【摘要】 目的 确认D-甲硫氨酸（D-Met）对美罗培南（MEM）抗万古霉素耐药肠球菌（VRE）的增敏作用，并初步探索其增敏机制。方法 通过肉汤微稀释法、棋盘法、杀菌曲线法检测D-Met对MEM抗VRE的体外增敏活性；通过大蜡螟幼虫感染模型、小鼠腿部感染模型和小鼠腿部口袋感染模型，以组织匀浆细菌计数和生存率为指标评价D-Met对MEM抗VRE的体内增敏活性；通过Van-FL二肽尾结合实验、DltA酶促反应速率测定等初步探索其增敏机制。结果 肉汤微稀释法和棋盘法实验结果显示D-Met可增敏MEM抗VRE活性，两者联用的分数抑菌浓度指数<0.5，提示协同抗VRE作用。1μg/mL MEM和20 mmol/L D-Met联用抗VRE时在2 h后开始呈现杀菌作用，并且能持续到24 h。大蜡螟幼虫模型中，MEM和D-Met联用组的抗VRE活性优于单用MEM，而小鼠腿部感染和腿部口袋模型均未表现出明显联用作用。初步机制探索结果提示D-Met作为dltABCD通路的底物参与壁磷壁酸合成的作用弱，并且D-Met无取代粪肠球菌肽聚糖二肽尾D-Ala-D-Ala的作用。结论 D-Met在体外能显著增加MEM的抗VRE活性，并且在大蜡螟幼虫感染模型中得到验证。D-Met对于壁磷壁酸合成及肽聚糖二肽尾D-Ala-D-Ala的影响弱，其增敏机制有待进一步探索。更多还原

【Abstract】 Objective To confirm the sensitizing effect of D-methionine(D-Met) on meropenem(MEM) against vancomycin-resistant Enterococcus(VRE), and to preliminarily explore its sensitizing mechanism. Methods The in vitro sensitization activity of D-Met on MEM against VRE was detected using broth microdilution method, checkerboard assay and bactericidal curve measurement. The in vivo sensitization activity was assessed utilizing various models, including Galleria mellonella larvae infection, mouse thigh infection, and mouse thigh pocket infection, using indicators such as survival rate or tissue homogenate bacterial count. The preliminary sensitization mechanism was explored through a Van-FL D-Ala-D-Ala dipeptide tail binding assay and the determination of the DltA enzyme-catalyzed reaction rate. Results The results from the in vitro antibacterial activity assay and checkerboard assay indicated that D-Met enhances the anti-VRE activity of MEM, with fractional inhibitory concentration index(FICI) values below 0.5, suggesting synergistic anti-VRE effects. Specifically, the combination of 1 μg/mL MEM and 20 mmol/L D-Met exhibited a bactericidal effect against VRE at 2 h and sustained this effect for up to 24 h. In the Galleria mellonella larvae infection model, the combination of MEM and D-Met improved anti-VRE activity; however, the mouse thigh infection and thigh pocket infection models did not reveal significant synergistic effects of D-Met and MEM. Preliminary investigations into the mechanism suggest that D-Met acts weakly as a substrate for the dltABCD pathway involved in the synthesis of wall teichoic acid, and D-Met does not play a role in substituting peptidoglycan D-Ala-D-Ala dipeptide tail of E. faecalis. Conclusion D-Met significantly enhances the anti-VRE activity of MEM in vitro, and this sensitization effect has been validated in Galleria mellonella larvae infection model. However, D-Met exhibits weak effects in the synthesis of wall teichoic acid and in substituting the D-Ala-D-Ala dipeptide tail, indicating that further exploration of the sensitization mechanism of D-Met is warranted.更多还原