龙胆泻肝汤对阴道加德纳菌的体外抑制作用

目的研究龙胆泻肝汤对阴道加德纳菌(GV)的体外抑制作用。方法利用稀释法检测龙胆泻肝汤对GV增殖的影响,并测定药物的最低抑菌浓度,通过革兰氏染色法观察龙胆泻肝汤对GV黏附人宫颈癌Hela细胞的影响,MTT法检测龙胆泻肝汤对GV细胞毒性的影响,96孔微量板法检测龙胆泻肝汤对GV生物膜形成的影响,qRT-PCR法检测龙胆泻肝汤对GV BAP、sialidase mRNA表达的影响。结果龙胆泻肝汤水提物、70%醇提物和90%醇提物对GV的最低抑菌浓度分别为(62.5±3.6)、(15.6±1.5)、(125.0±2.8) mg/mL。龙胆泻肝汤70%醇提物(1.56、15.6 mg/mL)能降低GV对Hela细胞的黏附,抑制GV生物膜的形成(P<0.01),下调BAP、sialidase mRNA表达(P<0.05,P<0.01),龙胆泻肝汤70%醇提物(15.6 mg/mL)抑制GV对Hela细胞的毒性作用(P<0.01)。结论龙胆泻肝汤70%乙醇提物能显著抑制GV增殖,并能抑制GV细胞毒性、粘附能力和生物膜形成。     

Potential impact of the sewer system on the applicability of alcohol and tobacco biomarkers in wastewater‐based epidemiology

Understanding the actual consumption of alcohol and tobacco in the population is important for forming public health policy. For this purpose, wastewater‐based epidemiology has been applied as a complementary method to estimate the overall alcohol and tobacco consumption in different communities. However, the stability of their consumption biomarkers – ethyl sulfate, ethyl glucuronide, cotinine, and trans‐3′‐hydroxycotinine – in the sewer system has not yet been assessed. This study aimed to conduct such assessment using sewer reactors mimicking conditions of rising main, gravity sewer, and wastewater alone, over a 12‐hour period. The results show that cotinine and trans‐3′‐hydroxycotinine are relatively stable under all sewer conditions while ethyl sulfate was only stable in wastewater alone and gradually degraded in rising main and gravity sewer conditions. Ethyl glucuronide quickly degraded in all reactors. These findings suggest that cotinine and trans‐3′‐hydroxycotinine are good biomarkers to estimate tobacco consumption; ethyl sulfate may be used as a biomarker to estimate alcohol consumption, but its in‐sewer loss should be accounted for in the calculation of consumption estimates. Ethyl glucuronide, and probably most of glucuronide compounds, are not suitable biomarkers to be used in wastewater‐based epidemiology due to their in‐sewer instability.     

A new in vitro model for the study of microbial microleakage around dental restorations: a preliminary qualitative evaluation

AIM: The aim of this study was to develop an in vitro model to replicate microbial microleakage at a tooth/ restoration interface using a constant depth film fermentor (CDFF). METHODOLOGY: Amalgam restorations were placed in machined bovine dentine cylinders and sealed externally with varnish, leaving a 1-mm perimeter exposed around the tooth/restoration interface. The dentine cylinders were housed in a CDFF and 300-microm thick microcosm dental plaques were grown over their exposed surfaces. The biofilms were maintained with a mucin-containing artificial saliva for up to 8 weeks. Cylinders were aseptically removed from the CDFF (at 1, 2, 4, & 8 weeks) and surface-decontaminated with validated protocols prior to splitting and sampling of apposing amalgam and dentine surfaces. Scanning electron microscopy (SEM) was used to ascertain the position and structure of the bacterial aggregates. Bacterial viability was determined by vital staining of the bacteria in situ. RESULTS: At all sampling times, SEM showed cocci, rods and filaments on both amalgam and dentine surfaces; some originated as cascades from the surface biofilm and extended into the tooth/restoration microspace. Vital staining showed the majority of bacteria from both dentine and amalgam surfaces to be viable. CONCLUSION: This preliminary investigation showed that the CDFF may be a valuable tool for the in vitro study of the dynamics of microbial microleakage around dental restorations.     

Characterization of in vitro oral bacterial biofilms by traditional and molecular methods

The aim of this study was to compare culture-based bacterial isolation methods with direct amplification and cloning of 16S rRNA genes from oral biofilms grown in an in vitro model. The model used was a constant depth film fermentor which was inoculated with pooled human saliva. The use of culture techniques and cloning resulted in the identification of 36 different bacterial species from the saliva inoculum and from the biofilms. Of these, only five were detected solely by molecular methods. Three taxa were detected which, according to the databases, were unidentified. Using the molecular methods of detection, differences in the number of species observed were found using different 16S rRNA gene primers and numbers of PCR cycles. We have shown that microcosm supragingival plaque biofilms grown in a fermentor consisted of a community most of the members of which could be cultivated on laboratory media.     

五倍子对口内菌斑生物膜活性影响的体外研究

目的应用口内菌斑生物膜模型观察五倍子水提取物对菌斑生物膜活性的影响。方法将牙釉质磨片粘结于两侧下颌第一恒磨牙颊侧获得24h口内菌斑生物膜模型,实验组和对照组分别用6mg/ml五倍子水提取物和生理盐水处理3min,应用溴化乙锭/荧光素双乙酸盐(ethidium bromide/fluorescein diacetate,EB/FDA)染色技术和激光共聚焦显微镜(confocal laser scan microscope,CLSM)观察五倍子水提取物对生物膜活性的影响。结果CLSM观察可见所有样本釉质表面均有细菌的定植,有早期的生物膜形成。对照组菌斑生物膜中细菌的活性明显高于实验组,两者相比具有显著差异(P<0.05)。结论五倍子水提取物在较短的时间内就能对生物膜中的细菌产生一定的杀伤效应,使其活性下降。     

Interaction between GIC and S. sanguis biofilms: antibacterial properties and changes of surface hardness

Objective

The aim of this study was to investigate the interaction of Streptococcus sanguis with two glass-ionomer formulations (GIC:A containing fluoride and GIC:B without fluoride) with particular reference to bacterial growth and changes in hardness of the cement with respect to time.

Materials and methods

Discs of two water activated glass-ionomer cements were prepared according to the manufacturer's instruction. Hydroxyapatite discs (HA) were used as controls. 3D laser scanning technique was used to characterize surface roughness and area of the substrate prior to growing biofilms. Surface hardness was evaluated before and after biofilm growth. A constant depth film fermenter system (CDFF) was used to grow S. sanguis biofilms on the specimens in a similar manner to that described previously by Wilson et al. in 1995. For susceptibility measurement, specimens were removed from CDFF aseptically over periods up to 14 d after the first colonization with bacteria. Counts of viable bacterial in the accumulating biofilm layer on each surface were measured and converted to colony forming units per unit surface area. To determine the effect of storage media, hardness discs were exposed to distilled water, lactic acid pH 4, lactic acid pH 5, citric acid pH 5, artificial saliva and S. sanguis biofilms. Twenty-four hours after preparing and subsequent autoclaving, specimens were transferred to a vessel containing 40 ml storage medium. The specimens were investigated for periods up to 7 d.

Results

The viable counts of S. sanguis per mm² on GIC:A were significantly less than those on HA and GIC:B during the first 5 d (p < 0.05). The viable counts of bacteria on the surface of GIC:B were lower during the initial 5 d when compared to HA. Exposure of GIC:A and GIC:B to different medium produced softening to the surface of cement. It is apparent that the effects of the biofilms are significantly greater than storage in water but similar to storage in lactic acid pH 5.

Conclusions

This investigation showed that the growth of S. sanguis biofilms were significantly affected by both glass-ionomer formulations, the greater reduction being noted on the surface of the fluoride containing GIC. S. sanguis biofilms produced reduction on the surface hardness of the cement equivalent to that seen after immersion in lactic acid at pH 5. This indicates that while S. sanguis biofilm is affected by the GIC, there is also a decrease in hardness of the cement indicating some cement degradation.     

Effects of apigenin and tt‐farnesol on glucosyltransferase activity,biofilm viability and caries development in rats

Propolis, a resinous hive product secreted by Apis mellifera bees, has been shown to reduce the incidence of dental caries in rats. Several compounds, mainly polyphenolics, have been identified in propolis. Apigenin and tt‐farnesol demonstrated biological activity against mutans streptococci. We determined here their effects, alone or in combination, on glucosyltransferase activity, biofilm viability, and development of caries in rats. Sprague‐Dawley rats were infected with Streptococcus sobrinus 6715 and treated topically twice daily as follows: (1) tt‐farnesol, (2) apigenin, (3) vehicle control, (4) fluoride, (5) apigenin +tt‐farnesol, and (6) chlorhexidine. Apigenin (1.33 mM) inhibited the activity of glucosyltransferases in solution (90–95%) and on the surface of saliva‐coated hydroxyapatite beads (35–58%); it was devoid of antibacterial activity. tt‐Farnesol (1.33 mM) showed modest antibacterial activity against biofilms and its effects on glucosyltransferases were minimal. The incidence of smooth‐surface caries was significantly reduced by apigenin +tt‐farnesol (60%), fluoride (70%), and chlorhexidine (72%) treatments compared to control (P < 0.05).