Newly identified pathogens associated with periodontitis: a systematic review.

There is substantial evidence supporting the role of certain oral bacteria species in the onset and progression of periodontitis. Nevertheless, results of independent-culture diagnostic methods introduced about a decade ago have pointed to the existence of new periodontal pathogens. However, the data of these studies have not been evaluated together, which may generate some misunderstanding on the actual role of these microorganisms in the etiology of periodontitis. The aim of this systematic review was to determine the current weight of evidence for newly identified periodontal pathogens based on the results of "association" studies. This review was conducted and reported in accordance with the PRISMA statement. The MEDLINE, EMBASE, and Cochrane databases were searched up to September 2013 for studies (1) comparing microbial data of subgingival plaque samples collected from subjects with periodontitis and periodontal health and (2) evaluating at least 1 microorganism other than the already-known periodontal pathogens. From 1,450 papers identified, 41 studies were eligible. The data were extracted and registered in predefined piloted forms. The results suggested that there is moderate evidence in the literature to support the association of 17 species or phylotypes from the phyla Bacteroidetes, Candidatus Saccharibacteria, Firmicutes, Proteobacteria, Spirochaetes, and Synergistetes. The phylum Candidatus Saccharibacteria and the Archaea domain also seem to have an association with disease. These data point out the importance of previously unidentified species in the etiology of periodontitis and might guide future investigations on the actual role of these suspected new pathogens in the onset and progression of this infection.

Utilidad de la espectrometría de masas MALDI-TOF en la identificación de bacterias anaerobias / Utility of MALDI-TOF MS for the identification of anaerobic bacteria

El análisis de espectrometría de masas mediante la metodología hoy conocida como MALDI-TOF MS (Matrix-assited laser desorption/ionization time-of-flight mass spectrometry) se ha convertido en un recurso de referencia para la identificación de microorganismos en microbiología clínica. No obstante, los datos relativos a algunos grupos de microorganismos son todavía controvertidos. El objetivo del presente estudio fue determinar la utilidad del MALDI-TOF MS para la identificación de aislamientos clínicos de bacterias anaerobias. Se analizaron 106 aislamientos de bacterias anaerobias mediante MALDI-TOF MS y por pruebas bioquímicas convencionales. En aquellos casos en los que la identificación por metodología convencional no era aplicable o frente a una discordancia de resultados entre las metodologías citadas, se realizó la secuenciación del gen 16S del ARNr. El método convencional y el MALDI-TOF MS coincidieron a nivel de género y especie en un 95,3 % de los casos considerando la totalidad de los aislamientos estudiados. Al considerar solo el conjunto de los bacilos gram negativos, la coincidencia fue del 91,4 %; entre los bacilos gram positivos, fue del 100 %; los 8 aislados de cocos gram positivos estudiados coincidieron y también hubo coincidencia en el único coco gram negativo incluido. Los datos obtenidos en este estudio demuestran que el MALDI-TOF MS ofrece la posibilidad de llegar a una adecuada identificación de bacterias anaerobias

The analysis by MALDI-TOF MS (Matrix-assited laser desorption/ionization time-of-flight mass spectrometry) has become a reference method for the identification of microorganisms in Clinical Microbiology. However, data on some groups of microorganisms are still controversial. The aim of this study is to determine the utility of MALDI-TOF MS for the identification of clinical isolates of anaerobic bacteria. One-hundred and six anaerobic bacteria isolates were analyzed by MALDI-TOF MS and by conventional biochemical tests. In those cases where identification by conventional methodology was not applicable or in the face of discordance between sequencing methodologies, 16 S rRNA gene sequence analysis was performed. The conventional method and MALDI-TOF MS agreed at genus and species level by 95.3 %. Concordance in gram-negative bacilli was 91.4% and 100% among gram-positive bacilli; there was also concordance both in the 8 isolates studied in gram-positive cocci and in the single gram-negative cocci included. The data obtained in this study demonstrate that MALDI-TOF MS offers the possibility of adequate identification of anaerobic bacteria

Susceptibility of as-yet-uncultivated and difficult-to-culture bacteria to chemomechanical procedures.

INTRODUCTION: A significant portion of the bacteria taking part of the microbiome associated with apical periodontitis still remain to be cultivated and phenotypically characterized. This molecular study evaluated the prevalence of selected as-yet-uncultivated and difficult-to-culture bacterial taxa in infected root canals and their susceptibility to chemomechanical procedures. METHODS: Root canals of single-rooted teeth with apical periodontitis were prepared using rotary nickel-titanium instruments and 2.5% sodium hypochlorite as the irrigant. DNA extracts from samples taken before (S1) and after (S2) chemomechanical preparation were surveyed for the presence of 7 as-yet-uncultivated phylotypes and 1 difficult-to-culture species using end-point polymerase chain reaction. Samples were also subjected to quantitative analysis of total bacteria and levels of the 2 most prevalent taxa. RESULTS: Bacteroidaceae sp. HOT-272 (24%) and Fretibacterium fastidiosum (20%) were the most prevalent taxa in S1. Their mean counts in S1 were 8.25 × 10(3) and 2.13 × 10(3) rRNA gene copies, corresponding to 0.18% and 0.55% of the total bacteria. Chemomechanical debridement promoted a highly statistically significant reduction in total bacterial counts (P < .001), but 64% of the canals were still positive for bacterial presence. Of the target taxa, only Bacteroidaceae sp. HOT-272 and F. fastidiosum were detected in S2 (each one in 1 sample). The reduction in counts of both taxa was also highly significant (P < .001). CONCLUSIONS: Findings confirmed that several as-yet-uncultivated and difficult-to-grow bacterial taxa can participate in the microbiome associated with apical periodontitis. Two of them were found in relatively high prevalence but rarely as a dominant species. Chemomechanical procedures were highly effective in completely eliminating these taxa or at least substantially reducing their numbers.

Microbiological analysis of infected root canals from symptomatic and asymptomatic teeth with periapical periodontitis and the antimicrobial susceptibility of some isolated anaerobic bacteria.

The purpose of the present study was to investigate the correlation between the composition of the bacterial flora isolated from infected root canals of teeth with apical periodontitis with the presence of clinical signs and symptoms, and to test the antibiotic susceptibility of five anaerobic bacteria mostly commonly found in the root canals of symptomatic teeth against various substances using the E-test. Microbial samples were taken from 48 root canals, 29 symptomatic and 19 asymptomatic, using adequate techniques. A total of 218 cultivable isolates were recovered from 48 different microbial species and 19 different genera. Root canals from symptomatic teeth harbored more obligate anaerobes and a bigger number of bacterial species than the asymptomatic teeth. More than 70% of the bacterial isolates were strict anaerobes. Statistical analysis used a Pearson Chi-squared test or a one-sided Fisher's Exact test as appropriate. Suggested relationships were found between specific microorganisms, especially gram-negative anaerobes, and the presence of spontaneous or previous pain, tenderness to percussion, pain on palpation and swelling amoxicillin, amoxicillin + clavulanate and cephaclor were effective against all the strains tested. The lowest susceptibility rate was presented by Prevotella intermedia/nigrescens against Penicillin G. Our results suggested that specific bacteria are associated with endodontic symptoms of infected teeth with periapical periodontitis and the majority of the anaerobic bacterial species tested were susceptible to all antibiotics studied.

PCR methodology as a valuable tool for identification of endodontic pathogens.

OBJECTIVES: This paper reviews the principles of polymerase chain reaction (PCR) methodology, its application in identification of endodontic pathogens and the perspectives regarding the knowledge to be reached with the use of this highly sensitive, specific and accurate methodology as a microbial identification test. DATA SOURCES: Studies published in the medical, dental and biological literature. STUDY SELECTION: Evaluation of published epidemiological studies examining the endodontic microbiota through PCR methodology. CONCLUSIONS: PCR technology has enabled the detection of bacterial species that are difficult or even impossible to culture as well as cultivable bacterial strains showing a phenotypically divergent or convergent behaviour. Moreover, PCR is more rapid, much more sensitive, and more accurate when compared with culture. Its use in endodontics to investigate the microbiota associated with infected root canals has expanded the knowledge on the bacteria involved in the pathogenesis of periradicular diseases. For instance, Tannerella forsythensis (formerly Bacteroides forsythus), Treponema denticola, other Treponema species, Dialister pneumosintes, and Prevotella tannerae were detected in infected root canals for the first time and in high prevalence when using PCR analysis. The diversity of endodontic microbiota has been demonstrated by studies using PCR amplification, cloning and sequencing of the PCR products. Moreover, other fastidious bacterial species, such as Porphyromonas endodontalis, Porphyromonas gingivalis and some Eubacterium spp., have been reported in endodontic infections at a higher prevalence than those reported by culture procedures.

Avaliaçäo clínica e microbiológica da saúde periodontal de crianças portadoras de fissura completa de lábio e palato unilateral comparada a de crianças sem fissuras na mesma faixa etária (5-6 anos) / Clinical and microbiological evaluation of periodontal condition in 5-6 years old children with unilateral complete cleft lip and palate compared to normal controls

Crianças portadoras de fissura de lábio e palato apresentam alteraçöes anatômicas decorrentes da própria anomalia e dos procedimentos terapêuticos para a sua reabilitaçäo, que podem predispô-las a um maior acúmulo de placa bacteriana, principalmente na regiäo anterior da maxila. Com o objetivo de comparar as condiçöes clínicas, através da utilizaçäo dos índices de placa (IP) e gengival (IG) de Loe (1967) e a presença das bactérias P. gingivalis, P. nigrescens e T. denticola, através da técnica "Slot Immunoblot", foram avaliadas 57 crianças com dentiçäo decídua e idades entre 5 e 6 anos, sendo 30 portadoras de fissura completa de lábio e palato unilateral (grupo experimental) e 27 crianças sem fissura (grupo controle). O IP médio do grupo experimental (1,82 ñ 0,38) foi superior ao do grupo controle (1,63 ñ 0,38), embora sem diferença estatisticamente significante, enquanto o IG médio do grupo experimental (1,05 ñ 0,22) foi significantemente superior ao do grupo controle (0,79 ñ 0,33) (p<0,05). No grupo experimental a área da fissura (2 sítios), com IP médio de 2,04 ñ 0,58 e IG médio de 1,11 ñ 0,26, quando comparada à regiäo posterior (4 sítios), com IP médio de 1,74 ñ 0,37 e IG médio de 1,04 ñ 0,26, mostrou difrença estatisticamente significante somente em relaçäo ao IP. Considerando a severidade, a maioria das crianças dos dois grupos apresentou IP de grau moderado, 73,33 por cento para o grupo experimental e 81,48 por cento para o grupo controle; enquanto, para o IG, a maioria apresentou severidade de grau baixo, sendo de 53,33 por cento para o grupo experimental e de 70,37 por cento para a grupo controle...

Coprothermobacter platensis sp. nov., a new anaerobic proteolytic thermophilic bacterium isolated from an anaerobic mesophilic sludge.

A new anaerobic, proteolytic, moderately thermophilic bacterium, strain 3RT, was isolated from a methanogenic mesophilic reactor treating protein-rich wastewater. The cells were Gram-negative, non-spore-forming, non-motile rods. The DNA base composition was 43 mol% G + C. The optimum pH and temperature for growth were 7.0 and 55 degrees C respectively. The bacterium fermented gelatin, casein, bovine albumin, peptone and yeast extract. Glucose, fructose, sucrose, maltose and starch were poorly fermented. The major fermentation products from glucose were acetate, CO2 and H2 and, from gelatin, propionate was also detected. Growth on glucose was stimulated by thiosulfate, which was reduced to sulfide. Sulfate and nitrate were not reduced. 16S rRNA gene analysis revealed that the isolated bacterial strain was phylogenetically related to Coprothermobacter proteolyticus (96.3% sequence similarity), the only known species within the genus. DNA-DNA hybridization analysis demonstrated a very low level of homology, indicating that the isolated strain and C. proteolyticus were not related at species level. Therefore, it is proposed to classify the described strain in the genus Coprothermobacter as a new species, Coprothermobacter platensis. The type strain of C. platensis is strain 3RT (= DSM 11748T).

12alpha- and 7alpha-hydroxysteroid dehydrogenase activities from Fusobacterium spp.

On screening fecal organisms for hydroxysteroid dehydrogenase activities applicable to bile acid metabolism studies, we have isolated a gram negative "Bacteroides-like" anaerobe which yields both 12alpha- and 7alpha-hydroxysteroid dehydrogenase (HSDH) activities in cell-free preparation. At the optimal harvest time of 36 hours, approximately 4500 units 12alpha-HSDH and 360 units 7alpha-HSDH were produced per 10(10) viable cells. The two enzymes appear to be separate entities in the basis of their stabilities on freezing, and prolonged storage at room temperature and elution volumes on Sephadex G 200. Thin layer chromatography studies on oxidation products confirmed the respective sites of oxidation to be the 12alpha-OH and 7alpha-OH position. No 3alpha-OH oriented activity was measurable. Preliminary kinetic studies of the 12alpha-HSDH revealed a broad pH curve with optimal activity at pH 9.5. Michaelis constants for glycodeoxycholate and NADP were estimated at 1.5 x 10(-4)M and 3.3 x 10(-5)M respectively.