和厚朴酚抑制耐甲氧西林金黄色葡萄球菌生物被膜形成

Inhibition of methicillin-resistant Staphylococcus aureus biofilm by honokiol

摘要:

[目的] 研究和厚朴酚（HNK）抑制MRSA生物被膜（BF）形成的作用机制。[方法] 使用TTC法测定了HNK对供试菌株BF的形成和成熟BF的抑制作用；刚果红平板法定性检测了HNK对PIA合成的影响；分光光度法测定了HNK对供试菌株eDNA释放量的影响；RT-PCR技术检测了HNK对供试菌株icaA、cidA以及agrA基因表达量的影响。[结果] HNK对MRSA 41573 BF的形成和成熟BF均有较强的抑制作用，其中，HNK抑制MRSA 41573 BF形成的MIC和MBC分别为10 μg/mL和20 μg/mL；抑制成熟BF的MIC和MBC分别为50 μg/mL和100 μg/mL。当用亚抑菌浓度的HNK与万古霉素联合作用后，可显著提高成熟BF对万古霉素的敏感性。HNK能显著抑制PIA的合成，且呈浓度剂量依赖。HNK能抑制供试菌株eDNA的释放量，其中1/8 MIC的HNK作用供试菌株16 h后，与对照组相比，eDNA的释放量降低了28.3%。HNK可抑制供试菌株BF形成的相关基因，其中1/2 MIC的HNK作用供试菌株16 h后，与对照相比，icaA的表达量降低了59.1%，cidA的表达量降低了56%，agrA的表达量降低了72.3%。[结论] HNK能显著抑制MRSA 41573 BF的形成，其作用机制主要是通过抑制icaA和cidA基因表达量，影响PIA和eDNA的合成，进而抑制BF的形成。此外HNK也可通过调控细菌的QS系统影响BF的形成。

Abstract:

[Objective] To study the inhibition of methicillin-resistant Staphylococcus aureas (MRSA) biofilm by honokiol.[Methods] We used triphenyl tetrazolium chloride method to evaluate the inhibition of biofilm formation and mature by honokiol. We used congo red agar and spectrophotometer to detect the influence of honokiol on polysaccharide intercellular adhesion formation and extracellular DNA release. RT-PCR analysis was used to determine the effect of honokiol on expression of icaA, cidA and agrA.[Results] Honokiol showed strong antimicrobial activity both on biofilm formation and mature biofilm of MRSA 41573. Minimum inhibitory concentration was 10 μg/mL for biofilm formation and 50 μg/mL for mature biofilm. Minimum bactericidal concentration was 20 μg/mL for biofilm formation and 100 μg/mL for mature biofilm. Honokiol showed synergy effect with vancomycin and it significantly increased the sensitivity of mature biofilm to vancomycin. Polysaccharide intercellular adhesion formation and extracellular DNA release were effectively inhibited by honokiol. Extracellular DNA release decreased by 28.3% when honokiol at 1/8 MIC. After incubated with 1/2 MIC of honokiol for 16 h, the relative expression of icaA, cidA and agrA of MRSA41573 was reduced by 59.1%, 56% and 72.3%, respectively.[Conclusion] Honokiol can significantly inhibit biofilm formation of MRSA41573 and its mechanism is mainly the inhibited expression of icaA and cidA to influence the synthesis of polysaccharide intercellular adhesion and extracellular DNA. Moreover, it also affect biofilm formation by QS system.