Bacteria in the cavity-restoration interface after varying periods of clinical service - SEM description of distribution and 16S rRNA gene sequence identification of isolates.

OBJECTIVES: To use extracted human teeth with amalgam (n = 26) or GIC (n = 3) restorations in service up to 20 years to evaluate microbiota at the cavity/restoration interface by SEM or culture. MATERIALS AND METHODS: Extracted teeth with intracoronal restorations (n = 20) of known history (2-20 years) were fixed, split, and prepared for SEM to ascertain the pattern and structure of bacterial aggregates on cavity and restoration surfaces. Another 9 teeth were anaerobically decontaminated, split and sampled (cavity/restorations), and cultured (anaerobically, aerobically); recovered isolates were identified by 16S rRNA gene sequencing. RESULTS: SEM showed rods, cocci, and filaments in 11/20 teeth (55%) on cavity and corresponding restoration surfaces; 4/20 (20%) on neither surface; 1/20 (5%) on just cavity; and 4/20 (20%) on just restoration. Microbial growth extended from marginal openings into the deeper interfacial microspace to varying extents but was not always evident. Restoration size or age did not predict bacterial presence. Bacteria-free surfaces (cavity/amalgam) showed possible calcification. Cultivation yielded 160 isolates, mainly Gram-positive (86%) and facultative (81%); and morphotypes of rods (43%), cocci (36%), and cocco-bacilli (18%) belonging to Actinobacteria (45%) and Firmicutes (50%). The most frequent genera were Staphylococcus, Streptococcus, Actinomyces, and Lactobacillus. Biofilms on cavity and restoration appeared independent of each other. CONCLUSIONS: Cavity and amalgam surfaces were independently colonised and some not. The penetration of microbiota into marginal gaps varied; resembled root caries and was dominated by Gram-positive species. CLINICAL RELEVANCE: Marginal gaps around restorations are unavoidable but are not always colonised by bacteria after long-term clinical service. Calcification of biofilms in the restorative interface may prevent further colonisation. The viable microbiota in the restorative interface resembled root caries and may be subject to ecological fluxes of activity and arrest and therefore preventative management.

Reassessment of Routine Midstream Culture in Diagnosis of Urinary Tract Infection.

Midstream urine (MSU) culture remains the gold standard diagnostic test for confirming urinary tract infection (UTI). We previously showed that patients with chronic lower urinary tract symptoms (LUTS) below the diagnostic cutoff on MSU culture may still harbor bacterial infection and that their antibiotic treatment was associated with symptom resolution. Here, we evaluated the results of the United Kingdom's MSU culture in symptomatic patients and controls. Next, we compared the bacterial enrichment capabilities of the MSU culture with those of a 50-µl uncentrifuged culture, a 30-ml centrifuged sediment culture, and 16S rRNA gene sequencing. This study was conducted on urine specimens from 33 LUTS patients attending their first clinical appointment (mean age, 48.7 years; standard deviation [SD], 16.5 years), 30 LUTS patients on treatment (mean age, 47.8 years; SD, 16.5 years) whose symptoms had relapsed, and 29 asymptomatic controls (mean age, 40.7 years, SD, 15.7 years). We showed that the routine MSU culture, adopting the UK interpretation criteria tailored to acute UTI, failed to detect a variety of bacterial species, including recognized uropathogens. Moreover, the diagnostic MSU culture was unable to discriminate between patients and controls. In contrast, genomic analysis of urine enriched by centrifugation discriminated between the groups, generating a more accurate understanding of species richness. In conclusion, the United Kingdom's MSU protocol misses a significant proportion of bacteria, which include recognized uropathogens, and may be unsuitable for excluding UTI in patients with LUTS.

Components in Lentinus edodes mushroom with anti-biofilm activity directed against bacteria involved in caries and gingivitis.

The present study investigated the compounds present in the low molecular mass fraction of Lentinus edodes mushroom (shiitake) extract and their anti-virulence activity against oral pathogens (reference and clinical Streptococcus mutans, Actinomyces naeslundii, and Prevotella intermedia strains). Oxalic, succinic, and quinic acids, and adenine, inosine, and uridine were identified by HPLC-DAD-ESI-MS/MS. Their anti-biofilm production and preformed biofilm disaggregation activities were studied using commercial standard compounds at different concentrations. As regards S. mutans, the highest activity was shown by adenine at 5 mg mL-1 both in the biofilm inhibition (BI 50%) and biofilm disaggregation tests (BD 20%). Considering A. naeslundii, BI values close to 80% were registered for oxalic acid at 1 mg mL-1 and 2 mg mL-1 and BD 50% for quinic acid at 3 mg mL-1. A weaker activity was found against P. intermedia. Furthermore, different mixtures of the commercial standards were tested showing that the activity of a compound can be strongly and sometimes negatively affected by the presence of the other compounds.

Same Exposure but Two Radically Different Responses to Antibiotics: Resilience of the Salivary Microbiome versus Long-Term Microbial Shifts in Feces.

UNLABELLED: Due to the spread of resistance, antibiotic exposure receives increasing attention. Ecological consequences for the different niches of individual microbiomes are, however, largely ignored. Here, we report the effects of widely used antibiotics (clindamycin, ciprofloxacin, amoxicillin, and minocycline) with different modes of action on the ecology of both the gut and the oral microbiomes in 66 healthy adults from the United Kingdom and Sweden in a two-center randomized placebo-controlled clinical trial. Feces and saliva were collected at baseline, immediately after exposure, and 1, 2, 4, and 12 months after administration of antibiotics or placebo. Sequences of 16S rRNA gene amplicons from all samples and metagenomic shotgun sequences from selected baseline and post-antibiotic-treatment sample pairs were analyzed. Additionally, metagenomic predictions based on 16S rRNA gene amplicon data were performed using PICRUSt. The salivary microbiome was found to be significantly more robust, whereas the antibiotics negatively affected the fecal microbiome: in particular, health-associated butyrate-producing species became strongly underrepresented. Additionally, exposure to different antibiotics enriched genes associated with antibiotic resistance. In conclusion, healthy individuals, exposed to a single antibiotic treatment, undergo considerable microbial shifts and enrichment in antibiotic resistance in their feces, while their salivary microbiome composition remains unexpectedly stable. The health-related consequences for the gut microbiome should increase the awareness of the individual risks involved with antibiotic use, especially in a (diseased) population with an already dysregulated microbiome. On the other hand, understanding the mechanisms behind the resilience of the oral microbiome toward ecological collapse might prove useful in combating microbial dysbiosis elsewhere in the body. IMPORTANCE: Many health care professionals use antibiotic prophylaxis strategies to prevent infection after surgery. This practice is under debate since it enhances the spread of antibiotic resistance. Another important reason to avoid nonessential use of antibiotics, the impact on our microbiome, has hardly received attention. In this study, we assessed the impact of antibiotics on the human microbial ecology at two niches. We followed the oral and gut microbiomes in 66 individuals from before, immediately after, and up to 12 months after exposure to different antibiotic classes. The salivary microbiome recovered quickly and was surprisingly robust toward antibiotic-induced disturbance. The fecal microbiome was severely affected by most antibiotics: for months, health-associated butyrate-producing species became strongly underrepresented. Additionally, there was an enrichment of genes associated with antibiotic resistance. Clearly, even a single antibiotic treatment in healthy individuals contributes to the risk of resistance development and leads to long-lasting detrimental shifts in the gut microbiome.

How clean is clean--is a new microbiology standard required?

The role of environment in the spread of nosocomial infection has been acknowledged. One way to control the spread of infection is to control and monitor patient environments to prevent transmission. Studies applying the suggested aerobic colony count standards to monitor environmental contamination were undertaken over an 18-month period at both a London pediatric hospital and in adult intensive care units. The resulting data demonstrate that a large proportion of sites screened for bacterial contamination would fail if using the criteria suggested by previous authors-particularly those sites closest to patients-suggesting a new standard might be required.

Environmental viral contamination in a pediatric hospital outpatient waiting area: implications for infection control.

BACKGROUND: Nosocomial outbreaks of viral etiology are costly and can have a major impact on patient care. Many viruses are known to persist in the inanimate environment and may pose a risk to patients and health care workers. We investigate the frequency of environmental contamination with common health care-associated viruses and explore the use of torque-teno virus as a marker of environmental contamination. METHODS: Environmental screening for a variety of clinically relevant viruses was carried out over 3 months in a UK pediatric hospital using air sampling and surface swabbing. Swabs were tested for the presence of virus nucleic acid by quantitative polymerase chain reactions. RESULTS: Viral nucleic acid was found on surfaces and in the air throughout the screening period, with adenovirus DNA being the most frequent. Door handles were frequently contaminated. Torque-teno virus was also found at numerous sites. CONCLUSION: Evidence of environmental contamination with viral pathogens is present in health care environments and may be indicative of an infectious virus being present. Screening for viruses should be included in infection control strategies. Torque-teno virus may provide a better marker of contamination and reduce time and cost of screening for individual viruses.

The anti-adhesive mode of action of a purified mushroom (Lentinus edodes) extract with anticaries and antigingivitis properties in two oral bacterial phatogens.

BACKGROUND: In previous works we have shown that a low-molecular-mass (LMM) fraction from mushroom (Lentinus edodes) homogenate interferes with binding of Streptococcus mutans to hydroxyapatite and Prevotella intermedia to gingival cells. Additionally, inhibition of biofilm formation of both odonto- and periodonto-pathogenic bacteria and detachment from preformed biofilms have been described for this compound. Further purification of mushroom extract has been recently achieved and a sub-fraction (i.e. # 5) has been identified as containing the majority of the mentioned biological activities. The aim of this study was to characterise the bacterial receptors for the purified mushroom sub-fraction #5 in order to better elucidate the mode of action of this compound when interfering with bacterial adhesion to host surfaces or with bacteria-bacteria interactions in the biofilm state. METHODS: Candidate bacterial molecules to act as target of this compound were bacterial surface molecules involved in cell adhesion and biofilm formation, and, thus, we have considered cell wall associated proteins (CWPs), teichoic acid (TA) and lipoteichoic acid (LTA) of S. mutans, and outer membrane proteins (OMPs) and lipopolysaccharide (LPS) of P. intermedia. RESULTS: Fifteen S. mutans CWPs and TA were capable of binding sub-fraction #5, while LTA did not. As far as P. intermedia is concerned, we show that five OMPs interact with sub-fraction # 5. Capacity of binding to P. intermedia LPS was also studied but in this case negative results were obtained. CONCLUSIONS: Binding sub-fraction # 5 to surface molecules of S. mutans or P. intermedia may result in inactivation of their physiological functions. As a whole, these results indicate, at molecular level, the bacterial surface alterations affecting adhesion and biofim formation. For these antimicrobial properties, the compound may find use in daily oral hygiene.

Effects of mushroom and chicory extracts on the shape, physiology and proteome of the cariogenic bacterium Streptococcus mutans.

BACKGROUND: Dental caries is an infectious disease which results from the acidic demineralisation of the tooth enamel and dentine as a consequence of the dental plaque (a microbial biofilm) accumulation. Research showed that several foods contain some components with antibacterial and antiplaque activity. Previous studies indicated antimicrobial and antiplaque activities in a low-molecular-mass (LMM) fraction of extracts from either an edible mushroom (Lentinus edodes) or from Italian red chicory (Cichorium intybus). METHODS: We have evaluated the antimicrobial mode of action of these fractions on Streptococcus mutans, the etiological agent of human dental caries. The effects on shape, macromolecular syntheses and cell proteome were analysed. RESULTS: The best antimicrobial activity has been displayed by the LMM mushroom extract with a bacteriostatic effect. At the MIC of both extracts DNA synthesis was the main macromolecular synthesis inhibited, RNA synthesis was less inhibited than that of DNA and protein synthesis was inhibited only by roughly 50%. The partial inhibition of protein synthesis is compatible with the observed significant increase in cell mass. The increase in these parameters is linked to the morphological alteration with transition from cocci of the untreated control to elongated cells. Interestingly, these modifications were also observed at sub-MIC concentrations. Finally, membrane and cytosol proteome analysis was conducted under LMM mushroom extract treatment in comparison with untreated S. mutans cells. Significant changes were observed for 31 membrane proteins and 20 of the cytosol fractions. The possible role of the changed proteins is discussed. CONCLUSIONS: This report has shown an antibiotic-like mode of action of mushroom and chicory extracts as demonstrated by induced morphogenetic effects and inhibition of specific macromolecular synthesis. This feature as well as the safe use of this extract as result of its natural origin render the LMM both mushroom and chicory extracts suitable for the formulation into products for daily oral hygiene such as mouthwashes or toothpastes.

Adhesive microbeads for the targeting delivery of anticaries agents of vegetable origin.

The formulation of quinic acid, a food constituent demonstrating potential anticaries and antigingivitis properties, was investigated in an adhesive microparticulate delivery system with the goal of improving its effect by prolonging its residence time at the site of action. Alginate and chitosan were selected as mucoadhesive polymers. The microspheres were prepared by coacervation. Different types of alginates, polymers blends and crosslinking agent concentrations were considered and evaluated. The best results in terms of encapsulation efficiency, in vitro active agent release profile and in vitro adhesive properties, both to oral mucosa and to teeth surface, were obtained with a blend of Alginate Protanal LF200S: Alginate Protanal LF120LS 1:1.5 w/w, 0.1M CaCl(2), and chitosan coating, prepared by a one-step complex coacervation method. This microparticulate delivery system showed prolonged release of quinic acid, and could be used as an active component in chewing gums or mouthwashes for both caries and gingivitis prevention.

Identification of organic acids in Cichorium intybus inhibiting virulence-related properties of oral pathogenic bacteria.

The low molecular mass (LMM) extract of Cichorium intybus var. silvestre (red chicory) has been shown to inhibit virulence-linked properties of oral pathogens including Streptococcus mutans, Actinomyces naeslundii and Prevotella intermedia. In the present study HPLC-DAD-ESI/MS(2) was used to investigate the compounds contained in this extract for their anti-virulence activity. The extract contained a number of components, including oxalic, succinic, shikimic and quinic acids, which interfere with the growth and virulence traits (i.e., biofilm formation, adherence to epithelial cells and hydroxyapatite) of oral pathogens involved in gingivitis and tooth decay. Succinic and quinic acid seem to be the most potent, mainly by interfering with the ability of oral pathogens to form biofilms (either through inhibition of their development or promotion of their disruption). Our findings suggest that one or more of these compounds may modulate plaque formation in vivo, which is a prerequisite for the development of both caries and gingivitis.

High strength re-mineralizing, antibacterial dental composites with reactive calcium phosphates.

OBJECTIVE: Development of high strength dental composites with adhesive, antibacterial and re-mineralizing potential. MATERIALS: Urethane and triethylene glycol dimethacrylates were combined with HEMA (10 or 20wt%) and 2MP (2 or 10wt%), antibacterial chlorhexidine (2.5wt%) and chemical cure initiators. Reactive mono/tri calcium phosphate (CP) mixed with silica/silicon carbide nanoparticles (S) (CP:S weight ratio 1:2 or 2:1) was added (50wt%). RESULTS: Decreasing CP/S ratio and HEMA content reduced monomer conversion at 15min from 93 to 63%. Conversely, decreasing CP/S increased initial "dry" compressive (137-203MPa) and flexural (79-116MPa) strength. With high HEMA content, these decreased by ∼15-20MPa upon 24h water storage. With low HEMA content, average decline was <8MPa due to reduced water sorption. Early water sorption induced mass increase, volume expansion, mono calcium phosphate dissolution and chlorhexidine release, were proportional to the initial calcium phosphate content. Furthermore, they increased ∼1.5 fold upon raising HEMA wt%. These diffusion controlled processes and strength decline slowed after 24h as phosphates reaction bound water within the materials. Increasing 2MP concentration reduced calcium release but did not affect strength. Formulations with high CP/S indicated greater antibacterial activity in agar diffusion and in vitro biofilm tests. SIGNIFICANCE: New material use beneath a conventional composite could potentially reduce high failure rates associated with residual caries and bacterial microleakage.

Maternal oral origin of Fusobacterium nucleatum in adverse pregnancy outcomes as determined using the 16S-23S rRNA gene intergenic transcribed spacer region.

Fusobacterium nucleatum, a common Gram-negative anaerobe prevalent in the oral cavity, possesses the ability to colonize the amniotic cavity and the fetus. However, F. nucleatum may also be part of the vaginal microbiota from where it could reach the amniotic tissues. Due to the heterogeneity of F. nucleatum, consisting of five subspecies, analysis at the subspecies/strain level is desirable to determine its precise origin. The aims of this study were: (i) to evaluate the use of the 16S-23S rRNA gene intergenic transcribed spacer (ITS) region as a tool to differentiate subspecies of F. nucleatum, and (ii) to design a simplified technique based on the ITS to determine the origin of F. nucleatum strains associated with adverse pregnancy outcomes. Amplified fragments of the 16S-23S rRNA gene ITS region corresponding to the five subspecies of F. nucleatum were subjected to cloning and sequencing to characterize the different ribosomal operons of the subspecies. Distinctive length and sequence patterns with potential to be used for identification of the subspecies/strain were identified. These were used to evaluate the origin of F. nucleatum identified in neonatal gastric aspirates (swallowed amniotic fluid) by sequence comparisons with the respective oral and vaginal maternal samples. A simplified technique using a strain-specific primer in a more sensitive nested PCR was subsequently developed to analyse ten paired neonatal-maternal samples. Analysing the variable fragment of the ITS region allowed the identification of F. nucleatum subsp. polymorphum from an oral origin as potentially being involved in neonatal infections. Using a strain-specific primer, the F. nucleatum subsp. polymorphum strain was detected in both neonatal gastric aspirates and maternal oral samples in cases of preterm birth from mothers presenting with localized periodontal pockets. Interestingly, the same strain was not present in the vaginal sample of any case investigated. The 16S-23S rRNA gene ITS can be a useful tool to determine the origin of F. nucleatum. The results of this study strongly indicate that F. nucleatum subsp. polymorphum of oral origin could be involved with pregnancy complications.

Identification of bacteria and potential sources in neonates at risk of infection delivered by Caesarean and vaginal birth.

Neonatal gastric aspirates (NGA) are routinely screened in UK hospitals to investigate fetal/neonatal infections associated with cases of adverse pregnancy outcome (APO). The aim of this study was to describe and compare the microbiology of NGA from Caesarean and vaginal deliveries using molecular methods, and to evaluate other possible clinical and non-clinical variables that may have determined the presence of the bacteria in the samples. The value of using NGA and molecular methods to investigate potential pathogens associated with the risk of early infection was also evaluated. Bacteria were identified by a combined molecular approach on the basis of the 16S rRNA gene using both clone analysis and denaturing gradient gel electrophoresis. A total of 43 and 34 different species were identified in the vaginal (n = 121) and Caesarean (n = 119) deliveries, respectively; 26 of the species observed (51 %) were common to both modalities, although usually less prevalent in the Caesarean cases. Multivariate analysis confirmed an association between infection and prolonged rupture of membranes in vaginal deliveries (odds ratio = 5.7, 95 % confidence interval = 1.1-29.0). Various associations between infection and given variables were also shown, including labour, intrapartum antibiotic prophylaxis, and time and place of sample collection. The molecular methods allowed identification of a range of bacteria and potential sources not previously observed in NGA, including possible genito-urinary, gastrointestinal and oral pathogens. NGA represents a valuable sample for investigating potential pathogens associated with APO and the risk of early infection in neonates using molecular methods.

Plant and fungal food components with potential activity on the development of microbial oral diseases.

This paper reports the content in macronutrients, free sugars, polyphenols, and inorganic ions, known to exert any positive or negative action on microbial oral disease such as caries and gingivitis, of seven food/beverages (red chicory, mushroom, raspberry, green and black tea, cranberry juice, dark beer). Tea leaves resulted the richest material in all the detected ions, anyway tea beverages resulted the richest just in fluoride. The highest content in zinc was in chicory, raspberry and mushroom. Raspberry is the richest food in strontium and boron, beer in selenium, raspberry and mushroom in copper. Beer, cranberry juice and, especially green and black tea are very rich in polyphenols, confirming these beverages as important sources of such healthy substances. The fractionation, carried out on the basis of the molecular mass (MM), of the water soluble components occurring in raspberry, chicory, and mushroom extracts (which in microbiological assays revealed the highest potential action against oral pathogens), showed that both the high and low MM fractions are active, with the low MM fractions displaying the highest potential action for all the fractionated extracts. Our findings show that more compounds that can play a different active role occur in these foods.

Effects of mushroom and chicory extracts on the physiology and shape of Prevotella intermedia, a periodontopathogenic bacterium.

Contrary to the common assumption that food has a negative impact on oral health, research has shown that several foods contain a number of components with antibacterial and antiplaque activity. These natural compounds may be useful for improving daily oral hygiene. In this study we evaluate the mode of antimicrobial action of fractions of mushroom and red chicory extracts on Prevotella intermedia, a periodontopathogenic bacterium. The minimal inhibitory concentration corresponded to 0.5x compared to the natural food concentration for both extracts. This concentration resulted in a bacteriostatic effect in mushroom extract and in a slightly bactericidal effect in chicory extract. Cell mass continued to increase even after division stopped. As regards macromolecular synthesis, DNA was almost totally inhibited upon addition of either mushroom or chicory extract, and RNA to a lesser extent, while protein synthesis continued. Cell elongation occurred after septum inhibition as documented by scanning electron microscopy and cell measurement. The morphogenetic effects are reminiscent of the mode of action of antibiotics such as quinolones or ß-lactams. The discovery of an antibiotic-like mode of action suggests that these extracts can be advantageously employed for daily oral hygiene in formulations of cosmetic products such as mouthwashes and toothpastes.

Testing a low molecular mass fraction of a mushroom (Lentinus edodes) extract formulated as an oral rinse in a cohort of volunteers.

Although foods are considered enhancing factors for dental caries and periodontitis, laboratory researches indicate that several foods and beverages contain components endowed with antimicrobial and antiplaque activities. A low molecular mass (LMM) fraction of an aqueous mushroom extract has been found to exert these activities in in vitro experiments against potential oral pathogens. We therefore conducted a clinical trial in which we tested an LMM fraction of shiitake mushroom extract formulated in a mouthrinse in 30 young volunteers, comparing the results with those obtained in two identical cohorts, one of which received water (placebo) and the other Listerine. Plaque index, gingival index and bacterial counts in plaque samples were determined in all volunteers over the 11 days of the clinical trial. Statistically significant differences (P < 0.05) were obtained for the plaque index on day 12 in subjects treated with mushroom versus placebo, while for the gingival index significant differences were found for both mushroom versus placebo and mushroom versus Listerine. Decreases in total bacterial counts and in counts of specific oral pathogens were observed for both mushroom extract and Listerine in comparison with placebo. The data suggest that a mushroom extract may prove beneficial in controlling dental caries and/or gingivitis/periodontitis.

The effects of fractions from shiitake mushroom on composition and cariogenicity of dental plaque microcosms in an in vitro caries model.

The aim of the current study was to investigate the anticariogenic potential of the (sub)fractions obtained from the edible mushroom shiitake (Lentinula edodes) in in vitro caries model. We used a modified constant depth film fermentor (CDFF) with pooled saliva as the inoculum and bovine dentin as a substratum. The test compounds were low molecular weight fraction (MLMW) of the shiitake extract and subfractions 4 and 5 (SF4 and SF5) of this fraction. Chlorhexidine (CHX) and water served as a positive and a negative control, respectively. Dentin mineral loss was quantified (TMR), microbial shifts within the microcosms were determined (qPCR), and the acidogenicity of the microcosms was assessed (CIA). From the compounds tested, the SF4 of shiitake showed strong inhibiting effect on dentin demineralization and induced microbial shifts that could be associated with oral health. The acid producing potential was increased, suggesting uncoupling of the glycolysis of the microbiota by the exposure to SF4. In conclusion, the results suggest that SF4 of shiitake has an anticariogenic potential.

Effects of fruit and vegetable low molecular mass fractions on gene expression in gingival cells challenged with Prevotella intermedia and Actinomyces naeslundii.

Low molecular mass (LMM) fractions obtained from extracts of raspberry, red chicory, and Shiitake mushrooms have been shown to be an useful source of specific antibacterial, antiadhesion/coaggregation, and antibiofilm agent(s) that might be used for protection towards caries and gingivitis. In this paper, the effects of such LMM fractions on human gingival KB cells exposed to the periodontal pathogens Prevotella intermedia and Actinomyces naeslundii were evaluated. Expression of cytokeratin 18 (CK18) and ß4 integrin (ß4INT) genes, that are involved in cell proliferation/differentiation and adhesion, and of the antimicrobial peptide ß2 defensin (HßD2) in KB cells was increased upon exposure to either live or heat-killed bacteria. All LMM fractions tested prevented or reduced the induction of gene expression by P. intermedia and A. naeslundii depending on the experimental conditions. Overall, the results suggested that LMM fractions could modulate the effects of bacteria associated with periodontal disease in gingival cells.

A molecular analysis of the bacteria present within oral squamous cell carcinoma.

In order to characterize the bacterial microbiota present within oral cancerous lesions, tumorous and non-tumorous mucosal tissue specimens (approx. 1 cm(3)) were harvested from ten oral squamous cell carcinoma (OSCC) patients at the time of surgery. Any microbial contamination on the surface of the specimens was eliminated by immersion in Betadine and washing with PBS. Bacteria were visualized within sections of the OSCC by performing fluorescent in situ hybridization with the universal oligonucleotide probe, EUB338. DNA was extracted from each aseptically macerated tissue specimen using a commercial kit. This was then used as template for PCR with three sets of primers, targeting the 16S rRNA genes of Spirochaetes, Bacteroidetes and the domain Bacteria. PCR products were differentiated by TA cloning and bacterial species were identified by partial sequencing of the 16S rRNA gene fragments. A total of 70 distinct taxa was detected: 52 different phylotypes isolated from the tumorous tissues, and 37 taxa from within the non-tumorous specimens. Differences between the composition of the microbiotas within the tumorous and non-tumorous mucosae were apparent, possibly indicating selective growth of bacteria within carcinoma tissue. Most taxa isolated from within the tumour tissue represented saccharolytic and aciduric species. Whether the presence of these bacteria within the mucosa has any bearing on the carcinogenic process is a concept worthy of further investigation.

Aggregative behavior of bacteria isolated from canine dental plaque.

Interbacterial adhesion of bacteria isolated from canine dental plaque was assessed by performing a visual coaggregation assay. Using conditions mimicking those likely to be encountered in vivo, the entire cultivable plaque microbiota from a single dog was assessed, and eight (6.7%) unique coaggregation interactions were detected for 120 crosses. Transmission electron microscopy was used to visualize several of the bacteria in isolation and as coaggregates, which revealed surface structures that may be involved in adhesion and coaggregation. The results of this study indicate that the prevalence of coaggregating pairs of dental plaque bacteria in dogs is similar to the prevalence of coaggregating pairs of dental plaque bacteria reported in humans. In addition, genera found in both hosts generally exhibited similar coaggregation reactions; however, autoaggregation was found to be more common among oral bacteria isolated from dogs.