In vitro attachment of oral microorganisms to solid surfaces: Evaluation of a controlled flow method

Abstract – A flow cell method was modified to provide shear-controlled experimental conditions for monitoring initial attachment and detachment of oral microorganisms, to solid surfaces. Whole unstimulated human saliva was collected and circulated at a flow rate of 1 ml/min, through a cell composed of two parallel test plates. Infrared-transparent plates of medium surface energy served as test substrata in these initial calibration experiments. The plates presented a similar distribution of polar forces and dispersion forces at the surface as that of human tooth enamel and some restorative dental materials. Internal reflection infrared spectroscopy verified the presence of deposited organic material. After saliva had been circulated through the flow cells for 15 min at 37 ° C, sterilized distilled water was introduced at the same flow rate and time of exposure to remove unattached microorganisms. Morphologic characterizations and counts of adherent Gram-stainable microorganisms were performed using incident light microscopy. Three different surface zones corresponding to the inlet area, the middle area and the outlet area of the flow cell were analyzed, and compared with enumerations of microorganisms in the whole saliva samples. Numbers of attached microorganisms in the three zones followed predictions from the laminar flow conditions, with a positive correlation shown between total numbers of microorganisms in saliva and total numbers of microorganisms attached. Cocci and rods were the only morphotypes observed on the plates, and no significant difference could be detected between the percentage of cocci and rods attached in the three different zones. The results showed high deposition efficiency from the zone immediately adjacent to the test surfaces. No apparent morphological selectivity was induced. This flow cell method has numerous advantages for monitoring the influence on cellular adhesion of varying liquid phase and solid surface combinations, under dynamically controlled experimental conditions. It is particularly useful for small sample volumes and short exposure times.     

Effect of critical surface tension on retention of oral microorganisms

The effect of critical surface tension on the initial retention of microorganisms from unstimulated human saliva was tested in a flow cell system. Prior to each experiment the total numbers and the morphotypes of microorganisms present in saliva were recorded. The test surfaces were prepared to display known increasing critical surface tensions, as verified and standardized by contact angle measurements. Surfaces of initially low (20-22 mN/m), medium (35-38 mN/m) and high (greater than 50 mN/m) critical surface tension were exposed to saliva at a flow rate of 1 ml/min. Microbiota and biofilm material associated with the test surface after 15 min of salivary exposure, were then subjected to standard detachment forces, by introducing a cell-free rinsing fluid at two different shear rates. Both the attachment and the detachment phases were executed at room temperature or 37 degrees C. The retained population was counted in three different zones of the test surfaces with a light microscope and statistically tested for correlation to the main variables (critical surface tension, flow rate and temperature) and interactions. Retention success was significantly dependent on the initial critical surface tension and the flow rate. Surfaces of medium critical surface tension, representative of human tooth surfaces and most restorative dental materials, retained the highest numbers of microorganisms in comparison with the other surfaces tested, with no statistically verified selectivity in proportions of retained coccoid and rodshaped microorganisms for any surface. A 30-fold increase of the flow rate resulted in a 70-80% reduction of the retention success, with a higher relative number of cocci present on all the test surfaces. These results demonstrate that initial retention of microorganisms to surfaces is non-specific with regard to morphotypes, but is strongly related both to the mechanical removal forces and the surface energetic state of the solid surface exposed. Retention of microbial populations at interfaces might, therefore, be controllable by advance selection of the critical surface tensions and predicted if shear forces at given sites are known.     

口腔混合唾液与下颌总义齿固位力

目的：探讨口腔混合唾液黏度和分泌量对下颌总义齿固位力的影响。方法：对31例患进行唾液黏度，分泌量及下颌总义齿固位力的测试。结果：切变率的改变可明显影响唾液黏度（P＜0．01），不同唾液黏度的两组间和不同唾液分泌量的两组间，下颌总义齿固位力略有差异，但差异均没有显性（P＞0．05）。结论：口腔混合唾液黏度和分泌量是影响下颌总义齿固位力的因素之一，但不是决定性因素。     

表面粗糙度对水与唾液在纯钛及PMMA上润湿作用的影响

目的:评价全口义齿基托材料及其表面粗糙度对义齿固位的影响。方法:测定水与唾液在表面粗糙度不同的纯钛和聚甲基丙烯酸甲酯(PMMA)上的前进接触角和后退接触角,通过两者的差值θh反映水与唾液对它们的润湿效果。结果:1、随着表面粗糙度增加,唾液在纯钛上的θh逐渐降低,但当Ra≥1.0,Rz≥1.5时,θh突然增大;而PMMA组,当Ra<1.2,Rz<2时,粗糙度对θh无明显影响,当Ra≥1.2,Rz≥2时θh也显著增加,多数情况下,唾液在相同粗糙度的纯钛上的θh大于在PMMA上的θh。2、水在光滑的PMMA和纯钛上的前进接触角比唾液的大。结论:唾液对纯钛的润湿效果,可以通过高度磨光或形成较为粗糙的表面(Ra≥1.0,Rz≥1.5)得到提高,多数情况下这种润湿效果好于PMMA。提示从润湿效果看,对于不能高度研磨和抛光的义齿组织面,全口义齿上颌腭托选择纯钛并形成较为粗糙的表面可能更有利于固位。     

牙体充填材料表面粗糙度对常见口腔链球菌黏附力的影响

目的 采用原子力显微镜（AFM）检测常见口腔链球菌属与不同表面粗糙度的光固化复合树脂及玻璃离子水门汀（GIC）之间的黏附力。方法 将光固化复合树脂和GIC样本表面梯度抛光,根据最终表面粗糙度不同分为300、200、100和10 nm组,使用AFM观察其表面形貌。采用先锋菌（血链球菌、缓症链球菌）和致龋菌（变异链球菌、表兄链球菌）制作细菌改性探针,通过AFM获得力-距曲线测量细菌与树脂和GIC样本表面的黏附力。对材料表面粗糙度测量值进行方差分析,细菌黏附力进行Kruskal-Wallis非参数检验,同时采用Dunn’s进行组间两两比较,并对表面粗糙度与细菌黏附力进行相关性分析。结果 随材料表面粗糙度增加,细菌的黏附力增大,4种细菌的黏附力均在300 nm的材料表面达到最大值;在10和300 nm组的GIC表面,变异链球菌的黏附力由0.578 nN增加到2.876 nN。4种细菌在树脂表面的黏附力略大于GIC,先锋菌的黏附力略大于致龋菌,组间差异均在200和300 nm组时较明显。结论 材料表面粗糙度对细菌黏附力的影响较大,二者有明显的相关性;GIC对细菌的黏附性较复合树脂低;材料表面粗糙度对致龋菌的影响小于先锋菌。