壳聚糖及其季铵盐对铜绿假单胞菌生物膜形成的影响

检验医学 ›› 2012, Vol. 27 ›› Issue (10): 854-857.

壳聚糖及其季铵盐对铜绿假单胞菌生物膜形成的影响

林萍¹,赵雨璠²,³,吴悦²,³,张骁²,³,郭晓奎⁴,谭红略⁵,李擎天²,³,孙阳⁶

1. 上海交通大学医学院附属上海市精神卫生中心检验科，上海 200030；2. 上海交通大学医学院附属瑞金医院检验系，上海200025；3. 上海交通大学医学院病原生物学教研室，上海 200025；4. 上海交通大学医学院附属上海市第九人民医院，上海 200011；5.上海市同济医院医学科学研究所，上海 200065

收稿日期:2012-09-04 修回日期:2012-09-14 出版日期:2012-10-30 发布日期:2012-10-12
通讯作者: 孙阳，联系电话：021-66111431。
作者简介:林萍，女，1968年生，硕士，主任技师，主要从事临床医学检验工作。
基金资助:
国家自然科学基金（81000708）；“十一五”国家科技重大专项（2009ZX10004-712）

The influence of chitosan and quaternized chitosan on Pseudomonas aeruginosa biofilm formation

1. Departemtn of Clinical Laboratory，Shanghai Mental Health Center，Shanghai Jiaotong University School of Medicine，Shanghai 200030，China； 2. Faculty of Medical Laboratory Science，Ruijin Hospital，Shanghai Jiaotong University School of Medicine，Shanghai 200025，China； 3. Laboratory of Pathogenic Biology，Shanghai Jiaotong University School of Medicine，Shanghai 200025，China； 4. Shanghai Nineth People's Hospital，Shanghai Jiaotong University School of Medicine，Shanghai 200011，China；5. Laboratory of Medical Science Institute，Shanghai Tongji Hospital，Shanghai 200065，China

Received:2012-09-04 Revised:2012-09-14 Online:2012-10-30 Published:2012-10-12
Contact: ping LIN

摘要/Abstract

摘要： 目的　探讨壳聚糖(CHI)及其衍生物可能具有的广谱抗菌作用。方法　采用CHI、壳聚糖季铵盐(QCHI)进行铜绿假单胞菌体外抑菌试验。采用CHI和QCHI分别作用0、3、6、9、12 h，每个时间点取1 mL菌液进行菌落计数并与对照孔比较，同时抽提铜绿假单胞菌RNA，扩增其ahpF、pa3187、aprA和rpoS基因，并检测各基因表达水平；剩余铜绿假单胞菌用LB培养基培养3 d，PBS缓冲液冲洗浮游细菌，结晶紫染色，95%乙醇脱色后用酶标仪570 nm检测并比较铜绿假单胞菌生物膜的形成能力。结果　与对照孔比较，自6 h起CHI和QCHI孔的存活菌量即有明显差异（P＜0.05）；但CHI孔和QCHI孔之间差异无统计学意义。CHI和QCHI均可使铜绿假单胞菌pa3187基因表达上调，aprA基因表达下调；QCHI可使ahpF基因表达下调，而CHI可使ahpF基因表达上调；QCHI对rpoS基因的表达有下调趋势。从6 h起，CHI和QCHI对铜绿假单胞菌生物膜的形成均有明显的抑制作用。结论　CHI及QCHI能在一定程度上抑制铜绿假单胞菌的增殖及其生物膜的形成。

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关键词: 壳聚糖, 壳聚糖季铵盐, 铜绿假单胞菌, 抑菌试验, 生物膜形成

Abstract: Objective　To investigate the broad-spectrum antibacterial efficacy of chitosan (CHI) and its derivatives. Methods　The antibacterial test of Pseudomonas aeruginosa was performed by CHI and quaternized chitosan (QCHI). The volumes of per 1 mL at 0，3，6，9 and 12 h by CHI and QCHI were determined. The results were compared with those of control hole volume. The Pseudomonas aeruginosa RNA was extracted simultaneously. The ahpF，pa3187，aprA and rpoS genes were amplified，and their expressions were determined. The remaining Pseudomonas aeruginosa were cultured by LB medium for 3 d. The floating bacteria were flushed by PBS buffer solution，and were stained by crystallization violet. After decolorizing by 95% ethanol，the biofilm formation function of Pseudomonas aeruginosa was analyzed by enzyme-labeled analyzer at 570 nm. Results　There were significant differences between CHI and QCHI hole volumes of live bacteria after culturing 6 h compared with control hole volumes，but there was no obvious difference CHI and QCHI volumes. The pa3187 expression was increased by CHI and QCHI，but aprA expression was decreased by them. QCHI can decrease ahpF expression，but CHI can improve ahpF expression. The rpoS expression was also decreased by QCHI. Biofilm formation showed that the significant inhibitory effect could be seen in Pseudomonas aeruginosa biofilm formation by CHI and QCHI from culturing 6 h. Conclusions　In a certain extent，CHI and QCHI can inhibit the proliferation and biofilm formation of Pseudomonas aeruginosa.

Key words: Chitosan, Quaternized chitosan, Pseudomonas aeruginosa, Antibacterial test, Biofilm formation

林萍1，赵雨璠2，吴悦2，张骁2，郭晓奎3，谭红略4，李擎天2，孙阳5. 壳聚糖及其季铵盐对铜绿假单胞菌生物膜形成的影响[J]. 检验医学, 2012, 27(10): 854-857.

LIN Ping 1，ZHAO Yufan 2，WU Yue 2，ZHANG Xiao 2，GUO Xiaokui 3，TAN Honglue 4，LI Qingtian 2，SUN Yang 5. The influence of chitosan and quaternized chitosan on Pseudomonas aeruginosa biofilm formation[J]. , 2012, 27(10): 854-857.

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