4-7, blaNDM-5 positive IncX3 plasmid, and mcr-1 positive IncI2 plasmid, respectively, were regarded as the donor strains and E. coli J53 as the recipient strain. Exposure to subinhibitory concentrations of colistin (1/4, 1/8, 1/16 MIC) significantly stimulated the conjugation frequency of RP-4 plasmid, wide-type IncI2 and IncX3 plasmid. Scanning electron microscopy revealed the shrunken cell membrane after colistin treatment, whereas propidium iodide dye and 1-N-Phenylnaphthylamine fluorescent probe showed the increased cell membrane permeability. Additionally, the expression level of the outer membrane proteins (ompF and ompC) was increased. These results indicate a break in the membrane barrier. The expression of the mating pair formation gene (trbBp) was promoted and the expression of the global regulatory genes (korA, trbA), which downregulates trbBp expression, was inhibited. Thus, the production of the mating pairing machine could be elevated after colistin exposure. These findings aid in understanding the hidden risks of colistin on the spread of antimicrobial resistance.

https://journals.asm.org/doi/full/10.1128/spectrum.02160-21