通过外排泵抑制结合苯丙氨酸-精氨酸β-萘胺诱导的外膜透化增强阿奇霉素对多重耐药大肠杆菌的抗菌活性,International Journal of Molecular Sciences

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通过外排泵抑制结合苯丙氨酸-精氨酸β-萘胺诱导的外膜透化增强阿奇霉素对多重耐药大肠杆菌的抗菌活性
International Journal of Molecular Sciences ( IF 4.9 ) Pub Date : 2023-05-12 , DOI: 10.3390/ijms24108662
Farah Al-Marzooq ₁ , Akela Ghazawi ₁ , Lana Daoud ₁ , Saeed Tariq ₂

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多重耐药 (MDR) 细菌的全球传播增加了对发现新抗生素和佐剂的需求。苯丙氨酸-精氨酸 β-萘胺 (PAβN) 是革兰氏阴性菌外排泵的抑制剂，例如大肠杆菌中的 AcrAB-TolC 复合物。我们旨在探讨 PAβN 联合阿奇霉素 (AZT) 对一组耐多药大肠杆菌菌株的协同作用和作用机制。对 56 株菌株进行了抗生素敏感性检测，并筛选出大环内酯类耐药基因。然后，使用棋盘分析法测试了 29 个菌株的协同作用。PAβN 在表达 mphA 基因和编码大环内酯磷酸转移酶的菌株中以剂量依赖性方式显着增强 AZT 活性，但在携带 ermB 基因和编码大环内酯甲基化酶的菌株中没有。在具有 mcr-1 基因的粘菌素抗性菌株中观察到早期细菌杀伤（6 小时），导致脂质重塑，从而导致外膜（OM）渗透性缺陷。在暴露于高剂量 PAβN 的细菌中，透射电子显微镜显示出明显的 OM 损伤。荧光测定法也证实了 OM 渗透性的增加，证实了 PAβN 对 OM 的作用。PAβN 在没有透化 OM 的情况下以低剂量保持其作为外排泵抑制剂的活性。在单独使用 PAβN 或使用 AZT 处理的细胞中，acrA、acrB 和 tolC 表达因长时间暴露于 PAβN 而出现非显着增加，这反映了细菌试图抵消泵抑制。因此，发现 PAβN 可通过剂量依赖性作用有效增强 AZT 对大肠杆菌的抗菌活性。这需要进一步研究其与其他抗生素联合使用对多种革兰氏阴性细菌的影响。协同组合将有助于对抗 MDR 病原体，为现有药物库增加新工具。

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Boosting the Antibacterial Activity of Azithromycin on Multidrug-Resistant Escherichia coli by Efflux Pump Inhibition Coupled with Outer Membrane Permeabilization Induced by Phenylalanine-Arginine β-Naphthylamide

The global spread of multidrug-resistant (MDR) bacteria increases the demand for the discovery of new antibiotics and adjuvants. Phenylalanine-arginine β-naphthylamide (PAβN) is an inhibitor of efflux pumps in Gram-negative bacteria, such as the AcrAB-TolC complex in Escherichia coli. We aimed to explore the synergistic effect and mechanism of action of PAβN combined with azithromycin (AZT) on a group of MDR E. coli strains. Antibiotic susceptibility was tested for 56 strains, which were screened for macrolide resistance genes. Then, 29 strains were tested for synergy using the checkerboard assay. PAβN significantly enhanced AZT activity in a dose-dependent manner in strains expressing the mphA gene and encoding macrolide phosphotransferase, but not in strains carrying the ermB gene and encoding macrolide methylase. Early bacterial killing (6 h) was observed in a colistin-resistant strain with the mcr-1 gene, leading to lipid remodeling, which caused outer membrane (OM) permeability defects. Clear OM damage was revealed by transmission electron microscopy in bacteria exposed to high doses of PAβN. Increased OM permeability was also proven by fluorometric assays, confirming the action of PAβN on OM. PAβN maintained its activity as an efflux pump inhibitor at low doses without permeabilizing OM. A non-significant increase in acrA, acrB, and tolC expression in response to prolonged exposure to PAβN was noted in cells treated with PAβN alone or with AZT, as a reflection of bacterial attempts to counteract pump inhibition. Thus, PAβN was found to be effective in potentiating the antibacterial activity of AZT on E. coli through dose-dependent action. This warrants further investigations of its effect combined with other antibiotics on multiple Gram-negative bacterial species. Synergetic combinations will help in the battle against MDR pathogens, adding new tools to the arsenal of existing medications.

更新日期：2023-05-12

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