Abstract

Carbapenem antibiotics are widely used for their broad antibacterial effects, but the emergence of carbapenem-resistant Enterobacterales has recently become a global problem. To solve this problem, research is needed to find compounds that increase antibiotic activity. Therefore, this study aimed to validate the antibacterial and anti-biofilm effects, as well as the inhibition of gene expression of thymoquinone, an extract of Nigella sativa commonly used as a spice in many dishes. The minimum inhibitory concentration of carbapenem antibiotics and thymoquinone was determined. Phenotypic analysis was performed to confirm the effect of thymoquinone on motility, which is one of the virulence factors of carbapenem-resistant uropathogenic Escherichia coli (CR-UPEC). Furthermore, quantitative real-time polymerase chain reaction analysis was used to determine the expression levels of carbapenemase gene (bla_KPC), efflux pump genes (acrA, acrB, acrD, tolC), as well as motility and adhesion genes (fliC, motA). In addition, biofilm inhibition and biofilm eradication assays were performed. All strains showed resistance to carbapenem antibiotics, while an antibacterial effect was confirmed at a concentration of 256 μg/mL of thymoquinone. Phenotypic analysis revealed a nearly 50% suppression in migration distance compared to the control group at 128 μg/mL of thymoquinone. Subsequent gene expression tests indicated the downregulation of carbapenemase-, efflux pump-, motility-, and adhesion genes by thymoquinone. Furthermore, our findings demonstrated that thymoquinone exhibits both biofilm formation inhibition and eradication effects. These findings suggest that thymoquinone may serve as a potential antibiotic adjuvant for treating CR-UPEC and could be a valuable resource in combating UTIs caused by multidrug-resistant bacteria.

Graphical Abstract

中文翻译：

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百里醌对耐碳青霉烯类尿路致病性大肠杆菌的抗菌和抗生物膜作用

摘要

碳青霉烯类抗生素因其广泛的抗菌作用而被广泛使用，但碳青霉烯类耐药肠杆菌的出现最近已成为一个全球性问题。为了解决这个问题，需要研究找到增加抗生素活性的化合物。因此，本研究旨在验证百里醌的抗菌和抗生物膜作用以及对基因表达的抑制作用，百里醌是黑种草的提取物，通常在许多菜肴中用作香料。测定了碳青霉烯类抗生素和百里醌的最低抑菌浓度。通过表型分析证实百里醌对运动性的影响，运动性是耐碳青霉烯类尿路致病性大肠杆菌(CR-UPEC) 的毒力因子之一。此外，采用实时定量聚合酶链式反应分析来测定碳青霉烯酶基因（bla _KPC）、外排泵基因（acrA、acrB、acrD、tolC）以及运动和粘附基因（fliC、motA）的表达水平。 。此外，还进行了生物膜抑制和生物膜根除测定。所有菌株均表现出对碳青霉烯类抗生素的耐药性，并且在浓度为 256 μg/mL 的百里醌时证实具有抗菌作用。表型分析显示，与对照组相比，在 128 μg/mL 百里醌浓度下，迁移距离受到近 50% 的抑制。随后的基因表达测试表明百里醌下调了碳青霉烯酶、外排泵、运动性和粘附基因。此外，我们的研究结果表明，百里醌具有生物膜形成抑制和根除作用。这些发现表明，百里醌可能作为治疗 CR-UPEC 的潜在抗生素佐剂，并且可能成为对抗多重耐药细菌引起的尿路感染的宝贵资源。

图形概要