替加环素联合亚胺培南对黏液型铜绿假单胞菌的体外抗菌活性及生物膜抑制作用分析

doi:10.3969/j.issn.1673-8640.2020.06.002

摘要/Abstract

摘要：

目的评价替加环素联合亚胺培南对黏液型铜绿假单胞菌（MPA）体外浮游菌和生物膜的抑制作用及联合抗菌效果。方法采用微量肉汤稀释法测定不同浓度组合的替加环素和亚胺培南对89株MPA的最小抑菌浓度（MIC）,采用棋盘法测定替加环素与亚胺培南的体外协同作用,并计算分级抑菌浓度指数（FICI）,采用时间杀菌曲线判定2种药物的联合效应,采用96孔板结合结晶紫半定量染色法判定生物膜清除效应。结果替加环素联合亚胺培南对MPA临床分离株浮游菌的协同抑制作用为46.07%（41株）,部分协同作用为14.6%（13株）,相加作用为12.36%（11株）,无关作用为26.97%（24株）,无拮抗作用。培养24 h,亚胺培南、替加环素联用与单独使用比较,菌量抑制>2 log₁₀CFU/mL。与单独使用1/4 MIC替加环素比较,加入1/4、1/2、1 MIC亚胺培南后,可明显降低生物膜黏附性（P<0.05）;与单独使用1/4 MIC亚胺培南比较,加入1/4、1/2、1 MIC替加环素后,可明显降低生物膜黏附性（P<0.05）。结论替加环素联合亚胺培南具有协同或部分协同抑制MPA的作用,联合用药比单独用药具有更高的抗菌效能,同时对于生物膜抑制作用具有一定的协同效应。

关键词: 黏液型铜绿假单胞菌, 生物膜, 时间杀菌曲线, 替加环素, 亚胺培南, 协同作用

Abstract:

Objective To evaluate the inhibitory and biofilm inhibitory effect of imipenem combined with tigecycline on mucoid Pseudomonas aeruginosa（MPA） in vitro. Methods The minimum inhibitory concentration（MIC） of different concentrations of tigecycline and imipenem against 89 MPA isolates were determined by micro-broth dilution method. The checkerboard method was used to determine and classify the fractional inhibitory concentration index（FICI）,the time-killing curve was used to determine the combined effect,and the 96-well plate combined with crystal violet semi-quantitative staining was used to determine the biofilm inhibitory effect. Results The synergistic effect of tigecycline combined with imipenem on MPA was 46.07%（41 isolates）,partial synergistic effect was 14.6%（13 isolates）,and additive effect was 12.36%（11 isolates）,irrelevant effect was 26.97%（24 isolates）,and there was no antagonistic effect. At culturing for 24 h,the combined therapy showed >2 log₁₀ CFU/mL reduction in viable bacteria compared with tigecycline or imipenem alone. Compared with the use of tigecycline（1/4 MIC）,the combination of tigecycline with imipenem（1/4,1/2,1 MIC） reduced the counts of viable bacteria on each biofilm（P<0.05）. Compared with the use of imipenem（1/4 MIC）,the combination of imipenem with tigecycline（1/4,1/2,1 MIC） reduced the counts of viable bacteria on each biofilm（P<0.05）. Conclusions The combination of tigecycline with imipenem showed synergistic or partial synergistic effects on MPA. Combined therapy has higher antibacterial efficiency than that alone,and it has a certain enhancement effect on the inhibitory of biofilm.

Key words: Mucoid Pseudomonas aeruginosa, Biofilm, Time-killing curve, Tigecycline, Imipenem, Synergistic effect

中图分类号:

R378.99

钟峰, 冯艺, 张晶晶, 许岩, 方咏梅, 陶秀彬, 吴竞. 替加环素联合亚胺培南对黏液型铜绿假单胞菌的体外抗菌活性及生物膜抑制作用分析[J]. 检验医学, 2020, 35(6): 519-523.

ZHONG Feng, FENG Yi, ZHANG Jingjing, XU Yan, FANG Yongmei, TAO Xiubin, WU Jing. In vitro antimicrobial activity and biofilm inhibitory effect of imipenem combined with tigecycline against mucoid Pseudomonas aeruginosa[J]. Laboratory Medicine, 2020, 35(6): 519-523.

图/表 4

图1 替加环素与亚胺培南单用及联用对MPA的MIC CIR曲线注： ● 亚胺培南; ▲ 替加环素; ■ 亚胺培南+替加环素

表1 替加环素与亚胺培南单用及联用对MPA的MIC μg/mL

抗菌药物	单用		联用
抗菌药物	MIC₅₀	MIC₉₀	MIC₅₀	MIC₉₀
亚胺培南	1	2	0.125^*	0.25^*
替加环素	0.25	0.5	0.062 5^*	0.125^*

图2 替加环素（1/2 MIC）与亚胺培南（1/2 MIC）单用及联用的时间杀菌曲线注：亚胺培南的MIC[（a）、（f）为0.5 μg/mL,（b）、（c）、（d）、（e）为0.25 μg/mL];替加环素的MIC[（a）、（b）、（c）、（f）为0.125 μg/mL,（e）为0.25 μg/mL,（d）为0.062 5 μg/mL]; ■ 对照; ● 亚胺培南; ▲ 替加环素; ▲ 联合用药

表2 替加环素联合亚胺培南对MPA生物膜的作用效应（A570nm值） $\bar{x}±s$

亚胺培南MIC	替加环素MIC
亚胺培南MIC	0	1/4	1/2	1
0	0.261±0.006	0.214±0.005^*	0.211±0.009^*	0.136±0.004^*
1/4	0.248±0.005	0.189±0.008^*#	0.175±0.006^*#	0.129±0.01^*
1/2	0.227±0.004	0.164±0.005^*#	0.136±0.001^*#	0.125±0.005^*
1	0.128±0.007^#	0.13±0.003^#	0.126±0.006^#	0.126±0.004

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