变形链球菌和远缘链球菌致龋性的研究近况

变形链球菌和远缘链球菌是人类牙齿的主要致龋菌。近年研究表明两种细菌的生物学特性存在一定差异，远缘链球菌与龋病的活跃性密切相关，本文对两种细菌的流行病学、粘附机理、多的合成产酸酸耐酸进行了综述，为今后龋病病因学研究，抗龋疫的制备提供参考。     

PCR同时检测变形链球菌和远缘链球菌

目的:建立一种快速、简便地同时检测变形链球菌和远缘链球菌的方法。方法:以变形链球菌Dextranase基因设计引物,用PCR检测变形链球菌群细菌。结果:变形链球菌(血清型c、e、f)的PCR扩增产物为720 bp;远缘链球菌(血清型d、g)的PCR扩增产物为460 bp;道勒链球菌(血清型h)910 bp;仓鼠链球菌(血清型a),鼠链球菌(血清型b)均为1 270 bp。其它异种菌均不能扩增出产物,因此该PCR检测具有高度的特异性。从细菌纯培养物中PCR检测的敏感性为105菌落形成单位(colony-form ing un its,CFU)。结论:PCR能同时检测变形链球菌和远缘链球菌。该检测方法具有较高的敏感性和特异性,有望运用于临床检测。     

变形链球菌与远缘链球菌致龋毒力因素的差异

变形链球菌与远缘链球菌均为重要的龋菌。但两者在一些生化特征和重要的毒力因素方面有所不同。本文对这两种细菌的粘附性和产酸，耐酸性等单方面的差异进行了综述，旨在解释两菌的致龋性和分布特点。     

套式PCR检测唾液中变形链球菌及远缘链球菌

目的 探索一种快速、简便地从人类唾液中同时检测变形链球菌和远缘链球菌的方法。方法 分别以变形链球菌gtfI和远缘链球菌gtfB基因设计两组成套引物，首先用套式PCR（二次PCR）检测变形链球菌和远缘链球菌标准株和临床株，然后用套式PCR直接从唾液中检测这两种细菌。结果 变链菌（血清型c,e,f)的标准株及临床株第1次PCR扩增产物为517bp，第2次扩增产物为468bp；远缘链球菌（血清型d,g)及道勒链球菌（血清型h)的标准株及临床株第1次PCR扩增产物为712bp，第2次扩增产物为663bp;其他异种菌均不能扩增出产物，因此该PCR检测具有高度的特异性。细菌纯培养物及唾液PCR检测的敏感性分别是：第1次PCR为10^5CFU，第2次PCR为10^3CFU。结论 套式PCR能快速在人类唾液中同时检测变形链球菌和远缘链球菌。该检测方法有望运用于临床检测，对揭示两种细菌与龋病发生关系的研究具有一定价值。     

变形链球菌和远缘链球菌耐氟菌株超微结构观察

目的：探讨变形链球菌和远缘链球菌耐氟突变后,其超微结构变化及氟化物对细菌结构的影响。方法：将变形链球菌和远缘链球菌分为3组：A为正常对照组;B为加氟孵育组;C为耐氟菌株组。透射电镜观察3组细菌的超微结构。结果：与正常亲代菌株相比,加氟孵育后的菌株及耐氟菌株菌体出现不同程度超微结构改变,包括胞浆电子密度减低,细胞肿胀,内容物外溢,远缘链球菌链状结构消失等。结论：变形链球菌和远缘链球菌与氟共同孵育及耐氟突变后,其超微结构发生了改变,表现为自溶活动及解链活动增强。     

聚合酶链反应(PCR)方法检测远缘链球菌

目的 :建立一种快速、特异、敏感的远缘链球菌PCR检测方法。方法 :根据已发表的远缘链球菌葡聚糖酶基因 (dexA)的特异片断设计一对寡核苷酸引物 ,对 12种变形链球菌群中包含a～h 8种血清型的细菌及 2 3种常见的口腔菌中进行PCR扩增 ,扩增产物电泳鉴定 ,并对特异片断进行回收 ,测序。结果 :变形链球菌群标准菌株中 ,只有远缘链球菌 (血清d、g型 )产生特异的扩增片断 ,测序结果与已发表的文献结果完全一致 ,且显示了极高的灵敏性 ,≥ 2× 10 2 个细胞均可以用该方法检出。结论 :PCR方法可以用于快速灵敏的检测远缘链球菌 ,比传统的培养鉴定方法显示了更大的优越性     

中药五倍子对变形链球菌及远缘链球菌抑制作用的体外研究

目的：研究五倍子及其主要成分鞣酸在体外对变链菌及远缘链球菌的抑制作用，并摸索适宜剂型和浓度。方法：本实验采用纸片扩散法研究五种浓度五倍子煎剂，五倍子浸剂，鞣酸标准品抑制变链菌及远缘链球菌生长的作用。结果：65.2mL/L以上浓度五倍子煎剂，浸剂，鞣酸对变链菌及远缘链球菌生长均有抑制作用，其中五倍子浸剂抑菌作用最强。抑菌作用随着各剂型浓度的增国而增强。结论：五倍子可以抑制变链菌及远缘链球菌生长，其中浸剂效果最优，鞣酸组效果较差，可初步推测五倍子中含有其他协同鞣酸抑菌的成分。     

婴儿期不同喂养方式儿童猛性龋病原菌分析

目的：比较不同喂养方式幼儿问变形链球菌族细菌(MS)定殖差异，为龋病预防提供依据。方法：采用细菌分离、培养、生化试验和免疫学血清凝集试验对幼儿乳前牙牙菌斑内细菌进行分离鉴定，分析病原菌的定殖情况以及与婴儿期喂养方式的关系。结果：幼儿猛性龋优势菌为变形链球菌。不同喂养方式猛性龋幼儿之间MS定殖水平无差别。母乳喂养无龋儿童牙菌斑内MS水平明显高于奶瓶喂养者。结论：幼儿猛性龋主要致病菌为变形链球菌，6个月以上的母乳喂养有利于MS定殖。     

变形链球菌和远缘链球菌的可溶性多糖和不溶性多糖的测定

目的 :对变形链球菌和远缘链球菌产生的可溶性多糖和不溶性多糖进行测定。方法 :酚硫酸法。结果 :发现变形链球菌产生的可溶性多糖显著高于远缘链球菌 ,而远缘链球菌产生的不溶性多糖高于变形链球菌。结论 :表明在对牙面的粘附作用中远缘链球菌更依赖于蔗糖及其产生的不溶性葡聚糖 ,而变链菌则较少依赖于葡聚糖 ,而可能主要依赖于唾液的介导。     

五倍子对变形链球菌及远缘链球菌产酸影响的体外研究

目的 研究五倍子及其主要成分鞣酸在体外对变链菌及远缘链球菌的产酸作用有何影响。方法 本实验采用气相色谱研究50％五倍子浸剂、37％鞣酸标准品、2％氟化钠液对变链菌及远缘链球菌产酸作用的影响。结果 50％五倍子浸剂有机酸含量最少,其次分别为2％氟化钠组、37％鞣酸标准品组,3组间无统计学的差异。结论 五倍子对变链菌及远缘链球菌产酸有较强的抑制作用,其作用与氟化钠相当。     

高发龋和无龋儿童菌斑中变形链球菌及远缘链球菌的任意引物PCR检测

目的采用任意引物聚合酶链反应（arbitrarily primed-polymerase chain reaction，AP-PCR）法探讨高发龋和无龋儿童牙菌斑致龋菌的检出情况，分析变形链球菌及远缘链球菌与乳牙龋齿发生的关系。方法从20例高发龋患儿和20名无龋儿童的牙菌斑中分离、鉴定变形链球菌和远缘链球菌，以Shiroza的DNA提取方法提取细菌基因组DNA，以形态学、生化和AP-PCR的方法鉴定变形链球菌和远缘链球菌。结果AP-PCR法检测发现高发龋患儿变形链球菌和远缘链球菌的检出率分别为100％和40％，而无龋儿童两种细菌的检出率分别为75％和5％，差异有统计学意义。结论用AP-PCR法检测发现高发龋患儿变形链球菌和远缘链球菌的检出率均明显高于无龋儿童，口腔中定植的变形链球菌和远缘链球菌是乳牙龋高发的危险因素。     

不同龋敏感人群牙菌斑中变形链球菌的检测

目的 :探讨不同龋敏感人群牙菌斑中变形链球菌的检出情况和血清分型与龋病的关系。方法 :采用经典生化和血清学鉴定方法 ,对不同龋敏感人群菌斑中的变形链球菌进行分离、鉴定。结果 :血清c型变形链球菌在牙菌斑中检出率最高 ,在高龋组中变形链球菌群细菌和变形链球菌培养阳性率明显高于无龋组 (P <0 .0 0 5 )。结论 :变形链球菌群细菌尤其是变形链球菌的数量与龋病发生有直接关系     

Effect of a bacteriocin-producing strain of Streptococcus sobrinus on infection and establishment of Streptococcus mutans on tooth surfaces in rats

The effect of bacteriocin produced by Streptococcus sobrinus MT6223 on infection and establishment of Streptococcus mutans MT6222 was studied in specific pathogen-free rats. These strains were isolated from a carious lesion of a single subject. S. mutans MT6222 was found to be susceptible to the growth inhibitory action of S. sobrinus MT6223. When simultaneously inoculated into the oral cavity of rats, even a small inoculum (10(5) CFU) of S. sobrinus MT6223 completely inhibited colonization of S. mutans MT6222 on the tooth surface. Also, S. sobrinus MT6223 eliminated S. mutans MT6222 when MT6223 (10(8) CFU) was inoculated 2 days after the inoculation of 10(8) CFU cells of MT6222. Similar results were obtained in dental plaque samples from the tooth surface and the fissures of the upper molars at the end of the experiment. However, when S. sobrinus MT6223 (10(8) CFU) was inoculated 2 weeks after the inoculation of S. mutans MT6222 (10(8) CFU), MT6223 coexisted with MT6222. However, the plaque samples showed that MT6223 inhibited the establishment of MT6222 on smooth surfaces, but not in fissures. In addition, MT6223 protected against subsequent infection with MT6222. However, a nonbacteriocinogenic mutant of S. sobrinus MT6223 did not inhibit the infection and establishment of S. mutans MT6222.     

幼儿猛性龋病原菌的分离鉴定

目的:确定幼儿猛性龋的优势病原菌,为其防治提供依据。方法:采用细菌分离培养、形态学、生理生化学和 DNAG+C mol%测定方法,对30名2~5岁猛性龋患儿牙菌斑菌丛进行分离鉴定,采样部位为上颌患龋乳切牙龋损部位及邻近健康釉质表面,对照组的非猛性龋和无龋儿童则采集上颌乳前牙唇面颈1/3处的菌斑。结果:猛性龋儿童龋损部位变链菌和远缘链球菌的检出率及两个采样部位菌斑标本中变链菌和远缘链球菌的检出水平均显著高于非猛性龋和无龋儿童(P<0105)。结论:变链菌和远缘链球菌为幼儿猛性龋的优势病原菌。关键词 幼儿猛性龋变链菌远缘链球菌。     

Mutans streptococci, lactobacilli and caries prevalence in 11-and 12-year-old Icelandic children

Since the caries prevalence has not declined in Iceland as it has in other European countries, it was of interest to study the presence of cariogenic bacteria in Icelandic children. The prevalence of salivary mutans streptococci and lactobacilli was examined in 217 11- and 12-yr-old Icelandic children and was related to their caries prevalence. In 2% of the children mutans streptococci were not found whereas 31% and 35% of the 11-yr-olds and 12-yr-olds respectively carried more than 10(6) CFU per ml saliva. Lactobacilli were not detected in 8% of the children whereas 28% of the 11-yr-olds and 23% of the 12-yr-olds had more than 10(5) CFU per ml saliva. The mean caries prevalence (DFS), initial caries included, for 11-yr-olds was 21.6 and for 12-yr-olds 28.8. Both mutans streptococci and lactobacilli were significantly correlated to each other as well as to caries prevalence. An increased number of these microorganisms, especially the mutans streptococci, were associated with an increased DFS. Children with high salivary counts of mutans streptococci and lactobacilli showed four times higher DFS than children with low numbers. The magnitude of salivary cariogenic bacteria as well as caries prevalence was found to resemble the situation in Sweden 10 yr ago. S. mutans (serotype c/e/f) was carried by all mutans streptococci positive children. S. sobrinus (serotype d/g) was found in 60 children (30.2%). Children with both S. mutans and S. sobrinus had significantly higher salivary counts of mutans streptococci and lactobacilli as well as caries prevalence than children with only S. mutans.     

Simple and rapid detection of Streptococcus mutans and Streptococcus sobrinus in human saliva by polymerase chain reaction

Streptococcus mutans and Streptococcus sobrinus are major pathogens causing dental caries in humans. A simple and rapid method to detect these species in human saliva simultaneously was developed using the polymerase chain reaction (PCR). Chromosomal DNA was extracted by boiling bacterial cells in lysis solution containing 1% Triton X-100. Oligonucleotide primers specific for portions of the glucosyltransferase genes (gtfB of S. mutans and gtfI of S. sobrinus) were designed. After PCR using two sets of these primers, S. mutans and S. sobrinus were specifically identified. The method was capable of amplifying DNA fragments specific for these species from chromosomal DNA extracted from 1 x 10(3) cells, or from 10 microliters of clinical saliva samples containing 1 x 10(3) colony-forming units of either streptococcal species. A second PCR, using the first PCR product as a template with newly designed internal primers, made it possible to detect 1 x 10(2) colony-forming units of either streptococcal species in 10 microliters of saliva samples. These results indicate that the PCR method developed in this study is useful for detecting S. mutans and S. sobrinus in saliva and that it can be used in epidemiological studies to evaluate the prevalence level of these organisms.     

In vitro inhibitory effects of Polygonum cuspidatum on bacterial viability and virulence factors of Streptococcus mutans and Streptococcus sobrinus

OBJECTIVES: Polygonum cuspidatum has been used in Korean folk medicine to improve oral hygiene. This study was performed to evaluate the effects of methanol extract from root of P. cuspidatum (MEP) on bacterial viability and the virulence factors of Streptococcus mutans and Streptococcus sobrinus. METHODS: To test the effects of MEP on bacterial viability, we determined the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against 20 bacterial strains, including S. mutans and S. sobrinus, using a micro-dilution assay. In case of S. mutans and S. sobrinus, the assays for time-kill and bacterial growth rate at sub-MIC concentrations were also performed. To determine effects of the extract on the virulence factors of S. mutans and S. sobrinus, the assays for sucrose-dependent adherence, water-insoluble glucan formation, glycolytic acid production, and acid tolerance were performed at sub-MIC levels. Phytochemical analysis for constituents of MEP was carried out. RESULTS: MEP showed a broad antibacterial range (MIC 0.5-4 mg/ml). The MBC was two to four times higher than the MIC. The time-kill curves showed S. mutans and S. sobrinus were significantly killed after 1h of incubation. At sub-MIC levels, doubling times of S. mutans and S. sobrinus dose-dependently increased up to 211% and 123%, respectively. At sub-MIC levels, MEP also showed inhibitory effects on the virulence factors of S. mutans and S. sobrinus in a dose-dependent fashion. Phytochemical analysis revealed the presence of alkaloids, sterol/terpenes, tannins, flavonoids, and carbohydrates. CONCLUSION: These data indicate that MEP has inhibitory effects on bacterial viability at higher concentrations (> or =MIC) and the virulence factors of S. mutans and S. sobrinus at sub-MIC concentrations, suggesting that it might be useful for the control of dental plaque formation and subsequent dental caries formation.     

儿童猛性龋变链菌分离株的产酸性

目的：确定儿童猛性龋变链菌和远缘链球菌临床分离株的产酸性。方法：采用酸度计和自动气相色谱仪比较儿童猛性龋、非猛性龋、无龋变链菌（各6株）和远缘链球菌（儿童猛性龋6株,非猛性龋和无龋各3株）的临床株降低环境pH值的能力（ΔpH）和乳酸产量,并以此推测其致龋力。结果：远缘链球菌ΔpH及乳酸产量均高于变链菌,特别是在低pH水平下,二者差异更为显著（P＜0．05）。儿童猛性龋远缘链球菌分离株ΔpH和乳酸产量显著大于非猛性龋和无龋分离菌株（P＜0．05）。结论：远缘链球菌的产酸能力强于变链菌;儿童猛性龋远缘链球菌分离株较非猛性龋和无龋儿童分离菌株产酸性更强。