β-内酰胺类抗生素、D-Ser、DAAO抑制剂三药联用抗MRSA作用

Anti-MRSA effect of theβ-lactams,D-Ser and DAAO inhibitor combinations

目的研究β-内酰胺类抗生素、D-Ser及D-氨基酸氧化酶(DAAO)抑制剂三药联用抗耐甲氧西林金黄色葡萄球菌(MRSA)感染的疗效。方法体外利用肉汤微稀释法和结晶紫染色法评价DAAO抑制剂5-氯苯并[d]异恶唑-3-醇(CBIO)对细菌生长和生物被膜的影响;建立小鼠全身感染模型和大鼠腿部感染模型,以生存率和组织匀浆细菌计数为指标评价三药联用疗效;小鼠皮下给药评价CBIO的安全性。结果在体外MIC测定时CBIO没有抑菌作用,但分别与苯唑西林和美罗培南联用时可对两种抗菌药物的抗菌活性产生明显的拮抗作用。D-Ser和CBIO单用对于MRSA N315所产生物被膜的抑制和清除作用较弱,当与抗生素联用时可显著提升生物被膜的抑制率和清除率;皮下单独给予健康小鼠30mg/kg CBIO后5d内,小鼠生存状况良好(n=5);三药(β-内酰胺类抗生素、D-Ser和CBIO)联用相比单用抗生素以及抗生素和D-Ser两药联用,可一定程度上提高小鼠存活率、降低大鼠腿部肌肉组织持菌量。结论β-内酰胺类抗生素、D-Ser及CBIO联用在体内具有良好的抗MRSA感染作用,且CBIO体内安全性良好,但CBIO会与抗生素产生一定程度的拮抗作用。一定浓度的CBIO与β-内酰胺类抗生素、D-Ser联用或许能为MRSA感染的临床用药提供新的选择。

Objective To study the activity ofβ-lactams,D-Ser and D-amino acid oxidase inhibitor combinations in treating infections caused by methicillin-resistant Staphylococcus aureus(MRSA).Methods The effect of D-amino acid oxidase inhibitor 5-chlorobenzo[d]isoxazol-3-ol(CBIO)on bacterial growth and biofilm was assessed in vitro by broth micro-dilution method and crystal violet staining method.The mouse systemic infection model and rat leg infection model were established,and the survival rates and tissue homogenate bacterial counts were used to evaluate the antibacterial effect of the three-drug combinations.The safety of CBIO in mice was evaluated by subcutaneous administration.Results In vitro MIC determination demonstrated that CBIO has no bacteriostatic effect,but it obviously antagonized the antibacterial activities of oxacillin and meropenem compounds when combined with them.D-Ser and CBIO alone have weak effects on biofilm formation and disassembly by MRSA N315.When combined with antibiotics,it can significantly increase the inhibition and clearance rates of biofilms.Within 5 days after subcutaneous administration of 30mg/kg CBIO to healthy mice,the mice survived well(n=5).Compared with single antibiotic administrations and antibiotic-D-Ser combinations,the three-drug(β-lactams,D-Ser and CBIO)combinations can improve the survival rate of mice and reduce the bacterial counts in the rat legs to some extent.Conclusion Theβ-lactams,D-Ser and CBIO combinations have good anti-MRSA infection effect in vivo,and CBIO has good safety in mice,but CBIO will have antagonistic effect to some extent with antibiotics.A certain concentration of CBIO combined withβ-lactam antibiotics and D-Ser may provide a new choice for the clinical drugs of MRSA infections.