完善根管充填后治疗失败感染根管中粪肠球菌的活菌研究

目的:将叠氮溴化丙锭(PMA)与定量PCR(q PCR)技术结合,用于计算根管治疗失败感染根管中粪肠球菌活菌数量。方法:选取34名根管治疗失败患者,提取感染根管内细菌DNA,每个样本分成2份,一份经PMA处理(实验组),一份未用PMA处理(对照组),然后通过q PCR方法检测样本中粪肠球菌的活菌数量和细菌总量。结果:20例患者(58.8%)细菌样本中检测出粪肠球菌。实验组和对照组Ct值分别为25.12±2.04和24.62±2.02(P=0.001)。结论:对于根管治疗失败感染根管内活菌的研究使用PMA结合定量PCR可能是一种有效的方法。     

纳米银对牙本质表面粪肠球菌生物膜的杀菌作用

目的 研究纳米银对牙本质表面粪肠球菌生物膜的杀菌作用,为探寻新型有效的根管消毒药物提供实验依据.方法 制备牙本质片309个,随机分成3组,在牙本质表面形成粪肠球菌生物膜后,分别用0.1%纳米银溶液、1.313%次氯酸钠溶液和0.9%氯化钠溶液处理.3组分别于处理0(即处理前)、1、6、12、24 h时取出样本,应用活菌...     

三种无机纳米抗菌剂对粪肠球菌的抗菌作用

目的:研究纳米银、载银纳米二氧化钛、掺氮纳米二氧化钛对粪肠球菌的抗菌作用。方法:将粪肠球菌在MBECTMP&G Assay中培养24 h,构建粪肠球菌感染模型,扫描电镜观察MBECTMP&G Assay上粪肠球菌感染情况。应用MBECTMP&G Assay以连续稀释法检测纳米银、载银纳米二氧化钛、掺氮纳米二氧化钛的最小抑菌浓度(MIC)和最小生物膜清除浓度(MBEC)。结果:扫描电镜下见粪肠球菌在MBECTM P&G Assay上形成生物膜。纳米银对粪肠球菌的MIC和MBEC分别为62.5 mg/L和1 000 mg/L,载银纳米二氧化钛MIC为64 g/L,但在所用浓度范围内均不能清除生物膜细菌,即未测得MBEC;掺氮纳米二氧化钛的MIC、MBEC均未测得;次氯酸钠的MIC和MBEC分别为6.56 g/L和13.13 g/L。结论:纳米银对粪肠球菌有较强抗菌作用,能清除粪肠球菌生物膜;粪肠球菌对载银纳米二氧化钛和掺氮纳米二氧化钛有较强抵抗力。     

茶多酚、MTAD和次氯酸钠液抑制粪肠球菌及其生物膜的作用比较

目的:检测中药茶多酚、MTAD、52.5 g/L次氯酸钠抑制根管壁粪肠球菌生物膜的能力,并分析中药茶多酚作为根管冲洗液的可行性。方法:采用微孔板滴定法检测茶多酚,MTAD和52.5 g/L次氯酸钠液对粪肠球菌的最小抑菌浓度(MIC)和最小杀菌浓度(MBC),采用琼脂扩散实验检测3种抗菌剂对粪肠球菌的抑菌圈。通过建立粪肠球菌感染根管3周和6周的模型,分别用茶多酚、MTAD和52.5 g/L次氯酸钠液对粪肠球菌感染后的根管进行处理,然后对根管壁生物膜中的细菌生存情况进行定性定量分析,比较这3种根管冲洗液抗粪肠球菌生物膜的能力。结果:MIC、MBC和琼脂扩散实验显示:3组冲洗剂对粪肠球菌均有抑制作用。在粪肠球菌感染根管3周的生物膜中,MTAD和次氯酸钠液可以完全抑制粪肠球菌生物膜,茶多酚处理后虽仍有粪肠球菌生长,但明显低于生理盐水处理组的细菌生存数,差异有统计学意义(P<0.05)。在粪肠球菌感染根管6周的生物膜中,次氯酸钠液可以完全抑制细菌生长,茶多酚和MTAD作用后仍有活菌生长,但与对照组相比,细菌生存数均显示了8个log值的下降,差异有统计学意义(P<0.05)。结论:52.5 g/L次氯酸钠液具有很强的抑制根管内粪肠球菌生物膜作用,而茶多酚和MTAD也具有一定的抑制粪肠球菌生物膜的能力。     

胞外多糖对粪肠球菌单菌种生物膜形成的影响

目的:观察粪肠球菌单菌种生物膜的动态形成过程,探讨胞外多糖在粪肠球菌生物膜形成过程中的分布和作用。方法:粪肠球菌在盖玻片上形成单菌种生物膜,采用荧光技术标记胞外多糖和细菌,荧光显微镜观察生物膜结构和胞外多糖的分布;蒽酮法测定粪肠球菌在游离和粘附状态下产生水溶性胞外多糖(Water soluble exopolysaccharides,WSE)和水不溶性胞外多糖(Water insoluble exopolysaccharides,WIE)的能力变化。结果:粪肠球菌能在玻片表面粘附形成生物膜,生物膜中胞外多糖分布与菌落中的细菌分布一致,相互包裹成熟形成网络状结构。不同培养时间、不同生存状态粪肠球菌合成WSE和WIE的能力不同,差异有显著性(P<0.05)。粘附菌产生WSE的能力往往低于浮游菌,但是粘附菌产生WIE均高于浮游菌,差异具有统计学意义(P<0.05)。结论:胞外多糖在粪肠球菌生物膜形成过程中发挥重要作用。     

纳米银干预牙本质表面粪肠球菌粘附的实验研究

目的:研究纳米银对牙本质表面粪肠球菌粘附的干预作用。方法:制备牙本质片42个,随机均分成3组,分别置于0.1%纳米银溶液、1.313%次氯酸钠溶液和生理盐水内浸泡30min后,于200μL粪肠球菌悬液中培养1h,用扫描电镜观察牙本质表面形态和粪肠球菌粘附情况,荧光显微镜分析牙本质表面粘附的粪肠球菌数量并拍照记录,Image pro plus 6.0软件计算粪肠球菌总菌数、活菌数和活菌百分比。结果:0.1%纳米银溶液组和生理盐水组牙本质表面形态清晰无破坏,1.313%次氯酸钠溶液组牙本质溶融坍塌,胶原溶解破坏。牙本质经0.1%纳米银溶液处理后,其表面粘附的粪肠球菌总菌数、活菌数和活菌百分比均低于1.313%次氯酸钠溶液组和生理盐水组(P﹤0.01)。结论:0.1%纳米银溶液可有效减少粪肠球菌对牙本质的粘附,对牙本质表面形态无明显破坏。     

纳米银颗粒联合氢氧化钙对饥饿期粪肠球菌生物膜的作用

目的:将氢氧化钙和纳米银联合,探讨其对饥饿期粪肠球菌生物膜的抑制效果.方法:使用256个牙根样本建立饥饿期粪肠球菌生物膜体外模型,使用平板菌落计数法和结晶紫染色法测定不同药物[Ca(OH)2+AgNP,Ca(OH)2,AgNPy不同作用时间(1、7d)对饥饿期粪肠球菌生物膜的抑制效果.以灭菌水作为实验对照组.采用SPSS13.0软件包对数据进行统计学分析.结果:在1d和7d时,Ca (OH)2+AgNP对粪肠球菌生物膜的抑制效果强于Ca (OH)2和AgNP;同时,AgNP对粪肠球菌生物膜的抑制效果显著强于Ca(OH)2.Ca(OH)2在7d时对粪肠球菌生物膜的抑制效果强于1d时;Ca(OH)2+AgNP和AgNP在7d和1d时对粪肠球菌生物膜的抑制效果无显著差异.结论:纳米银颗粒联合氢氧化钙对饥饿期粪肠球菌生物膜具显著的抑制作用.     

电解质水对粪肠球菌生物膜的杀菌作用研究

目的:观察强酸性电解质水(SAEW)和碱性电解质水(AEW)对粪肠球菌生物膜的杀菌作用。方法:利用BioFluxTM200微流体系统建立粪肠球菌生物膜模型,并分别用SAEW、AEW、52.5 g/L次氯酸钠液(阳性对照)和生理盐水(阴性对照)处理1、5、10 min后,激光共聚焦显微镜(CLSM)观察并采集荧光图像;ImageJ图像分析软件分析并计算各处理组不同时间点的活菌比例。结果:SAEW、AEW和52.5 g/L次氯酸钠液处理1、5、10 min各时间点的活菌比例都显著低于生理盐水组(P<0.05),其中AEW组活菌比例下降幅度最小,与SAEW和52.5 g/L次氯酸钠液处理组相比,各时间点均有统计学差异(P<0.05);而SAEW与52.5 g/L次氯酸钠液处理组相比,各时间点的活菌比例均无统计学差异(P>0.05)。结论:SAEW和AEW对粪肠球菌均有杀菌作用,其中SAEW的作用与52.5 g/L次氯酸钠液相当;而AEW的杀菌效果较弱,且需要延长作用时间。     

饥饿期粪肠球菌生物膜毒力因子明胶酶E表达研究

目的:研究粪肠球菌生物膜形成相关毒力因子明胶酶E(gelE)在饥饿期及药物作用后的表达情况。方法:体外建立对数期、稳定期、饥饿期粪肠球菌生物膜模型,分别以1%、2.5%、5.25%次氯酸钠溶液作用于各时期粪肠球菌生物膜后,用Real-time PCR对gelE的基因表达相对量进行检测。结果:gelE基因表达相对量,饥饿期高于稳定期及对数期(P<0.05);用药后,3个期gelE基因表达相对量,5.25%次氯酸钠溶液组低于2.5%次氯酸钠溶液组和1%次氯酸钠溶液组(P<0.05)。结论:饥饿期粪肠球菌生物膜形成相关毒力因子gelE较对数期及稳定期表达增强。次氯酸钠浓度依赖性可抑制粪肠球菌毒力因子gelE的表达。     

PIPS联合纳米银对根管粪肠球菌抗菌效果研究

目的 探究激光引发光声流系统(PIPS)和纳米银(AgNPs)联合使用对根管内粪肠球菌生物膜的抗菌效果。方法 收集36颗单根管离体牙,建立粪肠球菌感染根管实验模型,将样本随机分成6组,采用0.9%NaCl、2%NaClO、0.1%AgNPs溶液分别联合传统手动冲洗(CNI)或PIPS对根管进行冲洗,使用菌落计数法测定治疗前后根管内粪肠球菌生物膜菌落数,并计算菌落计数减少的百分比。结果 所有实验组粪肠球菌生物膜的抑制效果均强于对照组(P<0.05),使用PIPS辅助0.9%NaCl、2%NaClO、0.1%AgNPs冲洗组的降幅均分别大于CNI辅助0.9%NaCl、2%NaClO、0.1%AgNPs冲洗组(P<0.05)。PIPS辅助0.1%AgNPs冲洗组的降幅明显大于PIPS辅助2%NaClO冲洗组(P<0.05)。结论 PIPS辅助AgNPs溶液冲洗可以显著提高根管内粪肠球菌生物膜的清除效果。     

乳酸链球菌素联合次氯酸钠对粪肠球菌的体外抗菌作用研究

目的    研究乳酸链球菌素（Nisin）联合次氯酸钠（NaClO）对粪肠球菌的体外抗菌作用。方法    采用微量稀释法测定Nisin和NaClO对悬浮态粪肠球菌的最低抑菌浓度（MIC）和最低杀菌浓度（MBC）;通过微量棋盘法计算分级抑制浓度指数（FICI）以判定两药联合效应;用Time-Kill实验来测定Nisin和NaClO单用或联合使用的杀菌动力学。结果    Nisin的MICNisin单药和MBCNisin单药分别为6、15 μg/mL,NaClO的MICNaClO单药和MBCNaClO单药分别为0.0625%、0.25%。两药联合应用时的FICI为0.504,对粪肠球菌的抗菌作用为相加作用。24 h内的Time-Kill曲线表明,在4 × MIC单药时,Nisin和NaClO分别单独应用12、8 h可完全杀灭粪肠球菌;Nisin与NaClO联合应用时2 × MIC单药即可在12 h后出现杀菌作用,从4 × MIC单药开始,在4 h内达到完全杀灭作用。结论    Nisin对粪肠球菌具有抗菌作用,且呈浓度依赖性。在体外将Nisin与NaClO联合应用时,对粪肠球菌的抗菌作用表现为相加作用,较单独应用时可在更短时间内以更低的浓度显示出杀菌作用。     

An in vitro evaluation of the ability of ozone to kill a strain of Enterococcus faecalis

AIM: To evaluate the potential of ozone as an antibacterial agent using Enterococcus faecalis as the test species. METHODOLOGY: Ozone was produced by a custom-made bench top generator and its solubility in water determined by ultraviolet (258 nm) spectrophotometric analysis of solutions through which ozone was sparged for various time-periods. The antibacterial efficacy of ozone was tested against both broth and biofilm cultures. Ozone was sparged for 30, 60, 120 and 240 s, through overnight broth cultures of a strain of E. faecalis (E78.2) and compared with those that were centrifuged, washed and resuspended in water. Enterococcus faecalis (E78.2) biofilms were grown on cellulose nitrate membrane filters for 48 h and suspended in water through which ozone gas was sparged with stirring for 60, 120 and 240 s in a standard fashion. In a separate test, biofilms were also exposed to gaseous ozone. Sodium hypochlorite (NaOCl) was used as a positive control. All experiments were repeated four times. RESULTS: There were significant (P < 0.05) reductions of bacteria in the unwashed (2 log(10) reductions) and washed (5 log(10) reductions) broth cultures following 240 s applications. Biofilms incubated for 240 s with ozonated water showed no significant reduction in cell viability attributable to ozone alone, whereas with NaOCl no viable cells were detected over the same time. Gaseous ozone applied for 300 s had no effect on these biofilms. CONCLUSIONS: Ozone had an antibacterial effect on planktonic E. faecalis cells and those suspended in fluid, but little effect when embedded in biofilms. Its antibacterial efficacy was not comparable with that of NaOCl under the test conditions used.     

Evaluation of the effectiveness of sodium hypochlorite used with three irrigation methods in the elimination of Enterococcus faecalis from the root canal, in vitro

The effectiveness of 4.0% sodium hypochlorite (NaOCl) used with three irrigation methods in the elimination of Enterococcus faecalis from the root canal was tested in vitro . Root canals contaminated with E. faecalis were treated as follows: (i) irrigation with 2 mL of NaOCl solution and agitation with hand files; (ii) irrigation with 2 mL of NaOCl solution and ultrasonic agitation; (iii) irrigation with NaOCl alternated with hydrogen peroxide. Contaminated canals irrigated with sterile saline solution served as the control. Paper points used to sample bacteria from the root canals were transferred to tubes containing 5 mL of brain heart infusion (BHI) broth. Tubes were incubated and the appearance of broth turbidity was indicative of bacteria remaining in the root canal. There were no statistically significant differences between the experimental groups. However, NaOCl applied by the three methods tested, was significantly more effective than the saline solution (control group) in disinfecting the root canal.     

次氯酸钠溶液抗菌体积分数的测定

目的评价次氯酸钠溶液对牙龈卟啉单胞菌、中间普雷沃菌、具核梭杆菌和粪肠球菌牙髓病原菌的体外抗菌性能。方法采用琼脂稀释法测定次氯酸钠溶液对牙龈卟啉单胞菌、中间普雷沃菌、具核梭杆菌和粪肠球菌的抗菌活性,其抗菌活性以最低抑菌体积分数(MIVF)和最低杀菌体积分数(MBVF)表示。结果次氯酸钠溶液对牙龈卟啉单胞菌、中间普雷沃菌、具核梭杆菌和粪肠球菌的MIVF分别为0.035%、0.07%、0.018%和0.28%,MBVF分别为0.035%、0.07%、0.035%和0.28%,其总的抗菌效果排序分别为具核梭杆菌、牙龈卟啉单胞菌、中间普雷沃菌和粪肠球菌。结论次氯酸钠对口腔常见厌氧菌有抑制和杀灭作用,具有理想的抗菌性能。0.5%MBVF的次氯酸钠溶液的抑菌性能可满足临床需求。     

Antimicrobial efficacy of ozonated water, gaseous ozone, sodium hypochlorite and chlorhexidine in infected human root canals

AIM: To determine the antimicrobial efficacy of ozonated water, gaseous ozone, sodium hypochlorite and chlorhexidine in human root canals infected by Enterococcus faecalis. METHODOLOGY: Thirty human maxillary anterior teeth were prepared and inoculated with E. faecalis for 60 days. Eppendorf tubes were connected to the coronal portion of the teeth. Urethane hoses were attached to the tubes and to the entrance of a peristaltic pump. The exit of the apparatus corresponded to the apical portion of the root canals. The test irrigating solutions were ozonated water, gaseous ozone, 2.5% sodium hypochlorite (NaOCl), 2% chlorhexidine that circulated at a constant flow of 50 mL min(-1) for 20 min. Samples from the root canals were collected and immersed in 7 mL Letheen Broth (LB), followed by incubation at 37 degrees C for 48 h. Bacterial growth was analysed by turbidity of the culture medium and subculture on a specific nutrient broth. A 0.1 mL inoculum obtained from LB was transferred to 7 mL of brain heart infusion and incubated at 37 degrees C for 48 h. Bacterial growth was checked by turbidity of the culture medium carried out in triplicate. RESULTS: No solution used as an irrigant over a 20-min contact time demonstrated an antimicrobial effect against E. faecalis. CONCLUSION: The irrigation of infected human root canals with ozonated water, 2.5% NaOCl, 2% chlorhexidine and the application of gaseous ozone for 20 min was not sufficient to inactivate E. faecalis.     

醋酸氯己定和次氯酸钠对粪肠球菌抑菌作用的体外研究

目的：体外药敏试验评价醋酸氯己定（CHA）和次氯酸钠（NaCIO）对粪肠球菌的抑菌效果。方法：采用纸片扩散法药敏试验观察不同浓度醋酸氯已定和次氯酸钠对粪肠球菌的抑菌环直径（mm）,方差分析评价药物对粪肠球菌的抑菌效果。结果：2％的CHA抑菌环最大;1％CHA、0．5％CHA和5．25％NaCl0的抑菌环直径无显著性差异,均大于2．5％NaCl0;NaCl0抑菌环直径随浓度的增大而增大。0．5％NaCl0抑菌环与对照组无显著性差异。结论：低浓度CHA即可达到高浓度NaCl0的抑菌效果,高浓度NaCl0的抑菌效果优于低浓度NaCl0。     

粪肠球菌体外根尖生物膜模型的建立及形态学研究

目的 通过体外实验建立根尖生物膜模型,观察粪肠球菌根尖生物膜模型的形态及结构。方法 选取人新鲜拔除的单根离体牙24颗,随机分为8组,每组3颗,浸泡于粪肠球菌 ATCC 29212菌悬液中,分别于1、2、7d建立根尖生物膜模型。使用扫描电子显微镜（ SEM）观察生物膜形成过程;使用碘化丙啶（PI）和刀豆球蛋白 A异硫氰酸荧光素（ConA-FITC）对生物膜细菌及细胞外基质进行双染色,通过激光共聚焦扫描电子显微镜（CLSM）观察其形态和结构;计算生物膜覆盖率。结果 随时间的增加,根尖样本表面细菌覆盖面积增加,7d组与1、2d组间的差异存在统计学意义（P<0.05）, 1d与2d组间的差异无统计学意义（P>0.05）。 7d组形成生物膜结构,其样本表面的覆盖率为 17.23%±1.52%,样本表面有大量细菌附着,细菌被细胞外基质包绕,并呈群落分布。生物膜中的细菌被 PI染成红色,细胞外基质被 ConA-FITC染成绿色,可见多层次的空间结构以及亲水性通道。结论 粪肠球菌根尖生物膜于培养7d时形成完整的生物膜结构,由细菌群落及其周围大量的不定型基质组成。     

An in vitro evaluation of the antimicrobial efficacy of irrigants on biofilms of root canal isolates

AIM: The bactericidal effect of four antimicrobial agents was investigated against single-species biofilms derived from a range of root canal isolates. METHODOLOGY: Single-species biofilms of Prevotella intermedia, Peptostreptococcus micros, Streptococcus intermedius, Fusobacterium nucleatum and Enterococcus faecalis were generated on membrane filter discs and subjected to 15 min or 1 h incubation with 5 p.p.m. colloidal silver, 2.25% sodium hypochlorite (NaOCl), 0.2% chlorhexidine, 10% iodine or phosphate buffered saline (PBS) as a control. The antimicrobial activity of the agents was neutralized and the bacterial cells were harvested from the discs by vortexing, serially diluted in reduced transport fluid, plated on fastidious anaerobe agar containing 5% horse blood, incubated anaerobically and colony-forming units calculated. RESULTS: Iodine and NaOCl were more effective than chlorhexidine except against P. micros and P. intermedia where they were all 100% effective. Iodine and NaOCl elicited a 100% kill after 1 h incubation for all strains used. However, after 15 min, they showed differing bactericidal effects depending on the strain. None of the agents were effective against F. nucleatum after 15 min but NaOCl, iodine and chlorhexidine were all effective after 1 h. Colloidal silver was generally ineffective. CONCLUSIONS: The effectiveness of a particular agent was dependent on the nature of the organism in the biofilm and on the contact time. NaOCl was generally the most effective agent tested, followed by iodine. However the clinical efficacy of these agents must be considered in light of the complex root canal anatomy and polymicrobial nature of root canal infections.