Supplementary file 1_Toward essential oil stewardship: strain-resolved evaluation of thyme oil activity against Pseudomonas aeruginosa.docx

The rising interest in essential oils (EOs) as antimicrobial agents demands evaluation frameworks that provide structured, reproducible assessments. In this study, we examined the strain-dependent response of Pseudomonas aeruginosa to pharmacopoeia-grade Thyme Essential Oil (obtained from Thymus vulgaris L., TEO) or polyhexamethylene biguanide antiseptic (PHMB) using a panel of ten genetically diversified strains in planktonic and biofilm forms, and by complementary in vitro models. Chemical composition of TEO was assessed using Gas Chromatography-Mass Spectrometry (GC–MS), and the main components were thymol, p-cymene, and γ-terpinene. Despite uniform test conditions, we observed striking inter-strain variability: TEO Minimal Inhibitory Concentrations (MICs) differed by up to 1,000-fold, and biofilm susceptibility profiles ranged from full tolerance to near-complete eradication. Notably, strains with low metabolic activity and sparse cell populations—but high matrix biomass—exhibited reduced responsiveness to TEO, while susceptibility to PHMB was more consistent, though not absolute. These findings highlight the critical influence of both microbial phenotype and agent formulation on antimicrobial outcomes. Rather than framing EOs as superior or inferior alternatives, our results advocate for their integration into a stewardship paradigm—one that values standardization, model-based evaluation, and informed formulation. In this context, we position essential oil stewardship not as a constraint but as a necessary evolution for their credible inclusion in antimicrobial strategies.

随着人们对作为抗菌剂的精油（Essential Oils，EOs）的兴趣日益高涨，亟需建立可提供结构化、可重复评估结果的评价体系。本研究采用10株遗传多样性各异的菌株，分别以浮游态和生物膜态形式，并结合互补体外模型，探究了铜绿假单胞菌（Pseudomonas aeruginosa）对药典级百里香精油（Thyme Essential Oil，TEO，提取自百里香<Thymus vulgaris L.>）或聚六亚甲基双胍消毒剂（polyhexamethylene biguanide antiseptic，PHMB）的菌株依赖性应答。本研究通过气相色谱-质谱联用法（Gas Chromatography-Mass Spectrometry，GC–MS）对TEO的化学成分进行了分析，其主要成分为麝香草酚、对伞花烃与γ-松油烯。尽管实验条件保持统一，我们仍观察到显著的菌株间差异：TEO的最低抑菌浓度（Minimal Inhibitory Concentrations，MICs）差异最高可达1000倍，且生物膜敏感性谱从完全耐受到近乎完全清除不等。值得注意的是，代谢活性较低、细胞数量稀少但基质生物量较高的菌株对TEO的响应性有所降低；而菌株对PHMB的敏感性则一致性更高，尽管并非完全一致。上述研究结果凸显了微生物表型与抗菌剂配方两者对抗菌效果的关键影响。本研究结果并未将精油视为优劣分明的替代抗菌剂，而是主张将其纳入抗菌管理范式——这一范式重视标准化、基于模型的评估与审慎的配方开发。在此背景下，我们认为精油管理并非限制其应用的桎梏，而是推动其可信地纳入抗菌策略的必要演进路径。