Relationship of Metabolic Dysfunction-Associated Steatohepatitis-Related Hepatocellular Carcinoma with Oral and Intestinal Microbiota: A Cross-Sectional Pilot Study.

Background and Objectives: The incidence of metabolic dysfunction-associated steatohepatitis (MASH)-related hepatocellular carcinoma (HCC) is increasing worldwide, alongside the epidemic of obesity and metabolic syndrome. Based on preliminary reports regarding the potential association of HCC and periodontitis, this study aimed to analyze the involvement of periodontal bacteria as well as the oral and intestinal bacterial flora in MASH-related HCC (MASH-HCC). Materials and Methods: Forty-one patients with MASH and nineteen with MASH-HCC participated in the study, completing survey questionnaires, undergoing periodontal examinations, and providing samples of saliva, mouth-rinsed water, feces, and peripheral blood. The oral and fecal microbiome profiles were analyzed by 16S ribosomal RNA sequencing. Bayesian network analysis was used to analyze the causation between various factors, including MASH-HCC, examinations, and bacteria. Results: The genus Fusobacterium had a significantly higher occupancy rate (p = 0.002) in the intestinal microflora of the MASH-HCC group compared to the MASH group. However, Butyricicoccus (p = 0.022) and Roseburia (p < 0.05) had significantly lower occupancy rates. The Bayesian network analysis revealed the absence of periodontal pathogenic bacteria and enteric bacteria affecting HCC. However, HCC directly affected the periodontal bacterial species Porphyromonas gingivalis, Tannerella forsythia, Fusobacterium nucleatum, and Prevotella intermedia in the saliva, as well as the genera Lactobacillus, Roseburia, Fusobacterium, Prevotella, Clostridium, Ruminococcus, Trabulsiella, and SMB53 in the intestine. Furthermore, P. gingivalis in the oral cavity directly affected the genera Lactobacillus and Streptococcus in the intestine. Conclusions: MASH-HCC directly affects periodontal pathogenic and intestinal bacteria, and P. gingivalis may affect the intestinal bacteria associated with gastrointestinal cancer.

Fusobacterium watanabei sp. nov. As additional species within the genus Fusobacerium, isolated from human clinical specimens.

Eight spindle-shaped bacteria were isolated from clinical samples in Japan and investigated for their taxonomic position. Phylogenetic trees (based on 16S rRNA, rpoB, zinc protease, and gyrB gene sequence comparisons) showed distinct clustering of eight strains with the type strain of Fusobacterium nucleatum and its closely related species. In silico whole genome comparison analysis based on average nucleotide index based on BLAST (ANIb) and digital DNA-DNA hybridization (dDDH) data between our clinical isolates (PAGU 1795, PAGU 1796T, and PAGU 1797) and the type strain of the closely related species showed values of less than 92.4% and 49.5%, respectively. On the basis of its phylogenetic and genomic distinctiveness together with differential phenotypic properties and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) characteristic signal patterns, we propose Fusobacterium watanabei sp. nov., with the type strain PAGU 1796T (= GTC 21791T = CCUG 74246T).

Postpartum Fusobacterium gonidiaformans bacteremia.

We present a case of a healthy 29 year-old female with an uneventful vaginal delivery who had transient, sudden onset of rigors and fever 36 hours postpartum. She was found to have Fusobacterium gonidiaformans bacteremia due to retained placental tissue. We report this organism as it is not well-described and rarely reported. It does bear some similarities to other Fusobacterium species that have been reported to cause septicemia in young otherwise healthy patients.

Involvement of Fusobacterium Species in Oral Cancer Progression: A Literature Review Including Other Types of Cancer.

Chronic inflammation caused by infections has been suggested to be one of the most important cause of cancers. It has recently been shown that there is correlation between intestinal bacteria and cancer development including metastasis. As over 700 bacterial species exist in an oral cavity, it has been concerning that bacterial infection may cause oral cancer. However, the role of bacteria regarding tumorigenesis of oral cancer remains unclear. Several papers have shown that Fusobacterium species deriving the oral cavities, especially, play a crucial role for the development of colorectal and esophageal cancer. F. nucleatum is a well-known oral bacterium involved in formation of typical dental plaque on human teeth and causing periodontal diseases. The greatest characteristic of F. nucleatum is its ability to adhere to various bacteria and host cells. Interestingly, F. nucleatum is frequently detected in oral cancer tissues. Moreover, detection of F. nucleatum is correlated with the clinical stage of oral cancer. Although the detailed mechanism is still unclear, Fusobacterium species have been suggested to be associated with cell adhesion, tumorigenesis, epithelial-to-mesenchymal transition, inflammasomes, cell cycle, etc. in oral cancer. In this review, we introduce the reports focused on the association of Fusobacterium species with cancer development and progression including oral, esophageal, and colon cancers.

Utilizing Whole Fusobacterium Genomes To Identify, Correct, and Characterize Potential Virulence Protein Families.

Fusobacterium spp. are Gram-negative, anaerobic, opportunistic pathogens involved in multiple diseases, including a link between the oral pathogen Fusobacterium nucleatum and the progression and severity of colorectal cancer. The identification and characterization of virulence factors in the genus Fusobacterium has been greatly hindered by a lack of properly assembled and annotated genomes. Using newly completed genomes from nine strains and seven species of Fusobacterium, we report the identification and corrected annotation of verified and potential virulence factors from the type 5 secreted autotransporter, FadA, and MORN2 protein families, with a focus on the genetically tractable strain F. nucleatum subsp. nucleatum ATCC 23726 and type strain F. nucleatum subsp. nucleatum ATCC 25586. Within the autotransporters, we used sequence similarity networks to identify protein subsets and show a clear differentiation between the prediction of outer membrane adhesins, serine proteases, and proteins with unknown function. These data have identified unique subsets of type 5a autotransporters, which are key proteins associated with virulence in F. nucleatum However, we coupled our bioinformatic data with bacterial binding assays to show that a predicted weakly invasive strain of F. necrophorum that lacks a Fap2 autotransporter adhesin strongly binds human colonocytes. These analyses confirm a gap in our understanding of how autotransporters, MORN2 domain proteins, and FadA adhesins contribute to host interactions and invasion. In summary, we identify candidate virulence genes in Fusobacterium, and caution that experimental validation of host-microbe interactions should complement bioinformatic predictions to increase our understanding of virulence protein contributions in Fusobacterium infections and disease.IMPORTANCEFusobacterium spp. are emerging pathogens that contribute to mammalian and human diseases, including colorectal cancer. Despite a validated connection with disease, few proteins have been characterized that define a direct molecular mechanism for Fusobacterium pathogenesis. We report a comprehensive examination of virulence-associated protein families in multiple Fusobacterium species and show that complete genomes facilitate the correction and identification of multiple, large type 5a secreted autotransporter genes in previously misannotated or fragmented genomes. In addition, we use protein sequence similarity networks and human cell interaction experiments to show that previously predicted noninvasive strains can indeed bind to and potentially invade human cells and that this could be due to the expansion of specific virulence proteins that drive Fusobacterium infections and disease.

Fusobacterium pseudoperiodonticum sp. nov., Isolated from the Human Oral Cavity.

In the present study, three strains (ChDC F213T, ChDC F251, and ChDC F267) were classified as novel species of genus Fusobacterium based on average nucleotide identity (ANI) and genome-to-genome distance (GGD) analysis and chemotaxonomic characterization. 16S rDNA sequences of strains ChDC F213T, ChDC F251, and ChDC F267 were highly similar to that of F. periodonticum ATCC 33693T (99.6, 99.4, and 99.4%, respectively). ANI and GGD values of the three isolates with F. periodonticum ATCC 33693T ranged from 92.5 to 92.6% and 47.7 to 48.2%, respectively. Considering that threshold of ANI and GGD values for bacterial species discrimination are 95-96% and 70%, respectively, these results indicate that the three isolates represent a novel Fusobacterium species. DNA G + C contents of the three isolates were 28.0 mol% each. Cellular fatty acid analysis of these strains revealed that C14:0, C16:0, and C16:1 ω6c/C16:1 ω7c were major fatty acids. Therefore, these three strains are novel species belonging to genus Fusobacterium. Strain ChDC F213T (= KCOM 1259T = KCTC 5677T = JCM 33009T) is the type strain of a novel species of genus Fusobacterium, for which a name of Fusobacterium pseudoperiodonticum sp. nov. is proposed.

Rare isolation of Fusobacterium varium from a case of Fournier's gangrene.

Fusobacterium is a gram negative obligate anaerobic bacilli, a normal inhabitant of gastrointestinal tract, oropharynx and female genital tract. Here we report a case of Fourniers gangrene from which Fusobacterium varium has been isolated along with certain other pathogens. There are only a few reported cases of Fusobacterium varium in literature and it has never been reported from Fournier's gangrene. Through this report we intend to shed some light on the pathogenic potential of anaerobes which are considered as normal flora.

Genomic analysis identifies association of Fusobacterium with colorectal carcinoma.

The tumor microenvironment of colorectal carcinoma is a complex community of genomically altered cancer cells, nonneoplastic cells, and a diverse collection of microorganisms. Each of these components may contribute to carcinogenesis; however, the role of the microbiota is the least well understood. We have characterized the composition of the microbiota in colorectal carcinoma using whole genome sequences from nine tumor/normal pairs. Fusobacterium sequences were enriched in carcinomas, confirmed by quantitative PCR and 16S rDNA sequence analysis of 95 carcinoma/normal DNA pairs, while the Bacteroidetes and Firmicutes phyla were depleted in tumors. Fusobacteria were also visualized within colorectal tumors using FISH. These findings reveal alterations in the colorectal cancer microbiota; however, the precise role of Fusobacteria in colorectal carcinoma pathogenesis requires further investigation.

Fusobacterium hwasookii sp. nov., Isolated from a Human Periodontitis Lesion.

In this study, we classified the five strains (ChDC F128(T), ChDC F145, ChDC F174, ChDC F206, and ChDC F300) as a novel species of genus Fusobacterium by DNA-DNA hybridization and multi-locus phylogenetic analysis (MLPA), based on a single sequence (24,715 bp) of 22 concatenated housekeeping genes, with morphological and chemotaxonomic characteristics. DNA-DNA hybridization data showed that the values of genomic relatedness between ChDC F128(T) and each of the other novel strains were ranged from 79.0 to 82.6 %, while those of genomic relatedness between ChDC F128(T) and type strain of each of subspecies of F. nucleatum or Fusobacterium periodonticum were ranged from 40.9 to 54.4 %. MLPA revealed that the 5 strains were clustered as one group and clearly discriminated with F. nucleatum and F. periodonticum with 100 % bootstrap value. The DNA G+C content of the five novel strains were ranged from 26.9 to 27.0 mol%. The cellular fatty acid analysis of clinical isolates and type strains revealed C14:0, C16:0, and cis-9 C16:1 as the major fatty acids. The cell wall peptidoglycan of the 5 strains was comprised of meso-lanthionine. These results show that the 5 strains are novel species and belong to the genus Fusobacterium. Strain ChDC F128(T) (=KCOM 1249(T) = KCTC 5108(T) = JCM 30218(T)) is suggested to be the type strain of a novel species of genus Fusobacterium, for which the name Fusobacterium hwasookii sp. nov. is proposed.

Fusobacterium necrophorum and other Fusobacterium spp. isolated from head and neck infections: A 10-year epidemiology study in an academic hospital.

BACKGROUND: Fusobacterium spp. from clinical specimens are increasingly reported. We sought to describe the epidemiology, the microbiological, and the clinical characteristics of head and neck infections caused by Fusobacterium necrophorum and other Fusobacterium spp. MATERIALS AND METHODS: Retrospective cohort study between October 1st, 2004 and September 30(th), 2014 performed in an academic hospital. Electronic patient charts and the laboratory information system were reviewed for demographic and microbiological data. The number and percentages of specific diagnosis and treatment among patients with positive Fusobacterium spp. culture were calculated. The incidence was calculated based on the number of specimens investigated each year. RESULTS: Included were 230 cultures of 230 patients (median age of 28 years, 61.7% men). F. necrophorum was often found in young patients with high C-reactive protein (CRP) and high number of leukocytes in blood. Other Fusobacterium spp. were often found in middle aged patients with relatively high CRP and slightly increased leukocytes. Three major causes of the isolation of F. necrophorum and other Fusobacterium spp. were acute tonsillitis (n = 18, incidence of 0.2%), peritonsillar abscess (n = 39, 0.5%) and acute otitis (n = 45, 0.1%). While F. necrophorum was found in majority (37/57) of patients with acute tonsillitis or peritonsillar abscess, Fusobacterium spp. other than F. necrophorum were found in the majority (35/45) of patients with acute otitis. Isolated fusobacteria were susceptible to beta-lactam antibiotics, clindamycin and metronidazole. The outcomes of patients with Fusobacterium spp. were good. CONCLUSION: F. necrophorum and other Fusobacterium spp. are rare cause of head and neck infections. The infections are well treated by combination of antibiotics and surgery.

Evaluation of antibiotic susceptibility of Bacteroides, Prevotella and Fusobacterium species isolated from patients of the N. N. Blokhin Cancer Research Center, Moscow, Russia.

In total 122 non-duplicate Bacteroides, Prevotella and Fusobacterium spp isolated from cancer patients between 2004 and 2014 were involved in this study. Most of the strains belonged to the B. fragilis group (55%), followed by Prevotella strains (34.4%) and Fusobacterium spp (10.6%). The species identification was carried out by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and they were identified on species level with a log (score) >2.0. The most common isolates were B. fragilis, B. thetaiotaomicron, B. ovatus and B. vulgatus. Among Prevotella species, the most frequently isolated species were P. buccae, P. buccalis, P. oris, P. denticola and P. nigrescens, and most of the Fusobacterium spp. were F. nucleatum. Susceptibilities of the strains were determined by the E-test methodology. The percentage of the susceptibility of B. fragilis group isolates were: metronidazole (MIC ≤4 µg/ml), 97%; imipenem (MIC ≤2 µg/ml), 95.5%; amoxicillin/clavulanate (MIC ≤4 µg/ml), 95.5% and clindamycin (MIC ≤4 µg/ml), 77.6%. Three B. fragilis isolates proved to be multidrug-resistant (parallel resistance to imipenem, amoxicillin/clavulanate and metronidazole or clindamycin was observed). All Prevotella strains tested were susceptible to imipenem and amoxicillin/clavulanate, whereas 78.6% of the pigmented Prevotella species and 46.4% of the non-pigmented species were resistant to penicillin (MIC >0.5 µg/ml). The susceptibility to metronidazole and clindamycin were 93% and 88%, respectively. All Fusobacterium strains were sensitive to all tested antibiotics, including penicillin.

Establishment of ruminal bacterial community in dairy calves from birth to weaning is sequential.

AIM: Establishment of ruminal bacterial community in dairy calves. METHODS AND RESULTS: Rumen bacterial community was analysed on 6 calves bred according to commercial practices from day one to weaning at day 83 of age, using 454 16S rRNA-based pyrosequencing. Samples taken at day 1 did not produce amplicons. Analysis of data revealed a three-stage implantation process with a progressive but important shift of composition. At day 2, the bacterial community was mainly composed of Proteobacteria (70%) and Bacteroidetes (14%), and Pasteurellaceae was the dominant family (58%). The bacterial community abruptly changed between days 2 and 3, and until day 12, dominant genera were Bacteroides (21%), Prevotella (11%), Fusobacterium (5%) and Streptococcus (4%). From 15 to 83 days, when solid food intake rapidly increased, Prevotella became dominant (42%) and many genera strongly decreased or were no longer detected. A limited number of bacteria genera correlated with feed intake, rumen volatile fatty acids and enzymatic activities. CONCLUSION: The ruminal bacterial community is established before intake of solid food, but solid food arrival in turn shapes this community. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides insight into the establishment of calves' rumen bacterial community and suggests a strong effect of diet.

Microbial signature profiles of periodontally healthy and diseased patients.

AIM: To determine microbial profiles that discriminate periodontal health from different forms of periodontal diseases. METHODS: Subgingival biofilm was obtained from patients with periodontal health (27), gingivitis (11), chronic periodontitis (35) and aggressive periodontitis (24), and analysed for the presence of >250 species/phylotypes using HOMIM. Microbial differences among groups were examined by Mann-Whitney U-test. Regression analyses were performed to determine microbial risk indicators of disease. RESULTS: Putative and potential new periodontal pathogens were more prevalent in subjects with periodontal diseases than periodontal health. Detection of Porphyromonas endodontalis/Porphyromonas spp. (OR 9.5 [1.2-73.1]) and Tannerella forsythia (OR 38.2 [3.2-450.6]), and absence of Neisseria polysaccharea (OR 0.004 [0-0.15]) and Prevotella denticola (OR 0.014 [0-0.49], p < 0.05) were risk indicators of periodontal disease. Presence of Aggregatibacter actinomycetemcomitans (OR 29.4 [3.4-176.5]), Cardiobacterium hominis (OR 14.9 [2.3-98.7]), Peptostreptococcaceae sp. (OR 35.9 [2.7-483.9]), P. alactolyticus (OR 31.3 [2.1-477.2]), and absence of Fretibacterium spp. (OR 0.024 [0.002-0.357]), Fusobacterium naviforme/Fusobacterium nucleatum ss vincentii (OR 0.015 [0.001-0.223]), Granulicatella adiacens/Granulicatella elegans (OR 0.013 [0.001-0.233], p < 0.05) were associated with aggressive periodontitis. CONCLUSION: There were specific microbial signatures of the subgingival biofilm that were able to distinguish between microbiomes of periodontal health and diseases. Such profiles may be used to establish risk of disease.

[Quantitation of intestinal Fusobacterium and butyrate- producing bacteria in patients with colorectal adenomas and colorectal cancer].

OBJECTIVE: To compare the abundance of 16S rRNA gene of intestinal Fusobacterium and butyrate-producing bacteria in patients with colorectal adenomas patients and colorectal cancer and to reveal the correlation between the target bacteria and the development of colorectal cancer. METHODS: Feces were collected from colorectal cancer patients (n=19), colorectal adenomas patients (n=12) and healthy subjects (n=19). Bacteria genome DNA from the fecal samples was used to quantitate the Fusobacterium, two butyrate-producing bacteria Eubacterium rectal, Faecalibacterium prausnitzii and total bacteria by real-time polymerase chain reaction. Then the variation of the target bacteria among different groups were assayed using Mann-Whitney U test. RESULTS: The abundance of Fusobacterium was significantly higher in colorectal cancer patients than that in healthy subjects (P = 0.000) and colorectal adenomas patients (P = 0.013), and it was significantly higher in colorectal cancer patients than that in colorectal adenomas patients (P = 0.002). F. prausnitzii was significantly lower in colorectal adenomas patients compared to healthy subjects (P = 0.033). The total bacteria count was significantly lower in the colorectal adenomas samples than that in the healthy samples (P = 0.002). There was no significantly difference of E. rectal between the three groups. CONCLUSIONS: The shifts in the colonic bacterial population may potentially contribute to the development of colorectal cancer.

Comparative genomics reveal a novel phylotaxonomic order in the genus Fusobacterium.

Fusobacteria have been associated to different diseases, including colorectal cancer (CRC), but knowledge of which taxonomic groups contribute to specific conditions is incomplete. We analyzed the genetic diversity and relationships within the Fusobacterium genus. We report recent and ancestral recombination in core genes, indicating that fusobacteria have mosaic genomes and emphasizing that taxonomic demarcation should not rely on single genes/gene regions. Across databases, we found ample evidence of species miss-classification and of undescribed species, which are both expected to complicate disease association. By focusing on a lineage that includes F. periodonticum/pseudoperiodonticum and F. nucleatum, we show that genomes belong to four modern populations, but most known species/subspecies emerged from individual ancestral populations. Of these, the F. periodonticum/pseudoperiodonticum population experienced the lowest drift and displays the highest genetic diversity, in line with the less specialized distribution of these bacteria in oral sites. A highly drifted ancestral population instead contributed genetic ancestry to a new species, which includes genomes classified within the F. nucleatum animalis diversity in a recent CRC study. Thus, evidence herein calls for a re-analysis of F. nucleatum animalis features associated to CRC. More generally, our data inform future molecular profiling approaches to investigate the epidemiology of Fusobacterium-associated diseases.

Comprehensive microbiological findings in peri-implantitis and periodontitis.

AIM: The microbial differences between peri-implantitis and periodontitis in the same subjects were examined using 16S rRNA gene clone library analysis and real-time polymerase chain reaction. MATERIALS AND METHODS: Subgingival plaque samples were taken from the deepest pockets of peri-implantitis and periodontitis sites in six subjects. The prevalence of bacteria was analysed using a 16S rRNA gene clone library and real-time polymerase chain reaction. RESULTS: A total of 333 different taxa were identified from 799 sequenced clones; 231 (69%) were uncultivated phylotypes, of which 75 were novel. The numbers of bacterial taxa identified at the sites of peri-implantitis and periodontitis were 192 and 148 respectively. The microbial composition of peri-implantitis was more diverse when compared with that of periodontitis. Fusobacterium spp. and Streptococcus spp. were predominant in both peri-implantitis and periodontitis, while bacteria such as Parvimonas micra were only detected in peri-implantitis. The prevalence of periodontopathic bacteria was not high, while quantitative evaluation revealed that, in most cases, prevalence was higher at peri-implantitis sites than at periodontitis sites. CONCLUSIONS: The biofilm in peri-implantitis showed a more complex microbial composition when compared with periodontitis. Common periodontopathic bacteria showed low prevalence, and several bacteria were identified as candidate pathogens in peri-implantitis.

Subgingival microbiome in smokers and non-smokers in periodontitis: an exploratory study using traditional targeted techniques and a next-generation sequencing.

AIM: To compare the results of two targeted techniques to an open-ended technique in periodontitis patients, differentiated on the basis of smoking habit. MATERIALS & METHODS: Thirty periodontitis patients (15 smokers and 15 non-smokers) provided subgingival plaque samples for 16S rRNA gene amplicon sequencing, culturing and quantitative polymerase chain reaction (qPCR). RESULTS: No differences were found in the composition of the subgingival microbiome between smokers and non-smokers with culture and qPCR. With pyrosequencing, operational taxonomic units (OTUs) classified to genera Fusobacterium, Prevotella and Selenomonas were more abundant in smokers, while OTUs belonging to the genera Peptococcus and Capnocytophaga were more abundant in non-smokers. Principal coordinate analysis identified two clusters; one was composed mainly of smokers (80%) and revealed significantly lower taxonomic diversity, higher attachment loss and higher proportion of the genera Fusobacterium, Paludibacter and Desulfobubus. CONCLUSION: In periodontitis, there is a difference in the composition of the subgingival microbiome between smokers and non-smokers, as revealed by pyrosequencing. This difference was not identified by the targeted techniques. Low taxonomic diversity was associated with higher disease severity, especially in smokers. This supports the hypothesis of the ecological microbial-host interaction in the severity of periodontal disease.

Bacterial biofilm composition in caries and caries-free subjects.

BACKGROUND: Certain major pathogens such as Streptococcus mutans, Lactobacillus spp. and others have been reported to be involved in caries initiation and progression. Yet, in addition to those leading pathogens, microbial communities seem to be much more diverse and individually differing. The aim of this study, therefore, was to analyze the bacterial composition of carious dentin and the plaque of caries-free patients by using a custom-made, real-time quantitative polymerase chain reaction assay (RQ-PCR). METHODS: The study included 26 patients with caries and 28 caries-free controls. Decayed tooth substance and plaque samples were harvested. Bacterial DNA was extracted and tested for the presence of 43 bacterial species or species groups using RQ-PCR. RESULTS: Relative quantification revealed that Propionibacterium acidifaciens was significantly more abundant in caries samples than were other microorganisms (fold change 169.12, p = 0.023). In the caries-free samples, typical health-associated species were significantly more prevalent. Unsupervised hierarchical cluster analysis showed a high abundance of P. acidifaciens in caries subjects and distinct but individually differing bacterial clusters in the caries-free subjects. The distribution of 11 bacteria allowed full discrimination between caries and caries-free subjects. CONCLUSION: Within the investigated cohort, P. acidifaciens was the only pathogen significantly more abundant in caries subjects. Cluster analysis yielded a diverse flora in caries-free subjects, whereas it was narrowed down to a small range of a few outcompeting members in caries subjects.

Predominant bacterial species in subgingival plaque in dogs.

BACKGROUND AND OBJECTIVE: The dog has been used extensively for experimental and microbiological studies on periodontitis and peri-implantitis without detailed knowledge about the predominant flora of the subgingival plaque. This study was designed to evaluate the predominant cultivable bacterial species in dogs and compare them phenotypically and genotypically with corresponding human species. MATERIAL AND METHODS: Four subgingival samples were taken from two upper premolars in each of six Labrador retrievers. The samples from each dog were processed for anaerobic culture. From the samples of each dog, the five or six predominating bacteria based on colony morphology were selected and pure cultured. Each of the strains was characterized by Gram stain, anaerobic/aerobic growth and API-ZYM test. Eighteen strains showing clear-cut phenotypic differences were further classified based on DNA sequencing technology. Cross-reactions of DNA probes from human and dog strains were also tested against a panel of both human and dog bacterial species. RESULTS: Thirty-one strains in the dogs were isolated and characterized. They represented 21 different species, of which six belonged to the genus Porphyromonas. No species was found consistently in the predominant flora of all six dogs. Porphyromonas crevioricanis and Fusobacterium canifelinum were the two most prevalent species in predominant flora in dogs. DNA probes from human and dog species cross-reacted to some extent with related strains from humans and dogs; however, distinct exceptions were found. CONCLUSION: The predominant cultural subgingival flora in dogs shows great similarities with the subgingival bacteria from humans at the genus level, but distinct differences at the species level; however, a genetic relatedness could be disclosed for most strains investigated.

Molecular profiling of oral microbiota in jawbone samples of bisphosphonate-related osteonecrosis of the jaw.

OBJECTIVE: Infection has been hypothesized as a contributing factor to bisphosphonate (BP)-related osteonecrosis of the jaw (BRONJ). The objective of this study was to determine the bacterial colonization of jawbone and identify the bacterial phylotypes associated with BRONJ. MATERIALS AND METHODS: Culture-independent 16S rRNA gene-based molecular techniques were used to determine and compare the total bacterial diversity in bone samples collected from 12 patients with cancer (six, BRONJ with history of BP; six, controls without BRONJ, no history of BP but have infection). RESULTS: Denaturing gradient gel electrophoresis profile and Dice coefficient displayed a statistically significant clustering of profiles, indicating different bacterial population in BRONJ subjects and control. The top three genera ranked among the BRONJ group were Streptococcus (29%), Eubacterium (9%), and Pseudoramibacter (8%), while in the control group were Parvimonas (17%), Streptococcus (15%), and Fusobacterium (15%). H&E sections of BRONJ bone revealed layers of bacteria along the surfaces and often are packed into the scalloped edges of the bone. CONCLUSION: This study using limited sample size indicated that the jawbone associated with BRONJ was heavily colonized by specific oral bacteria and there were apparent differences between the microbiota of BRONJ and controls.