Department of Bioengineering, Marmara University, 34722, Kadikoy, Istanbul, Turkey.
Department of Bioengineering, Uskudar University, 34662, Uskudar, Istanbul, Turkey.
Microb Pathog. 2020 Dec;149:104542. doi: 10.1016/j.micpath.2020.104542. Epub 2020 Sep 30.
A plethora of natural products emerges as attractive molecules in the struggle against antibiotic resistance. These molecules impose their bioactivities not only alone but also in combinations as well, which further enhances their effects. Berberine is a well-known isoquinoline alkaloid with antibacterial activity. Unfortunately, it is readily extruded, which significantly reduces its efficacy and restricts its potential. Thymol is a monoterpenic phenol that exhibits different biological activities but its major effect is observed only at relatively high concentrations, which raises concern on cytotoxicity. The aim of the study was to potentiate the antibacterial activity of berberine, in a combination treatment with thymol in the opportunistic pathogen Staphylococcus aureus and understand the antibacterial mechanism of the combination treatment. The synergism of berberine and thymol was first established by the checkerboard assay. Then the antibacterial mechanism of the synergistic combination was explored by growth curves, biofilm formation assay, SEM observation, and RNA-Seq based transcriptomic profiling. Checkerboard assay showed that 32 μg mL berberine and 64 μg mL thymol was a synergistic combination, both concentrations below their cytotoxicity limits for many cells. 32 μg mL berberine and 32 μg mL thymol was sufficient to inhibit biofilm formation. SEM images confirmed the morphological changes on the structure of combination treated cells. The major finding of the combination treatment from the transcriptomic analysis was the repression in the expression of virulence factors or genes related to virulence factors. Apart from the particular changes related to the cell envelope, the majority of expressional changes seemed to be similar to berberine-treated cells or to be resulting from general stress conditions. The findings of this work showed that when thymol was used in combination with berberine, it enhanced the antibacterial activity of berberine in a synergistic manner. Furthermore, thymol could be considered as an antivirulence agent, disarming S. aureus cells.
大量的天然产物在与抗生素耐药性的斗争中成为有吸引力的分子。这些分子不仅单独发挥其生物活性，而且还以组合形式发挥作用，从而进一步增强其效果。小檗碱是一种具有抗菌活性的已知异喹啉生物碱。不幸的是，它很容易被排出，这大大降低了它的功效，并限制了它的潜力。百里酚是一种单萜酚，具有不同的生物学活性，但只有在相对较高的浓度下才会观察到其主要作用，这引起了人们对细胞毒性的关注。本研究旨在增强小檗碱的抗菌活性，在机会性病原体金黄色葡萄球菌中与百里酚联合治疗，并了解联合治疗的抗菌机制。首先通过棋盘试验确定小檗碱和百里酚的协同作用。然后通过生长曲线、生物膜形成试验、SEM 观察和基于 RNA-Seq 的转录组分析探索协同组合的抗菌机制。棋盘试验表明，32μg mL 的小檗碱和 64μg mL 的百里酚是协同组合，两种浓度均低于其对许多细胞的细胞毒性限值。32μg mL 的小檗碱和 32μg mL 的百里酚足以抑制生物膜的形成。SEM 图像证实了组合处理细胞结构的形态变化。转录组分析的联合治疗的主要发现是抑制了毒力因子或与毒力因子相关的基因的表达。除了与细胞包膜有关的特定变化外，大多数表达变化似乎与小檗碱处理的细胞相似，或者是由一般应激条件引起的。这项工作的结果表明，当百里酚与小檗碱联合使用时，它以协同的方式增强了小檗碱的抗菌活性。此外，百里酚可以被认为是一种抗毒力剂，使金黄色葡萄球菌细胞失去毒力。
