Analysis of the effects of chlorhexidine on oral biofilm vitality and structure based on viability profiling and an indicator of membrane integrity
Analysis of the effects of chlorhexidine on oral biofilm vitality and structure based on viability profiling and an indicator of membrane integrity
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DOI：
10.1128/aac.48.5.1461-1468.2004
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发表时间：
2004-05-01
影响因子：
4.9
通讯作者：
Wilson, M
中科院分区：
文献类型：
--
作者：
Hope, CK;Wilson, M
在恒定深度的膜发酵罐中的羟基磷灰石基质上生长多物种生物膜建模邻间牙菌斑，然后浸入含有膜完整性的荧光指示剂的观察溶液中。共聚焦激光扫描显微镜（CLSM）揭示了生物膜内细胞活力的结构和空间分布。将葡萄糖酸氯己定（CHX）加入观察溶液中，以达到0.05和0.2%（重量/体积）的浓度，然后捕获进一步的CLSM延时系列。图像分析显示，暴露于0.2%CHX导致生物膜沿着z轴以1.176 μ m/min（-1）的速率收缩，并且在3至5分钟的延迟后，还影响总荧光测量和通过生物膜的生存力分布的变化。生物膜的总荧光测量在5分钟后显示出几乎检测不到的变化。然而，（荧光相对于时间相对于深度）清楚地表明存在时间依赖性效应，但是稀释CHX随时间的效应的最清楚的指示剂是活力分析。这些发现表明，有可能使用膜完整性的荧光指示剂与活力分析相结合，来评估膜活性抗菌化合物的杀菌作用对生物膜的渗透。
Multispecies biofilms modeling interproximal plaque were grown on a hydroxyapatite substratum in a constant-depth film fermentor and then immersed in a viewing solution containing fluorescent indicators of membrane integrity. Confocall laser scanning microscopy (CLSM) revealed the structure and spatial distribution of cell vitality within the biofilms. Chlorhexidine gluconate (CHX) was added to the viewing solution to achieve concentrations of 0.05 and 0.2% (wt/vol) before further CLSM time-lapse series were captured. Image analysis showed that exposure to 0.2% CHX caused the biofilm to contract at a rate of 1.176 mum min(-1) along the z axis and also effected changes in total fluorescence measurements and viability profiles through the biofilms after a delay of 3 to 5 min. At a concentration of 0.05% CHX, total fluorescence measurements for the biofilm exhibited barely detectable changes after 5 min. Fluorescence profiles (fluorescence versus time versus depth), however, clearly showed that a time-dependent effect was present, but the clearest indicator of the effect of dilute CHX over time was viability profiling. These findings suggest the possibility of using fluorescent indicators of membrane integrity in conjunction with viability profiling to evaluate the penetration of the bactericidal effects of membrane-active antimicrobial compounds into biofilm.