变形链球菌表面蛋白和葡糖基转移酶基因疫苗免疫防龋动物实验:体质量影响研究

目的 :了解变形链球菌基因疫苗单独及联合免疫对定菌鼠体质量的影响。方法 :2 8d龄Wistar大鼠 3 6只 ,随机分为pcDNA3 pac组、pcDNA3 gtfB组、pcDNA3 pac联合pcDNA3 gtfB组、变形链球菌灭活全菌阳性对照组、pcDNA3空载体阴性对照组和PBS液空白对照组 ,进行 3次双侧颌下腺腺周注射免疫 ,建立定菌鼠模型及诱龋实验 3个月 ,于实验开始和结束时测定体质量。结果 :基因疫苗免疫组体质量与 pcD NA3及PBS组无显著差异 (P >0 .0 5) ,而灭活全菌细胞免疫组体质量显著下降 (P <0 .0 1)。结论 :pcD NA3 gtfB和pcDNA3 pac对大鼠体质量无不良影响 ,较灭活全菌疫苗安全     

The inability of Streptococcus mutans and Lactobacillus acidophilus to form a biofilm in vitro on dentine pretreated with ozone

Background: The use of ozone therapy in the treatment of dental caries is equivocal. The aim of this study was to use an in vitro model to determine the effects of prior ozone application to dentine on biofilm formation and to measure any associated reduction in bacteria viability. Methods: Twenty dentine discs were bonded to the bases of 5 mL polycarbonate screw top vials. Ten dentine discs were infused with ozone for 40 seconds, 10 samples remained untreated as a control. The vials were filled with nutrient medium, sterilized and placed into the outflow from a continuous chemostat culture of Streptococcus mutans and Lactobacillus acidophilus for four weeks. At the conclusion of the experiment bacterial growth was monitored by taking optical density readings of the growth medium in each vial and the outer surface of the dentine specimens were examined by scanning electron microscopy as shown by SEM analysis. Results: Ozone infusion prevented biofilm formation on all the treated samples while there was substantial biofilm present on the control specimens. While the average optical density of the control specimens was almost twice that of the ozone infused dentine (0.710 for the control with a SD of 0.288 and 0.446 for the ozonated samples with a SD of 0.371), the results were not significant (p > 0.05). Conclusions: This preliminary study has shown that the infusion of ozone into non‐carious dentine prevented biofilm formation in vitro from S. mutans and L. acidophilus over a four‐week period. The possibility exists that ozone treatment may alter the surface wettability of dentine through reaction with organic constituents.     

Effects of Nidus Vespae extract and chemical fractions on glucosyltransferases, adherence and biofilm formation of Streptococcus mutans

Nidus Vespae (the honeycomb of Polistes olivaceous, P. japonicus Saussure and Parapolybiavaria fabricius) have been extensively used in traditional Chinese medicine, given their multiple pharmacological activities, including antimicrobial, anti-inflammatory, anti-virus, anti-tumor and anesthetic properties. The present study evaluated the anti-glucosyltransferases (GTFs) activity, anti-adherence and anti-biofilm properties of 95% ethanol/water extract, cyclohexane/ethyl acetate, petroleum ether/ethyl acetate and chloroform/methanol fractions of Nidus Vespae. Chloroform/methanol fraction showed a remarkable capacity for inhibiting the adherence of Streptococcus mutans ATCC 25175 to saliva-coated hydroxyapatite disc (S-HA) at sub-MC concentrations. In addition, the Nidus Vespae extract and chemical fractions significantly inhibited the activity of cell-associated and extracellular GTFs at sub-MIC concentrations, and the chloroform/methanol fraction was the most effective one. For the anti-biofilm activity assays, minimum biofilm inhibition concentrations (MBIC50) and minimum biofilm reduction concentrations (MBRC50) were determined using the microdilution method. The chloroform/methanol fraction showed the highest anti-biofilm activities with a MBIC50 of 8mg/ml and a MBRC(50) of 16mg/ml against Streptococcus mutans ATCC 25175. The significant inhibition of GTFs activity and biofilm formation demonstrated by Nidus Vespae shows it to be a promising natural product for the prevention of dental caries.     

Initial biofilm formation of Streptococcus sobrinus on various orthodontics appliances

Biofilms accumulate on hard and soft surface in the oral cavity. Accumulation of biofilms on orthodontic appliance bear scientific and clinical interest. The objection of this study was to examine the formation of dental biofilm by Streptococcus sobrinus on different types of orthodontics appliances, using a model consisting of host and bacterial constituents. The adsorption pattern of saliva to the orthodontics appliances was determined by means of gel electrophoresis coupled with computerized densitometry techniques. The amount of salivary proteins adsorbed onto the surfaces was measured using the Bradford method. Sucrose-dependent bacterial adhesion to the saliva-coated orthodontics appliances was tested by radioactive-labelled S. sobrinus. Our results show different adsorption patterns of salivary proteins to the various orthodontic appliances as modules, brackets, springs and intra oral elastics. Modules and brackets demonstrated the most affinity to salivary proteins. A surface dependent adhesion profile was recorded, showing a high affinity of albumin and amylase to modules. Bacterial accumulation was the highest on modules compared with springs which demonstrated the least bacterial adhesion. Our study demonstrates the specificity of biofilm formation on the different orthodontic appliances. Formation of a variety of dental biofilms has a significant impact on the progression of dental diseases associated with orthodontic treatment.     

壳寡聚糖对变形链球菌表面疏水性的影响

目的:研究壳寡聚糖对变形链球菌细胞表面疏水性的影响,从而进一步了解壳寡聚糖影响细菌黏附的机制。方法:使用不同浓度(10-2、10-1、1g/L)的壳寡聚糖溶液处理变形链球菌,并采用微生物黏着碳氢化合物法测试细胞表面疏水性。结果:随壳寡聚糖浓度增加,变形链球菌ATCC25175细胞表面疏水性有显著性降低。结论:壳寡聚糖可能通过降低细菌表面疏水作用而抑制变形链球菌的黏附。     

变形链球菌耐氟菌株与亲代菌株质子移位膜ATP酶活性的比较

目的:通过测定并比较变形链球菌耐氟菌株及其亲代菌株体内质子移位膜ATP酶的活性,以阐明耐氟菌株耐酸能力提高的原因。方法:将变链菌株Ingbritt及其耐氟突变株Ingbritt-FR通过甲苯处理,2个循环的液氮冷冻和37℃解冻,制成透性细胞。将透性细胞悬液加到含有10mmol/LMgSO4的50mmol/LTris-maleate测试缓冲液中(pH6.0),加温至37℃,再加入5mmol/LATP(pH6.0)起动反应。分别于10、20、60min取样,测定样品中水解ATP所释出的无机磷量。采用磷钼酸比色法在紫外分光光度计上进行比色分析(660nm),所得数据采用双因素方差分析。结果:在10、20和60min时,耐氟菌株H+-ATP酶活性分别为308.48、136.67和82.80μmolPi/g细胞干重/min,显著高于亲代菌株的相应酶活性:104.77,、64.69和30.70(P<0.01)。随时间推移,两类菌株的H+-ATP酶活性均逐渐降低,在酶的作用时间差别上有统计学意义(P<0.01)。结论:耐氟菌株ATP酶活性增高为其耐酸性增高的原因;H+-ATP酶活性及耐酸性的增高将会增加变链菌耐氟菌株的致龋潜能。     

壳寡聚糖对变形链球菌黏附的影响

目的:研究壳寡聚糖对变形链球菌蔗糖依赖性黏附的影响。方法:使用不同浓度的壳寡聚糖与变形链球菌S.mutansATCC25175共同培养,计算壳寡聚糖对变形链球菌的黏附抑制率的影响,同时扫描电镜下观察变形链球菌在玻片的形态学特征。结果:变形链球菌对玻片的黏附率随培养基中壳寡聚糖浓度的升高而下降。扫描电镜图片显示壳寡聚糖组细菌间基质少,菌细胞清晰。而空白组细菌则包裹有较多的胶冻状基质。结论:壳寡聚糖可以有效抑制变形链球菌的蔗糖依赖性黏附。     

口腔链球菌对白假丝酵母菌的拮抗作用

目的 观测3种口腔链球菌对白假丝酵母菌的生长有无抑制作用.方法 采用直接点种法、反点种菌落计数法和液体共培养法观测口腔变异链球菌、血链球菌、唾液链球菌对白假丝酵母菌的拮抗作用.结果 1)直接点种法中各组均未观察到清晰的抑菌圈;2)唾液链球菌软琼脂上的白假丝酵母菌菌落数较对照组减少(P<0.05);3)活菌与白假丝酵母菌...     

二价双启动子防龋DNA疫苗的构建及鉴定

目的：构建并鉴定具有在真、原核细胞中均可表达目的抗原并具备强免疫原性、针对致龋菌变形链球菌2种主要毒力因子的DNA疫苗.方法：利用PCR技术从载有变形链球菌gtfB基因的质粒中扩增葡聚糖结合区段（GBR）基因片断;将合成的组织纤溶酶原信号肽序列（tPA-SP）连接到GBR基因的上游;再将此基因（sGBR）定向克隆到载有sSBR基因的双启动子真核表达载体pCN-SSIE上,构建出pCN-SSISG;对重组质粒pCN-SSISG进行酶切图谱分析和DNA序列测定.结果：酶切图谱分析显示,PCR扩增的及插入到pCN-SSIE中的目的基因sGBR片断大小与预期相符;DNA序列测定结果确定：重组质粒pCN-SSISG开放性阅读框架和插入的信号肽序列均正确.结论：成功构建了具有在真核、原核宿主细胞均可表达的具有双启动子的质粒pCN-SSISG.