3种寡核苷酸探针对龈下菌斑中牙周致病菌的检测

目的　采用寡核苷酸探针研究龈下菌斑中3种牙周致病菌的分布。方法　利用3种寡核苷酸探针对 60例慢性牙周炎患者60个患病位点、10例健康人的10个健康对照位点龈下菌斑中的牙龈卟啉单胞菌、福赛类杆菌、牙密螺旋体进行检测。结果　牙周炎位点龈下菌斑中的牙龈卟啉单胞菌、福赛类杆菌、牙密螺旋体的检出率分别为91·67%,90·00%和95·67%,明显高于健康对照位点;有83·33%的牙周炎位点同时检出3种致病菌,3种细菌检出情况为两两正相关(P<0·01)。结论　牙龈卟啉单胞菌、福赛类杆菌、牙密螺旋体在慢性牙周炎患者龈下菌斑中的检出率很高,它们间可能存在相互协同致病作用。     

应用实时荧光定量PCR定量检测慢性牙周炎患者龈下菌斑中牙龈卟啉单胞菌

目的 检测慢性牙周交患者和牙周健康人龈下菌斑中牙龈卟啉单胞菌、总菌量和牙龈卟啉单胞菌所占比例,探讨牙龈卟啉单胞菌与牙周炎发生发展的关系.方法 采集经常规PCR方法检测牙龈卟啉单胞菌为阳性的76例慢性牙周炎患者和25例牙周健康者的龈下菌斑,应用TaqMan实时荧光定量PCR方法定量检测样本中牙龈卟啉单胞菌、总菌量;构建含有牙龈卟啉单胞菌和真细菌扩增片断的重组质粒,建立定量标准.结果 本研究设计的引物和探针具有良好的特异性及敏感性.病变位点龈下菌斑中牙龈卟啉单胞菌数量和总菌量均比健康位点高(P<0.001),两组位点牙龈卟啉单胞菌在菌斑中的比例没有差异(P>0.05);细菌数量与探诊深度间存在显著正相关关系(P<0.001),不同的探诊深度牙龈卟啉单胞菌所占比例无统计学差异(P>0.05).结论 龈下菌斑中牙龈卟啉单胞菌的数量水平及细菌总量与牙周状况、牙周炎发展有密切关系,实时荧光定量PCR对牙周病学研究具有广泛的应用前景.     

不同牙周状况龈下菌斑中牙龈卟啉单胞菌和福赛斯坦纳菌的分布

目的 分析中国牙周健康者和慢性牙周炎患者龈下菌斑中牙龈卟啉单胞菌和福赛斯坦纳菌的分布情况,探讨两种细菌在中国慢性牙周炎患者中的分布及其致病机理.方法 收集65例牙周健康者、62例慢性牙周炎患者的龈下菌斑,采用16S rRNA PCR方法检测牙龈卟啉单胞菌和福赛斯坦纳菌的分布.结果 牙周健康者牙龈卟啉单胞菌和福赛斯坦纳菌的检出率分别是21.6%、12.3%,慢性牙周炎患者病变位点两菌检出率分别为74.2%、58.1%,牙周炎健康位点两菌的检出率分别是22.6%、4.8%.牙周炎病变位点两种细菌的检出率均明显高于牙周健康者和牙周炎健康位点(P＜0.001);福赛斯坦纳菌在牙周健康者中的检出率高于牙周炎健康位点(P＜0.05);牙周健康者和牙周炎健康位点牙龈卟啉单胞菌的检出率没有差异.慢性牙周炎患者两菌联合检出率为51.6%.结论 牙龈卟啉单胞菌、福赛斯坦纳菌以及两种细菌联合感染与牙周炎密切相关.     

伴放线放线杆菌在龈下菌斑和颊黏膜分布研究

目的:比较伴放线放线杆菌(actinobac illus actinomycetem com itans,A.a)在不同类型牙周炎患者龈下菌斑和颊黏膜中的分布。方法:通过聚合酶链反应(polym erase chain reaction,PCR)对侵袭性牙周炎患者(AgP)、慢性牙周炎患者(CP)、牙周健康者口腔龈下菌斑和颊黏膜中的A.a进行检测,分析该菌分别在两部位的相对含量。结果:AgP组菌斑和颊黏膜样本中A.a阳性检出率均为41.7%,分别高于CP组(菌斑16.7%、颊黏膜10.0%)和牙周健康组(菌斑和颊黏膜均为0%)。AgP组A.a在菌斑和颊黏膜的相对含量分别为38.5%和22.2%,高于CP组(菌斑19%、颊黏膜12.75%)。结论:A.a不仅存在于龈下菌斑中,也能够粘附于颊黏膜;A.a是AgP的主要优势菌也参与了CP的菌群组成。     

维吾尔族慢性牙周炎中牙龈卟啉单胞菌菌毛fimA毒力基因型的分布

目的:分析维吾尔族慢性牙周炎患者龈下菌斑中牙龈卟啉单胞菌菌毛fimA毒力基因型的分布情况.方法:收集52例维吾尔族慢性牙周炎患者的龈下菌斑,采用16S rRNA PCR法检测牙龈卟啉单胞菌,并根据菌毛fimA毒力基因型的特异引物,用聚合酶链反应(PCR)检测Ⅱ型fimA和Ⅳ型fimA菌株的分布.结果:16S rRNA PCR法检测牙龈卟啉单胞菌在龈下菌斑中阳性检出率是76.9%.牙周袋PPD＞6 mm位点龈下菌斑标本的P.gingivalis检出率高于4＜PPD≤6 mm采样的位点,2组差异有统计学意义(P＜0.05).牙龈卟啉单胞菌菌毛fimA毒力基因型在牙龈卟啉单胞菌感染者的检出率分别是:ⅡfimA型为37.5%,ⅣfimA型为22.5%.结论:维吾尔族慢性牙周炎患者龈下菌斑中牙龈卟啉单胞菌有较高的检出率.牙龈卟啉单胞菌存在fimA毒力基因多态性.     

不同rag基因型牙龈卟啉单胞菌在慢性牙周炎患者中的分布

目的分析不同rag基因型牙龈卟啉单胞菌在慢性牙周炎患者中的分布状况。方法收集50例慢性牙周炎患者的150个龈下菌斑样本,采用16S rDNA 聚合酶链反应（PCR）法检测牙龈卟啉单胞菌在牙周炎病变位点的检出率,并根据各rag基因型的特异性引物检测牙龈卟啉单胞菌不同rag基因型在慢性牙周炎病变位点的分布。结果经16S rDNA PCR法检测,病变位点牙龈卟啉单胞菌阳性检出率为70.7%。各rag基因型在牙龈卟啉单胞菌阳性位点的总检出率：rag-1为60.4%,rag-2为23.6%,rag-3为44.3%,rag-4为15.1%;经统计学检验,rag-1和rag-3型检出率较高,高于rag-2和rag-4型（P<0.05）。结论慢性牙周炎患者龈下菌斑中的牙龈卟啉单胞菌存在rag基因多态性,rag-1和rag-3基因型牙龈卟啉单胞菌与中国辽宁地区人群慢性牙周炎的发生发展关系密切。     

不同fimA基因型牙龈卟啉单胞菌在慢性牙周炎患者中的分布

目的　分析不同fimA基因型牙龈卟啉单胞菌(P.gingivalis)在慢性牙周炎患者中的分布状况。方法　收集101例慢性牙周炎患者的龈下菌斑,采用常规培养法和16S rRNA PCR检测P.gingivalis,并根据各fimA基因型的特异引物,用聚合酶链反应(PCR)检测不同fimA基因型菌株的分布。结果　16S rRNA PCR检测P.gingivalis阳性检出率为88·1%。大多数受检牙龈下菌斑中只检测出一种fimA基因型菌株(65·1%),各fimA基因型的总检出率: ⅠfimA为24·7%;ⅡfimA为43·8%;ⅢfimA为15·7%;ⅣfimA为40·4%;VfimA为3·4%。结论　慢性牙周炎患者龈下菌斑中的牙龈卟啉单胞菌存在fimA基因多态性,ⅡfimA和ⅣfimA基因型P.gingivalis菌株与慢性牙周炎的发生发展关系密切。     

龈沟液中有机酸与龈下牙龈卟啉单胞菌间的关系

目的分析慢性牙周炎患者相同龈沟液样本中挥发性有机酸与牙龈卟啉单胞菌（Porphyromonas gingivalis,P.g）间的关系。方法应用高效毛细管电泳技术分析慢性牙周炎患者治疗前龈沟液中有机酸的浓度;应用多聚酶链反应（polymerase chain reaction,PCR）技术检测相同样本中牙龈卟啉单胞菌。结果慢性牙周炎患者龈下菌斑中牙龈卟啉单胞菌检出率显著高于牙周健康对照者;慢性牙周炎患者P.g阳性组龈沟液中丁酸与异戊酸浓度显著高于P.g阴性组。结论结果表明相同样本龈沟液中丁酸与异戊酸的表达与P.g的检出相关联,P.g是重要牙周致病菌     

牙龈卟啉单胞菌在龈下菌斑中的分布

研究牙龈卟啉单胞菌在牙周炎患者病变部位和健康部位龈下菌班中的分布情况,方法：选择６４例成年牙周炎患者,取龈下菌斑,经厌氧培养,挑战产黑色素菌落,经多聚酶链反应鉴定牙龈卟啉单胞菌。结果：产黑色素Ｇ＾－厌氧杆菌和牙龈卟啉单胞菌的患者检出率分别是６７．２％和６０．９％。牙龈卟啉单胞菌在病变部位和健康部位的检出率分别是３５．９％和２８．１％,二者差异无统计学意义;牙龈卟啉单胞菌在病变部位和健康部位的检出株     

龈下菌斑中牙龈卟啉单胞菌rgpB基因多态性研究

目的 探讨不同rgpB基因型牙龈卟啉单胞菌在慢性牙周炎发生发展中所起的作用。方法 选择不同牙周状态下的龈下菌斑样本104个，设计引物对牙龈卟啉单胞菌rgpB催化域基因进行扩增，用限制性片段长度多态性分析的方法将牙龈卟啉单胞菌分为4个基因型。结果 慢性牙周炎患者病变重部位rgpB-cdⅣ型检出率最高为52．78％，与Ⅰ、Ⅲ型比较差异有统计学意义（P〈0．05、P〈0．01），病变轻部位rgpB-cdⅡ型检出率为75．86％，显著高于其他型（P〈0．01）。结论rgpB-cdⅣ型可能与慢性牙周炎的关系最密切，在致病过程中起主要作用，而rgpB-cdⅡ型可能与慢性牙周炎无关，为健康人群或牙周健康部位的定居菌。     

Detection frequency of periodontitis-associated bacteria by polymerase chain reaction in subgingival and supragingival plaque of periodontitis and healthy subjects

The aim of this study was to compare the detection frequencies of 25 bacterial species in subgingival and supragingival plaque of 18 untreated periodontitis subjects and 12 periodontally healthy subjects. Genomic DNA was extracted from subgingival and supragingival plaque samples, and bacterial detection was performed by polymerase chain reaction of the 16S rRNA genes. Fourteen bacteria showed no relationship with periodontitis, and 11 of these 14 species were frequently detected (≥50%) in subgingival plaque in both periodontitis and healthy subjects. Nine bacteria such as Eubacterium saphenum, Prevotella intermedia, and Treponema denticola seemed to be related to periodontitis; their detection frequencies in subgingival plaque samples were higher in periodontitis than in healthy subjects, but these differences were not statistically significant by multiple comparisons (0.002≤ P< 0.05). Two species ( Mogibacterium timidum and Porphyromonas gingivalis ) were detected significantly more frequently in subgingival plaque of periodontitis subjects than of healthy subjects ( P< 0.002), with P. gingivalis being detected only in periodontitis subjects, suggesting that these two species are closely related to periodontitis. There were no significant differences in the detection frequencies of the 25 bacteria between subgingival and supragingival plaque, suggesting that the bacterial flora of supragingival plaque reflects that of subgingival plaque.     

Eikenella corrodens in the human oral cavity

The prevalence and distribution of the putative periodontal pathogen Eikenella corrodens in the human oral cavity was examined. A total of 508 oral bacterial samples were taken from 10 periodontally healthy adults (PH), 11 adult periodontitis patients (AP), and 6 localized juvenile periodontitis patients (LJP). From each subject, samples of supra- and subgingival plaque were obtained from six to eight teeth as well as samples from buccal mucosa, lateral and dorsal surfaces of tongue, tonsil, and saliva. E. corrodens was cultured from 70% of healthy subjects and 100% of periodontitis patients. Dental plaque appears to be the main oral ecological niche of E. corrodens in PH subjects since it was found in, respectively, 26% and 31% of supra- and subgingival plaque samples and rarely found in other oral sites in these subjects. It was found in 59% of both supra- and subgingival plaque samples from AP subjects, as well as 48% and 64% of supra- and subgingival plaque samples of LJP subjects. In contrast to healthy subjects, E. corrodens was found on the buccal mucosa, tongue, tonsil and in the saliva of patients with periodontitis. The microorganism constituted, on average, 1% to 2% of the total cultivable bacteria in supra- and subgingival plaque samples. The prevalence of E. corrodens in plaque samples was higher in AP and LJP subjects and was significantly different from PH subjects. Within the AP group, the prevalence of E. corrodens in subgingival plaque is significantly higher from sites with GI greater than or equal to 2. These data suggest that E. corrodens is an indigenous oral microorganism which may be an opportunistic pathogen associated with gingival inflammation.     

Comparison of the microbiota of supra- and subgingival plaque in health and periodontitis

BACKGROUND, AIMS: The purpose of the present investigation was to compare the microbial composition of supra and subgingival plaque in 22 periodontally healthy (mean age 32+/-16 years) and 23 adult periodontitis subjects (mean age 51+/-14 years). METHODS: A total of 2358 supra and separately subgingival plaque samples were collected from the mesial aspect of all teeth excluding 3rd molars in each subject. Samples were examined for the presence and levels of 40 bacterial taxa using whole genomic DNA probes and checkerboard DNA-DNA hybridization. Clinical assessments including dichotomous measures of gingival redness, bleeding on probing, plaque accumulation and suppuration, as well as duplicate measures of pocket depth and attachment level, were made at 6 sites per tooth. Mean counts (x10(5), % DNA probe count and % sites colonized for each species were determined separately for supra and subgingival samples in each subject and then averaged across subjects in the 2 clinical groups. Significance of differences between healthy and periodontitis subjects was determined using the Mann-Whitney test and adjusted for multiple comparisons. RESULTS: Mean total DNA probe counts (x10(5), +/-SEM) for healthy and periodontitis subjects in supragingival plaque were 72.1+/-11 and 132+/-17.5, respectively (p<0.01), and in subgingival plaque 22.1+/-6.6 and 100.3+/-18.4, (p<0.001). Porphyromonas gingivalis, Bacteroides forsythus and Treponema denticola could be detected in supragingival plaque samples of both healthy and periodontitis subjects. Actinomyces species were the dominant taxa in both supra- and subgingival plaque from healthy and periodontitis subjects. 4 Actinomyces species accounted for 63.2%, of supragingival and 47.2% of subgingival plaque in healthy subjects and 48.% and 37.8% in periodontitis subjects respectively. Increased proportions of P. gingivalis, B. forsythus, and species of Prevotella, Fusobacterium, Campylobacter and Treponema were detected subgingivally in the periodontitis subjects. P. gingivalis, B. forsythus and T. denticola were significantly more prevalent in both supra- and subgingival plaque samples from periodontitis subjects. CONCLUSIONS: The main differences between supra and subgingival plaque as well as between health and disease were in the proportions and to some extent levels of Actinomyces, "orange" and "red" complex species.     

Occurrence of Porphyromonas gingivalis and Tannerella forsythensis in periodontally diseased and healthy subjects

BACKGROUND: The purpose of this study was to compare the prevalence and level of Porphyromonas gingivalis (P. gingivalis) and Tannerella forsythensis (T. forsythensis) in subgingival plaque samples from both healthy individuals and periodontal patients in different age groups. METHODS: A total of 498 subgingival plaque samples were studied. These samples were collected from 407 individuals diagnosed with periodontal disease (210 adult periodontitis [AP], 78 rapidly progressive periodontitis [RPP], and 119 refractory periodontitis [Ref-P] cases) and 91 healthy (H) subjects. P. gingivalis and T. forsythensis were detected by indirect immunofluorescent assay using species-specific polyclonal antisera to P. gingivalis strain (FDC 381) and T. forsythensis strain (FDC 335). The prevalence of P. gingivalis and T. forsythensis was compared by chi-square analysis. Differences in P. gingivalis and T. forsythensis levels among various periodontal status and age groups was determined by one-way analysis of variance and Fisher's multiple comparison tests. The association between the presence of P. gingivalis or T. forsythensis in different periodontal status and age groups was measured using odds ratios. RESULTS: P. gingivalis was found in 85.7% (P < 0.0001) and T. forsythensis in 60.7% (P = 0.0002) of diseased subjects compared to 23.1% and 39.6%, respectively, in healthy subjects. P. gingivalis, but not T. forsythensis, was detected more frequently in any diseased group than in the H group in every age group (P<0.0001). No significant difference was found in the prevalence of P. gingivalis and T. forsythensis among age groups, except T. forsythensis was more prevalent in the age group of 40 to 59 years than in the age group < 20 years (chi2 = 3.93, P = 0.047) in the H group. The mean level of P. gingivalis and T. forsythensis was significantly higher in diseased groups than in the H group (P < 0.0001). The odds ratio for P. gingivalis in the AP group (25.0) was greater than any other group for P. gingivalis or T. forsythensis compared to the H group. CONCLUSIONS: These data suggested that P. gingivalis is closely associated with the pathogenesis of periodontitis and that it may not be a normal inhabitant of periodontally healthy subjects. T. forsythensis is also important in the pathogenesis of periodontal disease; however, whether it causes periodontal disease or is a secondary invader of periodontal lesions remains to be determined.     

Genotypic characterization of Porphyromonas gingivalis isolated from subgingival plaque and blood sample in positive bacteremia subjects with periodontitis

Aim: The objective of this study was to investigate clonal relationship among Porphyromonas gingivalis isolated from subgingival plaque and blood samples in positive transient bacteremia subjects with periodontitis.
Material and Methods: Unrelated patients with general chronic periodontitis or general aggressive periodontitis requiring scaling and root planing (SRP) were included in the study. Genotyping of each isolate was performed using pulsed field gel electrophoresis technique. Genetic relatedness of strains isolated within an individual or between different patients was determined by dendogram analysis.
Results: Following SRP, from 16 patients, seven patients showed positive P. gingivalis bacteremia and nine were negative. Thirty-two strains were isolated from subgingival plaque and blood samples before and during induced transient bacteremia. The majority of the patients harboured one clonal type. Two patients showed different clones in plaque and blood samples suggesting that more than one clone can be found in subgingival plaque. P. gingivalis isolates from periodontitis patients after transient bacteremia following SRP, revealed a high heterogeneity among isolates.
Conclusion: In 6/16 subjects the same P. gingivalis isolate was found in the blood and in oral cavity. P. gingivalis heterogeneity suggests no association of a unique clonal type with transient bacteremia.     

成人牙周健康状况与fimA基因型牙龈卟啉单胞菌的相关性

目的分析不同fimA基因型牙龈卟啉单胞菌（P.gingivalis）在牙周健康人群和慢性牙周炎人群中的分布,探讨不同fimA基因型P.gingivalis与成人牙周状况的相关关系。方法收集牙周健康组（136例）和慢性牙周炎组（115例）的龈下菌斑样本,采用16S rRNA PCR法检测P.gingivalis,并根据各fimA基因型（Ⅰ～Ⅴ和Ⅰb）的特异性引物检测不同fimA基因型P.gingivalis菌株的分布,计算OR值和95%可信区间。结果牙周健康组和慢性牙周炎组龈下菌斑样本中P.gingivalis阳性率分别为22.1%和81.7%,多数样本中只检测到1种fimA基因型。牙周健康组中ⅠfimA型的检出率最高（占66.7%）;慢性牙周炎组中则为ⅡfimA基因型（占43.6%）,其次为Ⅳ和Ⅰb fimA基因型。慢性牙周炎的发生与P.gingivalis的关系密切（OR=16.36）,Ⅰ、Ⅰb、Ⅱ、Ⅲ、Ⅳ、ⅤfimA基因型P.gingivalis与慢性牙周炎相关性的OR值分别为0.97、13.26、36.62、4.57、22.86、1.19;ⅡfimA基因型P.gingivalis与慢性牙周炎的相关性最强,其次为Ⅳ和Ⅰb型。结论P.gingivalis菌株的fimA基因型存在差异,特异性fimA基因型P.gingivalis可能与成人慢性牙周炎的发生关系密切。     

牙龈卟啉单胞菌PG0717基因与慢性牙周炎的相关性研究

目的检测慢性牙周炎患者和牙周健康者龈下菌斑中牙龈卟啉单胞菌（P.gingivalis）PG0717基因,探讨PG0717基因与牙周临床指数之间的关系。方法选取慢性牙周炎（CP）患者90例和牙周健康者90例,共采集龈下菌斑标本540个;记录临床牙周指数（牙周探诊深度、临床附着丧失和探诊出血）;设计特异性引物检测P.gingivalis阳性龈下菌斑标本的PG0717基因。结果在P.gingivalis阳性龈下菌斑中,CP组PG0717基因检出率显著高于对照组,分别为56.22%和41.27%（掊2=4.50,P＜0.05）;随着牙周探诊深度、临床附着丧失加重和探诊出血趋势的增加,CP组该基因检出率呈现增高趋势。结论PG0717基因与P.gingivalis的致病性有关。     

Description of the subgingival microbiota of periodontally untreated Mexican subjects: chronic periodontitis and periodontal health

BACKGROUND: Recent studies have suggested that changes in the prevalence and/or proportion of distinct microorganisms characterize the subgingival microbial profiles of populations around the world. At present, no information is available on the subgingival microbiota of Mexican subjects. The purpose of the present study was to determine the microbial composition of subgingival plaque in Mexican subjects with untreated chronic periodontitis. METHODS: A total of 44 chronic periodontitis and 20 periodontally healthy subjects (who were currently non-smokers) were selected. Clinical measurements including plaque accumulation, gingival erythema, bleeding on probing, suppuration, probing depth, and attachment level were recorded at six sites of every tooth. Up to 28 subgingival plaque samples were obtained from each subject and individually analyzed to determine the levels, proportion, and prevalence of 40 microbial species using the checkerboard DNA-DNA hybridization technique. RESULTS: Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythensis were the only species that presented higher mean levels in periodontitis subjects. The proportions of P. gingivalis (P<0.001), T. forsythensis (P<0.01), and red complex species (P. gingivalis, T. forsythensis, and T. denticola; P<0.001) as a group were also significantly higher in periodontitis subjects. Periodontally healthy subjects harbored a significantly larger proportion of Actinomyces species (P<0.05). No significant differences were detected in the percentage of carriers of any of the species tested. CONCLUSIONS: Our results revealed that the subgingival microbiota of untreated chronic periodontitis Mexican subjects was characterized by increases in the level, prevalence, and proportion of classic periodontal pathogens. However, the prevalence and proportion of specific microbial species varied significantly from the results of other reports on subjects from different geographical locations.