RESUMEN
Porphyromonas gingivalis has been extensively associated with both the onset and progression of periodontitis. We previously isolated and characterized two P. gingivalis strains, one from a patient exhibiting severe chronic periodontitis (CP3) and another from a periodontally healthy individual (H3). We previously showed that CP3 and H3 exhibit differences in virulence since H3 showed a lower resistance to cationic peptides compared with CP3, and a lower ability to induce proliferation in gingival epithelial cells. Here, we aimed to determine whether differences in virulence between these two strains are associated with the presence or absence of specific genes encoding virulence factors. We sequenced the whole genomes of both P. gingivalis CP3 and H3 and conducted a comparative analysis regarding P. gingivalis virulence genetic determinants. To do so, we performed a homology search of predicted protein sequences in CP3 and H3 genomes against the most characterized virulence genes for P. gingivalis available in the literature. In addition, we performed a genomic comparison of CP3 and H3 with all the 62 genomes of P. gingivalis found in NCBI's RefSeq database. This approach allowed us to determine the evolutionary relationships of CP3 and H3 with other virulent and avirulent strains; and additionally, to detect variability in presence/absence of virulence genes among P. gingivalis genomes. Our results show genetic variability in the hemagglutinin genes. While CP3 possesses one copy of hagA and two of hagC, H3 has no hagA and only one copy of hagC. Experimentally, this finding is related to lower in vitro hemmaglutination ability of H3 compared to CP3. Moreover, while CP3 encodes a gene for a major fimbrium subunit FimA type 4 (CP3_00160), H3 possess a FimA type 1 (H3_01400). Such genetic differences are in agreement with both lower biofilm formation ability and less intracellular invasion to oral epithelial cells exhibited by H3, compared with the virulent strain CP3. Therefore, here we provide new results on the genome sequences, comparative genomics analyses, and phenotypic analyses of two P. gingivalis strains. The genomics comparison of these two strains with the other 62 genomes included in the analysis provided relevant results regarding genetic determinants and their association with P. gingivalis virulence.
Asunto(s)
Periodontitis Crónica/patología , Regulación Bacteriana de la Expresión Génica , Genoma Bacteriano , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/patogenicidad , Factores de Virulencia/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biopelículas/crecimiento & desarrollo , Estudios de Casos y Controles , Línea Celular , Periodontitis Crónica/microbiología , Células Epiteliales/microbiología , Proteínas Fimbrias/genética , Proteínas Fimbrias/metabolismo , Ontología de Genes , Variación Genética , Genómica , Encía/microbiología , Humanos , Lectinas/genética , Lectinas/metabolismo , Anotación de Secuencia Molecular , Fenotipo , Filogenia , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Análisis de Secuencia de ADN , Virulencia , Factores de Virulencia/metabolismoRESUMEN
The aim of this study was to evaluate the relationship among nutritional status, gingival health and the composition of oral microbiota in children of a public school from a very poor area of San Miguel de Tucuman. Forty-five children ranging in age from 6 to 14 years old, 13 males and 32 females were studied. Twenty of these children were undernourished (Lejarraga-Morasso Table) and twenty-five were eutrophic. A clinical study that included DMF and dmf indexes, Löe Silness Plaque Index and bleeding on probing was performed. For microbiological study, saliva samples without stimulation were taken; aliquots of them were immediately placed in TAE buffer pH 7.6, adding NaOH (N and keeping at -70 °C until processed by checkerboard DNA-DNA hybridization method to check the presence of 40 oral microorganism species. Positive bleeding on probing was present in more than 80% of children, without significant differences between eutrophic and undernourished groups. Same result were obtain for the other clinical indexes (p > 0.05, Two Way ANOVA). Significant differences were found for some oral microorganism species, with a higher percentage of undernourished children harboring them. That was the case of S. gordonii (p < 0.05), Capnocitophaga gingivalis and C. ochraceae (p < 0.01 and p < 0.10, respectively), F. nucleatum ss nucleatum (p < 0.05), P. nigrescens (p < 0.10), Campylobacter gracilis (p < 0,05), and T. denticola (p < 0.10, multiple logistic regression). Significant differences were also found between children groups for E. saborreum (p < 0.001), P. acnes (p < 0.10), G. morbillorum (p < 0.05) and L. buccalis (p < 0.10). Gingivitis and bleeding on probing would not be related to nutritional status in the groups of children studied. There were significant differences for the presence of some of the main periodontal pathogen species between eutrophic and undernourished children. It would be important to study the meaning of significant differences found for the other microorganisms more deeply.
Asunto(s)
ADN Bacteriano/genética , Encía/microbiología , Gingivitis/microbiología , Desnutrición/microbiología , Microbiota/genética , Adolescente , Aggregatibacter actinomycetemcomitans/clasificación , Aggregatibacter actinomycetemcomitans/genética , Aggregatibacter actinomycetemcomitans/aislamiento & purificación , Argentina , Bacteroides/clasificación , Bacteroides/genética , Bacteroides/aislamiento & purificación , Campylobacter/clasificación , Campylobacter/genética , Campylobacter/aislamiento & purificación , Capnocytophaga/clasificación , Capnocytophaga/genética , Capnocytophaga/aislamiento & purificación , Estudios de Casos y Controles , Niño , Femenino , Fusobacterium nucleatum/clasificación , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/aislamiento & purificación , Gingivitis/fisiopatología , Humanos , Masculino , Desnutrición/fisiopatología , Hibridación de Ácido Nucleico , Peptostreptococcus/clasificación , Peptostreptococcus/genética , Peptostreptococcus/aislamiento & purificación , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/aislamiento & purificación , Saliva/microbiologíaRESUMEN
It is well known that strain and virulence diversity exist within the population structure of Porphyromonas gingivalis. In the present study we investigate intra- and inter-species variability in biofilm formation of Porphyromonas gingivalis and partners Prevotella intermedia and Prevotella nigrescens. All strains tested showed similar hydrophobicity, except for P. gingivalis W83 which has roughly half of the hydrophobicity of P. gingivalis ATCC33277. An intraspecies variability in coaggregation of P. gingivalis with P. intermedia was also found. The association P. gingivalis W83/P. intermedia 17 produced the thickest biofilm and strain 17 was prevalent. In a two-compartment system P. gingivalis W83 stimulates an increase in biomass of strain 17 and the latter did not stimulate the growth of P. gingivalis W83. In addition, P. gingivalis W83 also stimulates the growth of P. intermedia ATCC25611 although strain W83 was prevalent in the association with P. intermedia ATCC25611. P. gingivalis ATCC33277 was prevalent in both associations with P. intermedia and both strains of P. intermedia stimulate the growth of P. gingivalis ATCC33277. FISH images also showed variability in biofilm structure. Thus, the outcome of the association P. gingivalis/P. intermedia seems to be strain-dependent, and both soluble factors and physical contact are relevant. The association P. gingivalis-P. nigrescens ATCC33563 produced larger biomass than each monotypic biofilm, and P. gingivalis was favored in consortia, while no differences were found in the two-compartment system. Therefore, in consortia P. gingivalis-P. nigrescens physical contact seems to favor P. gingivalis growth. The intraspecies variability found in our study suggests strain-dependence in ability of microorganisms to recognize molecules in other bacteria which may further elucidate the dysbiosis event during periodontitis development giving additional explanation for periodontal bacteria, such as P. gingivalis and P. intermedia, among others, to persist and establish chronic infections in the host.
Asunto(s)
Biopelículas/crecimiento & desarrollo , Porphyromonas gingivalis/fisiología , Prevotella intermedia/fisiología , Adhesión Bacteriana , Biomasa , Disbiosis/microbiología , Humanos , Periodontitis/microbiología , Porphyromonas gingivalis/clasificación , Prevotella intermedia/clasificación , Prevotella intermedia/genéticaRESUMEN
PURPOSES: The primary aim of this cross-sectional study was to compare the levels of red complex bacteria between Afro-Brazilian and non Afro-Brazilian cohort. The secondary aim was to compare the distribution of both Aggregatibacter actinomycetemcomitans serotype b and its JP2 strains among participants who harboured this bacterial species. METHODS: A total of 84 individuals were included in this study: 42 Afro-descendants (mean age 35.9 ± 13.1 years) and 42 non-Afro-descendants (mean age 36.2 ± 13.1 years) matched (1:1) by periodontal diagnosis, age and gender. All participants received clinical examinations of periodontal pocket depth, clinical attachment level, and plaque and gingival indices. Subgingival samples were taken for microbial analysis. First, genomic DNA (gDNA) was extracted and purified and the quantification of total number of bacterial cells, A. actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia and Treponema denticola was carried out by qPCR. Then, A. actinomycetemcomitans strains were classified according to serotype b and JP2 profiles by conventional PCR. RESULTS: Clinically, mean PD, mean CAL and percentage of CAL ≥ 3 mm differed between groups (Student's t-test p<0.05). The levels of red complex bacteria between Afro-Brazilian and non-Afro-Brazilian populations were similar. The exception was verified to A. actinomycetemcomitans showing significantly higher levels among Afro-Brazilian descendants in comparison to non-Afro-Brazilian descendants. Afro-Brazilian descendants were clearly infected by more virulent serotype b and JP2 strains. CONCLUSIONS: Despite no statistically significant differences related to the red complex species, Afro-Brazilian descendants harboured higher levels of A. actinomycetemcomitans. Also, our findings confirm that Afro-descendant populations are preferably colonised by A. actinomycetemcomitans serotype b as well as JP2 strains.
Asunto(s)
Aggregatibacter actinomycetemcomitans/clasificación , Bacteroides/clasificación , Periodontitis/microbiología , Adulto , Población Negra , Brasil , Estudios Transversales , Femenino , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/clasificación , Serotipificación , Treponema denticola/clasificaciónRESUMEN
BACKGROUND: Different serotypes of Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis have been shown to induce differential dendritic cell (DC) responses. This study investigates whether cytokine and CC-chemokine receptor (CCR) production by DCs stimulated with different serotypes of A. actinomycetemcomitans or P. gingivalis is Toll-like receptor 2 (TLR2) and/or TLR4 dependent. METHODS: DCs were obtained from healthy individuals and primed at a multiplicity of infection (MOI) of 10(2) with different A. actinomycetemcomitans or P. gingivalis serotypes in the presence or absence of anti-TLR2 or anti-TLR4 blocking antibodies. TLR2 and TLR4 expression, CCR5 and CCR6 expression, and interleukin (IL)-1ß, IL-10, IL-12, and IL-23 expression and secretion were quantified by flow cytometry, real-time reverse-transcription polymerase chain reaction, and enzyme-linked immunosorbent assay. RESULTS: When DCs were stimulated with serotype b of A. actinomycetemcomitans or serotype K1 of P. gingivalis, higher levels of TLR2 or TLR4, respectively, were detected compared to DCs stimulated with the other serotypes. Similarly, higher levels of cytokines and CCRs were detected in serotype b- or serotype K1-primed DCs compared to the others, and these increased levels positively correlated with levels of TLR2 or TLR4. When TLR2 signaling was blocked using a specific anti-TLR2 monoclonal antibody, serotype b-induced cytokine and CCR expression was inhibited; when TLR4 signaling was blocked, serotype K1-induced response was inhibited. CONCLUSIONS: These results demonstrate that the variability of secretion of cytokines and expression of CCRs detected in DCs stimulated with different serotypes of A. actinomycetemcomitans or P. gingivalis is TLR2 or TLR4 dependent, respectively.
Asunto(s)
Aggregatibacter actinomycetemcomitans/inmunología , Células Dendríticas/microbiología , Porphyromonas gingivalis/inmunología , Receptor Toll-Like 2/inmunología , Receptor Toll-Like 4/inmunología , Adulto , Aggregatibacter actinomycetemcomitans/clasificación , Carga Bacteriana , Técnicas de Cultivo de Célula , Diferenciación Celular/fisiología , Células Cultivadas , Células Dendríticas/inmunología , Femenino , Humanos , Interleucina-10/análisis , Interleucina-12/análisis , Interleucina-1beta/análisis , Interleucina-23/análisis , Masculino , Monocitos/fisiología , Porphyromonas gingivalis/clasificación , Receptores CCR5/análisis , Receptores CCR6/análisis , Serogrupo , Receptor Toll-Like 2/análisis , Receptor Toll-Like 4/análisis , Adulto JovenRESUMEN
AIMS: To evaluate the association among serum immunoglobulin G (IgG) responses to Aggregatibacter actinomycetemcomitans (Aa) serotypes a, b and c, Porphyromonas gingivalis (Pg), Tannerella forsythia (Tf) and clinical parameters in Aggressive Periodontitis (AP) subjects. Associations between periodontal pathogens and clinical and immunological parameters were also evaluated. METHODS: Thirty-eight subjects diagnosed with generalized AP (GAP) and localized AP (LAP) were included. Ten healthy controls were also evaluated. Clinical parameters were assessed and percentages of subgingival levels of Aa, Pg and Tf (beyond bacterial load), were determined by quantitative real-time polymerase chain reaction. Serum IgG antibody levels against Aa, Pg and Tf were evaluated by enzyme-linked immunosorbent assay. RESULTS: Percentages of Aa, Pg and Tf were significantly higher in AP than in controls. The response to Aa serotype c was higher in LAP subjects than in controls. There were no differences in microbial composition or antibodies responses between GAP and LAP, except for IgG response to Tf. Pg levels were correlated with probing depth (PD), BoP and CAL in GAP but not in LAP subjects. Tf levels correlated with PD and CAL in GAP subjects. In GAP, the infection levels of Aa and Pg correlated with the corresponding IgG levels to Aa serotype c and Pg. CONCLUSION: Given the evidences that IgG response in AP patients correlated with bacterial infection level in GAP, but not in LAP, and that LAP patients lack a response to Tf, despite harbouring this species, our data suggest a difference in host immune defence between these two forms of aggressive periodontitis.
Asunto(s)
Periodontitis Agresiva/microbiología , Anticuerpos Antibacterianos/sangre , Bacterias Gramnegativas/inmunología , Inmunoglobulina G/sangre , Adulto , Aggregatibacter actinomycetemcomitans/clasificación , Aggregatibacter actinomycetemcomitans/inmunología , Periodontitis Agresiva/clasificación , Periodontitis Agresiva/inmunología , Pérdida de Hueso Alveolar/diagnóstico por imagen , Pérdida de Hueso Alveolar/microbiología , Carga Bacteriana , Bacteroides/clasificación , Bacteroides/inmunología , Estudios Transversales , Femenino , Interacciones Huésped-Patógeno/inmunología , Humanos , Masculino , Pérdida de la Inserción Periodontal/inmunología , Pérdida de la Inserción Periodontal/microbiología , Bolsa Periodontal/inmunología , Bolsa Periodontal/microbiología , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/inmunología , Radiografía , Serogrupo , Adulto JovenRESUMEN
AIM: Destructive periodontitis is associated with a Th1-Th17 immune response and activation of RANKL-induced osteoclasts. In addition, Porphyromonas gingivalis K1 and K2 serotypes induce a strong Th1-Th17 response. This study aimed to investigate whether these P. gingivalis serotypes induce higher osteoclasts activation, by increased Th17-associated RANKL production, and an antigen-specific memory T-lymphocyte response. MATERIAL AND METHODS: The RANKL production and TRAP(+) osteoclast induction were quantified on naïve T lymphocytes stimulated with dendritic cells primed with the P. gingivalis serotypes. The T-bet, GATA-3, RORC2 and Foxp3 expression was correlated with RANKL production. The frequency of proliferating memory T lymphocytes in response to P. gingivalis serotypes was determined in both periodontitis and healthy subjects. RESULTS: T lymphocytes stimulated by K1 or K2-primed dendritic cells elicited higher levels of RANKL and TRAP(+) osteoclasts than cells stimulated with the other serotypes. RANKL positively correlated with RORC2. Whereas periodontitis patients had a higher frequency of memory T lymphocytes responding to K1 or K2, healthy subjects had a higher frequency of memory T lymphocytes responding to K4 or K(-) . CONCLUSIONS: P. gingivalis serotypes K1 and K2, but not others, are associated with an increased production of the osteoclastogenesis-related factor RANKL. This important information suggests that these serotypes could elicit a greater bone resorption in vivo and have a role in the periodontitis pathogenesis.
Asunto(s)
Memoria Inmunológica/inmunología , Osteoclastos/inmunología , Porphyromonas gingivalis/inmunología , Ligando RANK/inmunología , Serogrupo , Linfocitos T/inmunología , Fosfatasa Ácida/análisis , Fosfatasa Ácida/inmunología , Animales , Antígenos Bacterianos/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/microbiología , Diferenciación Celular/inmunología , Línea Celular , Periodontitis Crónica/inmunología , Selección Clonal Mediada por Antígenos , Células Dendríticas/inmunología , Factores de Transcripción Forkhead/análisis , Factor de Transcripción GATA3/análisis , Humanos , Isoenzimas/análisis , Isoenzimas/inmunología , Macrófagos/inmunología , Ratones , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/análisis , Osteoclastos/efectos de los fármacos , Porphyromonas gingivalis/clasificación , Ligando RANK/análisis , Proteínas de Dominio T Box/análisis , Linfocitos T/microbiología , Fosfatasa Ácida Tartratorresistente , Células TH1/inmunología , Células Th17/inmunologíaRESUMEN
AIM: Porphyromonas gingivalis can synthesize an extracellular capsule and different serotypes have been described based on capsular antigenicity. On dendritic cells (DCs), the type of capsule present plays a role on the strength of the developed immune response. This study aimed to investigate the T-lymphocyte responses when stimulated with autologous mature DCs exposed to different P. gingivalis K-serotypes. MATERIALS AND METHODS: Naïve CD4(+) T-lymphocytes were obtained from healthy subjects and stimulated with autologous DCs primed with increasing multiplicity of infections of the different P. gingivalis K-serotypes. The Th1, Th2, Th17 and T-regulatory cytokines and transcription factor levels were quantified. RESULTS: Distinct types of response were detected when T-lymphocytes were stimulated by DCs primed with the different P. gingivalis K-serotypes. T-lymphocytes stimulated by K1 or K2-primed DCs elicited higher levels of Th1 and Th17-associated cytokines, T-bet and RORC2 than T-lymphocytes stimulated with DCs primed with the other serotypes. Conversely, the serotypes K3-K5 induced higher levels of Th2-associated cytokines and GATA-3 than the others. CONCLUSIONS: These results demonstrate that DCs primed with the different P. gingivalis K-serotypes elicited distinct T-cell responses. Strains K1 (W83) and K2 (HG184) induced a Th1/Th17 pattern of immune response and K3 (A7A1-28), K4 (ATCC(®49417™) ), and K5 (HG1690) a Th2 response.
Asunto(s)
Cápsulas Bacterianas/inmunología , Células Dendríticas/microbiología , Porphyromonas gingivalis/inmunología , Linfocitos T/inmunología , Técnicas Bacteriológicas , Citocinas/análisis , Células Dendríticas/inmunología , Factores de Transcripción Forkhead/análisis , Factor de Transcripción GATA3/análisis , Humanos , Interferón gamma/análisis , Subunidad alfa del Receptor de Interleucina-2/análisis , Interleucinas/análisis , Activación de Linfocitos/inmunología , Monocitos/inmunología , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/análisis , Porphyromonas gingivalis/clasificación , Serotipificación , Proteínas de Dominio T Box/análisis , Linfocitos T/microbiología , Linfocitos T Reguladores/inmunología , Células TH1/inmunología , Células Th17/inmunología , Células Th2/inmunología , Factor de Crecimiento Transformador beta1/análisis , Factores de Necrosis Tumoral/análisisRESUMEN
Peri-implant inflammation contributes for loss of secondary stability of orthodontic mini-implants. The investigation of microbial colonization in this area would benefit its control, and consequently favor the long-term success of mini-implants. Therefore, the aim of this study was to determine the establishment and the evolution of microbial colonization process in orthodontic mini-implants for 3 months, since the time of their installation. One-hundred and fifty samples collected from 15 mini-implants were investigated from baseline up to 3 months. The biological material was obtained from peri-implant area using paper points. Nonspecific, Streptococcus spp, Lactobacillus casei and Candida spp colonizations were analyzed by cell growth methods. Porphyromonas gingivalis colonization was observed by 16S rDNA-directed polymerase chain reaction. Data from cell growth were submitted to the Wilcoxon sign rank test and results from molecular analysis were presented in a descriptive way. There was no significant difference in the microbial colonization among the examined time intervals, except for Streptococcus spp, between baseline and 24 h, which characterized the initial colonization in this time interval. Lactobacillus casei and Candida spp colonizations were insignificant. No Porphyromonas gingivalis was detected among the analyzed samples. The microbial colonization of mini-implants did not significantly change during the study. However, it should be monitored by orthodontists, since it is an important factor for mini-implants success.
Asunto(s)
Bacterias/crecimiento & desarrollo , Implantes Dentales/microbiología , Métodos de Anclaje en Ortodoncia/instrumentación , Adolescente , Aleaciones , Antiinfecciosos Locales/uso terapéutico , Bacterias/clasificación , Carga Bacteriana , Técnicas Bacteriológicas , Candida/crecimiento & desarrollo , Clorhexidina/uso terapéutico , Aleaciones Dentales/química , Femenino , Estudios de Seguimiento , Humanos , Lacticaseibacillus casei/crecimiento & desarrollo , Masculino , Antisépticos Bucales/uso terapéutico , Higiene Bucal/educación , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/crecimiento & desarrollo , ARN Bacteriano/análisis , ARN Ribosómico 16S/análisis , Streptococcus/clasificación , Streptococcus/crecimiento & desarrollo , Titanio/química , Técnicas de Movimiento Dental/instrumentación , Cepillado Dental/métodos , Adulto JovenRESUMEN
Las periodontitis son un conjunto de patologías de naturaleza inflamatoria y etiología infecciosa producidas por el biofilm patogénico subgingival. Porphyromonas gingivalis y Aggregatibacter actinomycetemcomitans son bacterias periodonto-patógenas que pueden causar daño directo a las estructuras periodontales a través de los diversos factores de virulencia que expresan. Sobre la base de estos factores de virulencia, distintos genotipos y serotipos bacterianos se han descrito, cada uno de ellos con una potencial variable patogenicidad. En esta revisión bibliográfica se describen diferentes factores de virulencia de P. gingivalis y A. actinomycetemcomitans y se discute la variable inmunogenicidad y patogenicidad de los distintos genotipos y serotipos descritos para ellos. Tanto P. gingivalis como A. actinomycetemcomitans poseen diversos factores de virulencia asociados al inicio, progresión y severidad de las periodontitis. En P. gingivalis, los factores de virulencia para los cuales se describen distintos genotipos y/o serotipos son fimbria, LPS y cápsula bacteriana, y en A. actinomycetemcomitans son leucotoxina A, Cdt y LPS. Cada uno de estos distintos genotipos y serotipos induce una respuesta inmuno-inflamatoria diferente en el hospedero y, por lo tanto, se podrían asociar a una variable patogenicidad y podrían determinar las características clínicas de la enfermedad.
Periodontitis represents a heterogenic group of periodontal infections elicited by bacteria residing at the subgingival biofilm. Although this biofilm is constituted by a broad variety of bacterial species, only a limited number has been associated with the periodontitis aetiology, among them Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. Both P. gingivalis and A. actinomycetemcomitans express a number of virulence factors that contribute to direct tissue damage and, based on them, distinct genotypes and serotypes have been described, each one with a potential variable pathogenicity. This review aimed to analyze the different virulence factors described for P. gingivalis and A. actinomycetemcomitans and to discuss the variable immunogenicity and pathogenicity of their serotypes and genotypes. P. gingivalis and A. actinomycetemcomitans express different virulence factors and they determine the initiation, progression, and severity of periodontitis. In P. gingivalis, distinct serotypes and/or genotypes are described based on fimbriae, LPS, and capsule. Additionally, in A. actinomycetemcomitans distinct serotypes and/or genotypes are described based on leucotoxin A, Cdt, and LPS. These distinct serotypes and genotypes induce a differential immunoinflammatory response and, thus, could be associated with variations in pathogenicity and reflected in clinic characteristics of the disease.
Asunto(s)
Aggregatibacter actinomycetemcomitans/patogenicidad , Periodontitis/microbiología , Porphyromonas gingivalis/patogenicidad , Aggregatibacter actinomycetemcomitans/clasificación , Fimbrias Bacterianas , Genotipo , Lipopolisacáridos , Péptido Hidrolasas , Porphyromonas gingivalis/clasificación , Serotipificación , Factores de VirulenciaRESUMEN
Peri-implant inflammation contributes for loss of secondary stability of orthodontic mini-implants. The investigation of microbial colonization in this area would benefit its control, and consequently favor the long-term success of mini-implants. Therefore, the aim of this study was to determine the establishment and the evolution of microbial colonization process in orthodontic mini-implants for 3 months, since the time of their installation. One-hundred and fifty samples collected from 15 mini-implants were investigated from baseline up to 3 months. The biological material was obtained from peri-implant area using paper points. Nonspecific, Streptococcus spp, Lactobacillus casei and Candida spp colonizations were analyzed by cell growth methods. Porphyromonas gingivalis colonization was observed by 16S rDNA-directed polymerase chain reaction. Data from cell growth were submitted to the Wilcoxon sign rank test and results from molecular analysis were presented in a descriptive way. There was no significant difference in the microbial colonization among the examined time intervals, except for Streptococcus spp, between baseline and 24 h, which characterized the initial colonization in this time interval. Lactobacillus casei and Candida spp colonizations were insignificant. No Porphyromonas gingivalis was detected among the analyzed samples. The microbial colonization of mini-implants did not significantly change during the study. However, it should be monitored by orthodontists, since it is an important factor for mini-implants success.
A inflamação peri-implantar contribui para a perda da estabilidade secundária dos mini-implantes ortodônticos. A investigação da colonização microbiana desta área beneficiaria o seu controle e, consequentemente, favoreceria o sucesso dos mini-implantes a longo prazo. Portanto, o objetivo dos autores foi determinar o estabelecimento e evolução do processo de colonização microbiana em mini-implantes ortodônticos por três meses desde a instalação. Cento e cinquenta amostras coletadas de 15 mini-implantes foram investigadas desde o tempo inicial até 3 meses. O material biológico foi obtido da área peri-implantar com auxílio de cones de papel absorvente. As colonizações inespecíficas de Streptococcus spp, Lactobacillus casei e Candida spp foram analisadas por métodos de crescimento celular. A colonização por Porphyromonas gingivalis foi observada por meio da reação em cadeia da polimerase 16S rDNA. Os dados do crescimento celular foram submetidos ao teste de Wilcoxon sign rank e os resultados da biologia molecular foram apresentados de modo descritivo. Não houve diferença estatisticamente significante da colonização microbiana entre os intervalos de tempo avaliados, exceto para Streptococcus spp entre os tempos inicial e 24 h, o que caracterizou o início da colonização neste intervalo de tempo. As colonizações por Lactobacillus casei e Candida spp foram insignificantes. Não foi detectada a presença de Porphyromonas gingivalis nas amostras analisadas. A colonização microbiana nos mini-implantes não se alterou significativamente durante o estudo. No entanto, deve ser monitorada por ortodontistas, uma vez que é um fator importante para o sucesso dos mini-implantes.
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Adolescente , Femenino , Humanos , Masculino , Adulto Joven , Bacterias/crecimiento & desarrollo , Implantes Dentales/microbiología , Métodos de Anclaje en Ortodoncia/instrumentación , Antiinfecciosos Locales/uso terapéutico , Carga Bacteriana , Técnicas Bacteriológicas , Bacterias/clasificación , Candida/crecimiento & desarrollo , Clorhexidina/uso terapéutico , Aleaciones Dentales/química , Estudios de Seguimiento , Lacticaseibacillus casei/crecimiento & desarrollo , Antisépticos Bucales/uso terapéutico , Higiene Bucal/educación , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/crecimiento & desarrollo , ARN Bacteriano/análisis , /análisis , Streptococcus/clasificación , Streptococcus/crecimiento & desarrollo , Titanio/química , Técnicas de Movimiento Dental/instrumentación , Cepillado Dental/métodosRESUMEN
OBJECTIVE: Long fimbriae (FimA) are important virulence factors of Porphyromonas gingivalis. Based on the diversity of the fimA gene, this species is classified into 6 genotypes. This study surveyed samples from primary endodontic infections for the presence of these P. gingivalis fimA variants. STUDY DESIGN: Genomic DNA isolated from samples taken from 25 root canals of teeth with chronic apical periodontitis and 25 aspirates from acute apical abscess was used as template in polymerase chain reaction (PCR) assays directed toward the detection of the different P. gingivalis fimA genotypes. RESULTS: Porphyromonas gingivalis was detected by a 16S rRNA gene-based PCR in 36% of the total number of cases sampled (44% of chronic apical periodontitis and 28% of abscess aspirates). In cases of chronic apical periodontitis, P. gingivalis variant type IV was the most prevalent (24%), followed by types I (20%), II (16%), and III (8%). In acute abscess samples, variant type II was the most prevalent (12%), followed by types III and IV (8% of each) and type I (4%). Combinations of up to 3 different genotypes were detected in a few cases. No single fimA genotype variant or combination thereof was significantly associated with symptoms. Overall, fimA types IV (16%), II (14%), and I (12%) were the most prevalent. CONCLUSIONS: Findings demonstrated that different P. gingivalis fimA genotypes can be present in primary endodontic infections.
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Infecciones por Bacteroidaceae/microbiología , Periodontitis Crónica/microbiología , Proteínas Fimbrias/genética , Absceso Periapical/microbiología , Periodontitis Periapical/microbiología , Porphyromonas gingivalis/genética , Variación Genética , Genotipo , Humanos , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/aislamiento & purificación , ARN Bacteriano/análisisRESUMEN
BACKGROUND: This study compared the subgingival microbiota of subjects with refractory periodontitis (RP) to those in subjects with treatable periodontitis (GRs = good responders) or periodontal health (PH) using the Human Oral Microbe Identification Microarray (HOMIM). METHODS: At baseline, subgingival plaque samples were taken from 47 subjects with periodontitis and 20 individuals with PH and analyzed for the presence of 300 species by HOMIM. The subjects with periodontitis were classified as having RP (n = 17) based on mean attachment loss (AL) and/or more than three sites with AL >or=2.5 mm after scaling and root planing, surgery, and systemically administered amoxicillin and metronidazole or as GRs (n = 30) based on mean attachment gain and no sites with AL >or=2.5 mm after treatment. Significant differences in taxa among the groups were sought using the Kruskal-Wallis and chi(2) tests. RESULTS: More species were detected in patients with disease (GR or RP) than in those without disease (PH). Subjects with RP were distinguished from GRs or those with PH by a significantly higher frequency of putative periodontal pathogens, such as Parvimonas micra (previously Peptostreptococcus micros or Micromonas micros), Campylobacter gracilis, Eubacterium nodatum, Selenomonas noxia, Tannerella forsythia (previously T. forsythensis), Porphyromonas gingivalis, Prevotella spp., Treponema spp., and Eikenella corrodens, as well as unusual species (Pseudoramibacter alactolyticus, TM7 spp. oral taxon [OT] 346/356, Bacteroidetes sp. OT 272/274, Solobacterium moorei, Desulfobulbus sp. OT 041, Brevundimonas diminuta, Sphaerocytophaga sp. OT 337, Shuttleworthia satelles, Filifactor alocis, Dialister invisus/pneumosintes, Granulicatella adiacens, Mogibacterium timidum, Veillonella atypica, Mycoplasma salivarium, Synergistes sp. cluster II, and Acidaminococcaceae [G-1] sp. OT 132/150/155/148/135) (P <0.05). Species that were more prevalent in subjects with PH than in patients with periodontitis included Actinomyces sp. OT 170, Actinomyces spp. cluster I, Capnocytophaga sputigena, Cardiobacterium hominis, Haemophilus parainfluenzae, Lautropia mirabilis, Propionibacterium propionicum, Rothia dentocariosa/mucilaginosa, and Streptococcus sanguinis (P <0.05). CONCLUSION: As determined by HOMIM, patients with RP presented a distinct microbial profile compared to patients in the GR and PH groups.
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Bacterias/clasificación , Periodontitis Crónica/microbiología , Periodontitis/microbiología , Periodoncio/microbiología , Adulto , Amoxicilina/uso terapéutico , Antibacterianos/uso terapéutico , Bacteroides/clasificación , Bacteroidetes/clasificación , Campylobacter/clasificación , Periodontitis Crónica/terapia , Placa Dental/microbiología , Raspado Dental , Eikenella corrodens/clasificación , Eubacterium/clasificación , Femenino , Bacterias Gramnegativas/clasificación , Bacterias Grampositivas/clasificación , Humanos , Masculino , Metronidazol/uso terapéutico , Análisis por Micromatrices , Persona de Mediana Edad , Peptostreptococcus/clasificación , Periodontitis/terapia , Porphyromonas gingivalis/clasificación , Prevotella/clasificación , Proteobacteria/clasificación , Aplanamiento de la Raíz , Selenomonas/clasificación , Treponema/clasificaciónRESUMEN
AIM: Capsular polysaccharides play an important role in the virulence of Gram-positive and Gram-negative bacteria. In Porphyromonas gingivalis, six serotypes have been described based on capsular antigenicity and its pathogenicity has been correlated both in vitro and in animal models. This study aimed to investigate the differential response of human dendritic cells (DCs) when stimulated with different P. gingivalis capsular serotypes. MATERIALS AND METHODS: Using different multiplicity of infection (MOI) of the encapsulated strains K1-K6 and the non-encapsulated K(-) strain of P. gingivalis, the mRNA expression levels for interleukin (IL)-1beta, IL-2, IL-5, IL-6, IL-10, IL-12, IL-13, interferon (IFN)-gamma, tumour necrosis factor (TNF)-alpha, and TNF-beta in stimulated DCs were quantified by real-time reverse transcription-polymerase chain reaction. RESULTS: All P. gingivalis capsular serotypes induced a T-helper type 1 (Th1) pattern of cytokine expression. K1- and K2-stimulated DCs expressed higher levels of IL-1beta, IL-6, IL-12p35, IL-12p40, and IFN-gamma and at lower MOI than DCs stimulated with the other strains. CONCLUSIONS: These results demonstrate a differential potential of P. gingivalis capsular serotypes to induce DC responses and a higher capacity of strains K1 W83 and K2 HG184 than other K serotypes to trigger cytokine expression.
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Cápsulas Bacterianas/inmunología , Citocinas/inmunología , Células Dendríticas/inmunología , Porphyromonas gingivalis/inmunología , Antígenos Bacterianos/inmunología , Cápsulas Bacterianas/clasificación , Células Cultivadas , Citocinas/análisis , Humanos , Interferón gamma/análisis , Interleucina-10/análisis , Interleucina-12/análisis , Subunidad p35 de la Interleucina-12/análisis , Subunidad p40 de la Interleucina-12/análisis , Interleucina-13/análisis , Interleucina-1beta/análisis , Interleucina-2/análisis , Interleucina-5/análisis , Interleucina-6/análisis , Linfotoxina-alfa/análisis , Polisacáridos Bacterianos/inmunología , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/patogenicidad , Serotipificación , Células TH1/inmunología , Factor de Necrosis Tumoral alfa/análisis , VirulenciaRESUMEN
INTRODUCTION: Porphyromonas gingivalis is considered as a major etiological agent in the onset and progression of chronic destructive periodontitis. Porphyromonus gingivalis fimA type has been correlated to the virulence potential of the strain; therefore this gene could be involved in the ability of P. gingivalis to reach blood stream. OBJECTIVE: The classifications of P. gingivalis fimA types will be compared in subgingival plaque and blood samples collected after scaling and root root planing of periodontitis patients. MATERIALS AND METHODS: Fifteen periodontitis patients requiring scaling and root planing were enrolled. P. gingivalis isolates were classed to genotype with fimA type-specific PCR assay. fimA gene was sequenced if the isolate was listed as unclassifiable after PCR technique. RESULTS: Six patients showed positive P. gingivalis bacteremia. The most frequent fimA was fimA type II, followed by Ib, III and IV. In blood strains, type II was followed by IV, Ib and III. CONCLUSION: Type II was the most frequent genotype in blood samples and in subgingival plaque samples. However, no correlation was found between the frequency of any fimA type with SRP induced bacteremia. P. gingivalis fimA type appears to be conserved within individual patients throughout the times of sample collection, fimA gene sequence results were not in agreement with results of fimA genotyping by PCR.
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Bacteriemia/microbiología , Placa Dental/microbiología , Proteínas Fimbrias/genética , Genes Bacterianos , Periodontitis/microbiología , Porphyromonas gingivalis/genética , Adulto , Bacteriemia/etiología , ADN Bacteriano/genética , Raspado Dental/efectos adversos , Genotipo , Humanos , Periodontitis/terapia , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/aislamiento & purificación , Aplanamiento de la Raíz , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Ácido Nucleico , Curetaje Subgingival/efectos adversosRESUMEN
BACKGROUND/AIMS: The aim of this study was to evaluate the composition of the microbiota of primary endodontic infections in 111 selected cases of single-rooted teeth with necrotic pulp. METHODS: Samples were collected from the root canals using #15 Hedströen-type files and two sterile paper points, which were introduced 1 mm short of the apical foramen. The presence, levels, and proportions of 40 different bacterial species in each sample were determined using DNA probes and checkerboard DNA-DNA hybridization techniques. RESULTS: The mean number of species per sample was 22. Enterococcus faecalis (89.3%), Campylobacter gracilis (89.3%), Leptotrichia buccalis (89.3%), Neisseria mucosa (87.5%), Prevotella melaninogenica (86.6%), Fusobacterium nucleatum ssp. vincentii (85.7%), Eubacterium saburreum (75.9%), Streptococcus anginosus (75%), and Veillonella parvula (74.1%) were the most prevalent species. The species found in highest mean counts (over 10(5)) were F. nucleatum ssp. vincentii (13.14 x 10(5)), E. saburreum (5.67 x 10(5)), E. faecalis (5.38 x 10(5)), N. mucosa (4.19 x 10(5)), V. parvula (3.63 x 10(5)), C. gracilis (3.46 x 10(5)), Treponema socranskii (3.34 x 10(5)), Porphyromonas endodontalis (2.96 x 10(5)), Porphyromonas gingivalis (2.85 x 10(5)), Micromonas micros (2.81 x 10(5)), Prevotella nigrescens (2.68 x 10(5)) and Fusobacterium nucleatum ssp. nucleatum (2.64 x 10(5)). Most of these species were also found in high proportions. CONCLUSIONS: Our results suggest that several bacterial species considered to be oral pathogens seem to be implicated in the etiology of primary endodontic infections.
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ADN Bacteriano/análisis , Necrosis de la Pulpa Dental/microbiología , Infecciones por Bacterias Gramnegativas/diagnóstico , Infecciones por Bacterias Grampositivas/diagnóstico , Hibridación de Ácido Nucleico/métodos , Adolescente , Adulto , Anciano , Campylobacter/clasificación , Recuento de Colonia Microbiana , Sondas de ADN , Cavidad Pulpar/microbiología , Enterococcus faecalis/clasificación , Eubacterium/clasificación , Femenino , Fusobacterium nucleatum/clasificación , Humanos , Leptotrichia/clasificación , Masculino , Persona de Mediana Edad , Neisseria mucosa/clasificación , Peptostreptococcus/clasificación , Porphyromonas endodontalis/clasificación , Porphyromonas gingivalis/clasificación , Prevotella melaninogenica/clasificación , Prevotella nigrescens/clasificación , Streptococcus anginosus/clasificación , Treponema/clasificación , Veillonella/clasificaciónRESUMEN
BACKGROUND/AIMS: The aim of this study was to examine the diversity of bacterial species in the infected root canals of teeth associated with endodontic abscesses by cloning and sequencing techniques in concert with denaturing high-performance liquid chromatography. METHODS: Samples collected from five infected root canals were subjected to polymerase chain reaction (PCR) with universal 16S ribosomal DNA primers. Products of these PCRs were cloned and sequenced. Denaturing high-performance liquid chromatography (DHPLC) was used as a screening method to reduce the number of clones necessary for DNA sequencing. RESULTS: All samples were positive for the presence of bacteria and a range of 7-13 different bacteria were found per root canal sample. In total, 48 different oral clones were detected among the five root canal samples. Olsenella profusa was the only species present in all samples. Porphyromonas gingivalis, Dialister pneumosintes, Dialister invisus, Lachnospiraceae oral clone, Staphylococcus aureus, Pseudoramibacter alactolyticus, Peptostreptococcus micros and Enterococcus faecalis were found in two of the five samples. The majority of the taxa were present in only one sample, for example Tannerella forsythia, Shuttleworthia satelles and Filifactor alocis. Some facultative anaerobes that are frequently isolated from endodontic infections such as E. faecalis, Streptococcus anginosus and Lactobacillus spp. were also found in this study. CONCLUSION: Clonal analysis of the microflora associated with endodontic infections revealed a wide diversity of oral species.
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Absceso/microbiología , Bacterias/clasificación , Enfermedades de la Pulpa Dental/microbiología , Actinobacteria/clasificación , Bacillaceae/clasificación , Bacteroides/clasificación , Cromatografía Líquida de Alta Presión , Células Clonales , Cartilla de ADN , ADN Ribosómico , Enterococcus faecalis/clasificación , Eubacterium/clasificación , Fusobacterium/clasificación , Bacilos Gramnegativos Anaerobios Rectos, Curvos y Espirales/clasificación , Bacterias Grampositivas/clasificación , Humanos , Lactobacillus/clasificación , Desnaturalización de Ácido Nucleico , Peptostreptococcus/clasificación , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/clasificación , ARN Ribosómico 16S , Staphylococcus aureus/clasificación , Streptococcus anginosus/clasificaciónRESUMEN
BACKGROUND: An association between race/ethnicity and the composition of the subgingival microbiota has been found in chronic periodontitis. A study was undertaken to determine the characteristics of the subgingival microbiota of chronic periodontitis in Chileans residing in Santiago. METHODS: Twenty-six subjects (mean age 45 +/- 7 years) with chronic periodontitis, mean probing depth (PD) 2.63 +/- 0.5 mm, mean attachment level (AL) 3.70 +/- 0.77 mm, and without a history of periodontal therapy were selected. Measurements of PD, AL, bleeding on probing, and plaque accumulation were recorded at six sites per tooth. Subgingival plaque samples were taken from the mesial aspect of every tooth and evaluated for the presence, levels, and proportions of 40 bacterial taxa using whole genomic DNA probes and checkerboard DNA-DNA hybridization. The microbial data of the Chileans were compared with data from 115 chronic periodontitis patients from Boston, Massachusetts. Since several clinical and demographic parameters differed between the two populations, significance of differences for each species was determined using analysis of covariance, adjusting for age, plaque level, mean PD, gender, and smoking status. RESULTS: Each of the individual test species was present in at least 25 of the 26 subjects, and 12 subjects (46.1%) harbored all 40 test species. With the exception of Prevotella intermedia, all test species colonized more than 75% of sites, and 25 species colonized > or = 90% of sites including the co-colonizing species of advanced periodontal lesions, termed the red complex, composed of the three species Porphyromonas gingivalis, Tannerella forsythensis (formerly Bacteroides forsythus), and Treponema denticola as well as Fusobacterium nucleatum subspecies, Campylobacter rectus, Peptostreptococcus micros, and Treponerma socranskii. Sixteen of the 40 species differed significantly between Chilean and U.S. subjects. Red, yellow, and other complexes were significantly higher in the Chileans, while the Actinomyces were higher in the U.S. subjects. CONCLUSIONS: The composition of the subgingival plaque differs among different subject populations. Thus, care should be taken when extrapolating the findings of one study to different ethnic groups.
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Periodontitis/microbiología , Adulto , Factores de Edad , Anciano , Bacteroides/clasificación , Boston , Campylobacter/clasificación , Chile , Placa Dental/microbiología , Femenino , Fusobacterium nucleatum/clasificación , Encía/microbiología , Hemorragia Gingival/microbiología , Humanos , Masculino , Persona de Mediana Edad , Peptostreptococcus/clasificación , Pérdida de la Inserción Periodontal/microbiología , Bolsa Periodontal/microbiología , Periodontitis/etnología , Porphyromonas gingivalis/clasificación , Factores Sexuales , Fumar , Treponema/clasificaciónRESUMEN
A 16S rDNA-directed polymerase chain reaction method was used to assess the occurrence of four black-pigmented anaerobic rods, Treponema denticola, and Actinobacillus actinomycetemcomitans in acute periradicular abscesses. Pus was collected by aspiration from 10 cases diagnosed as acute abscesses of endodontic origin. DNA was extracted from the samples and analyzed using a polymerase chain reaction-based identification assay. The method allowed detecting black-pigmented anaerobes in 80% of the examined abscesses. Porphyromonas endodontalis was found in 70%, T. denticola in 50%, Porphyromonas gingivalis in 40%, and Prevotella intermedia in 10% of the cases. P. gingivalis was always found associated with P. endodontalis. Prevotella nigrescens and A. actinomycetemcomitans were not found in any pus sample. The high prevalence of P. endodontalis, T. denticola, and P. gingivalis suggests that they can play an important role in the etiology of acute periradicular abscesses.
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Aggregatibacter actinomycetemcomitans/clasificación , Absceso Periapical/microbiología , Porphyromonas/clasificación , Prevotella/clasificación , Treponema/clasificación , Adulto , Aggregatibacter actinomycetemcomitans/genética , Pérdida de Hueso Alveolar/microbiología , Recuento de Colonia Microbiana , Cartilla de ADN , ADN Bacteriano/análisis , ADN Bacteriano/genética , Caries Dental/microbiología , Necrosis de la Pulpa Dental/microbiología , Humanos , Reacción en Cadena de la Polimerasa , Porphyromonas/genética , Porphyromonas gingivalis/clasificación , Porphyromonas gingivalis/genética , Prevotella/genética , Prevotella intermedia/clasificación , Prevotella intermedia/genética , ARN Ribosómico 16S/análisis , ARN Ribosómico 16S/genética , Supuración , Treponema/genéticaRESUMEN
A random sample of sixty-two 11-15-year-old adolescents from 17 different locations in Guatemala were selected for this study. Pocket depth, Plaque Index, and bleeding upon probing were recorded from 6 randomly selected sites in each subject (a total of 372 sites). Subgingival plaque samples were subsequently collected from these sites and processed by several assays. For cost reasons, in each pair of sites different assays were performed as follows: sites #1, #2--BANA test for T. denticola, P. gingivalis, B. forsythus and screening of plaque samples with polyclonal antibodies (ELISA system) for A. actinomycetemcomitans; sites #3, #4--detection of yeasts by SAB agar; sites #5, #6--detection of Entamoeba gingivalis by the Heidenhain iron hematoxylin modified technique. A total of 66% of the children had at least one site that bled upon probing, 42% exhibited at least one site with pocket depth > 3 mm, and 79% exhibited a high Plaque Index, with the percent of sites affected being 30%, 12% and 41%, respectively. In sites #1, #2 (N = 124), the BANA test assay and A. actinomycetemcomitans tested positive in 77% and 47% of the children accounting for 59% and 31% of the sites, respectively. In sites #3, #4 (N = 124), yeasts were detected in 43% of the children and 29% of the sites. In sites #5, #6 (N = 124), Entamoeba gingivalis was detected in 21% of the children and in 11% of the sites. The risk for severe gingival inflammation and/or increased probing depth was 1.5 and 5.2 times higher if a positive BANA test or A. actinomycetemcomitans test was found in a particular site. No associations could be found for yeasts and Entamoeba gingivalis.