The oral microbiota is a reservoir for antimicrobial resistance: resistome and phenotypic resistance characteristics of oral biofilm in health, caries, and periodontitis.

BACKGROUND: Antimicrobial resistance (AMR) is an ever-growing threat to modern medicine and, according to the latest reports, it causes nearly twice as many deaths globally as AIDS or malaria. Elucidating reservoirs and dissemination routes of antimicrobial resistance genes (ARGs) are essential in fighting AMR. Human commensals represent an important reservoir, which is underexplored for the oral microbiota. Here, we set out to investigate the resistome and phenotypic resistance of oral biofilm microbiota from 179 orally healthy (H), caries active (C), and periodontally diseased (P) individuals (TRN: DRKS00013119, Registration date: 22.10.2022). The samples were analysed using shotgun metagenomic sequencing combined, for the first time, with culture technique. A selection of 997 isolates was tested for resistance to relevant antibiotics. RESULTS: The shotgun metagenomics sequencing resulted in 2,069,295,923 reads classified into 4856 species-level OTUs. PERMANOVA analysis of beta-diversity revealed significant differences between the groups regarding their microbiota composition and their ARG profile. The samples were clustered into three ecotypes based on their microbial composition. The bacterial composition of H and C samples greatly overlapped and was based on ecotypes 1 and 2 whereas ecotype 3 was only detected in periodontitis. We found 64 ARGs conveying resistance to 36 antibiotics, particularly to tetracycline, macrolide-lincosamide-streptogramin, and beta-lactam antibiotics, and a correspondingly high prevalence of phenotypic resistance. Based on the microbiota composition, these ARGs cluster in different resistotypes, and a higher prevalence is found in healthy and caries active than in periodontally diseased individuals. There was a significant association between the resistotypes and the ecotypes. Although numerous associations were found between specific antibiotic resistance and bacterial taxa, only a few taxa showed matching associations with both genotypic and phenotypic analyses. CONCLUSIONS: Our findings show the importance of the oral microbiota from different niches within the oral cavity as a reservoir for antibiotic resistance. Additionally, the present study showed the need for using more than one method to reveal antibiotic resistance within the total oral biofilm, as a clear mismatch between the shotgun metagenomics method and the phenotypic resistance characterization was shown.

Awareness of electronic crimes related to E-learning among students at the University of Jordan.

The spread of e-learning as an alternative to traditional or face-to-face education has faced many problems and challenges in general and ethical and legal challenges in particular. This study aims to measure students' awareness of the safe use of technology and its tools in e-learning that is consistent with ethical and legal standards. The study attempts to reveal the degree of awareness of students of the University of Jordan about electronic crimes related to e-learning and the legal procedures and penalties related to electronic crimes in e-learning. Quantitative research methods were used. A questionnaire was established and distributed to students enrolled in the following online courses: Ethics and Human Values, Communication Skills, and National Culture. Analysis of the data revealed that students had a high awareness about cybercrime due to the widespread use of the internet by students as it became an integral part of their daily lives. The degree of awareness of student about legal procedures and penalties related to electronic crimes in e-learning was medium. This indicates students' lack of awareness of the effectiveness of procedures and penalties for electronic crimes that can be applied in e-learning due to the rapid transition in the learning process at the University of Jordan from traditional learning to distance e-learning that was imposed during the Corona pandemic. Based on these findings, the study presented a set of recommendations that could be implemented to increase awareness and maximize the benefit of using e-learning.

Influence of the long-term use of oral hygiene products containing stannous ions on the salivary microbiome - a randomized controlled trial.

Oral hygiene products containing tin are suitable to prevent erosive tooth wear, yet effects on the oral microbiota are not known yet. Therefore, this study determined the salivary microbiome of 16 participants using products with stannous ions for three years (TG) compared with a control group (CG) to assess their influence on the microbiota. Participants were included in a randomized controlled clinical trial (RCT) with biannual visits. Illumina Miseq sequencing revealed as most abundant genera: Streptococcus (TG 14.3%; CG 13.0%), Veillonella (TG 11.3%; CG 10.9%), Prevotella (TG 7.0%; CG 9.8%), Haemophilus (TG 6.6%; CG 7.2%), Porphyromonas (TG 5.9%, CG 5.1%), Leptotrichia (TG 5.8%; CG 4.9%), Actinomyces (TG 4.0%; CG 4.6%) and Neisseria (TG 5.4%; CG 4.2%). Beta-Diversity was not significantly different between groups at both time points, although significant differences between groups were found for certain taxa after three years. The genus Prevotella was found in higher abundance in CG whereas Neisseria and Granulicatella, health-associated taxa, were found more abundantly in TG. Salivary microbiota after three years reflected a composition associated with oral health, hence continual use as a preventive measure for dental erosion can be considered safe and benefitting oral health for patients with a high risk of erosion.

Initial Bacterial Adhesion Properties of Anodically Oxidized Ti6Al4V.

This paper reports about the initial interaction of bacteria with anodically oxidized Ti6Al4V for the use as dental implant abutment surfaces. Ti6Al4V samples are anodically oxidized in hydrofluoric acid using different voltages. The resulting nanotopographies are characterized by atomic force microscopy, scanning electron microscopy and contact angle measurements. The topographies reach from micro-porous structures with small nanoporosities on top to fully hexagonally aligned nanotubes. For initial bacterial adhesion tests, Escherichia coli and Staphylococcus aureus are used. Samples are incubated for 2 h and afterwards non-adherent cells are washed off. The results of live/dead staining and cell counts are presented. Gram-negative and Gram-positive strains show different behavior in respect to total number of initially adherent cells on different micro/nanotopographies. The observed reduction of adhered microorganisms is mainly based on underlying microporous topographies.

An oral health optimized diet can reduce gingival and periodontal inflammation in humans - a randomized controlled pilot study.

BACKGROUND: The aim of this pilot study was to investigate the effects of four weeks of an oral health optimized diet on periodontal clinical parameters in a randomized controlled trial. METHODS: The experimental group (n = 10) had to change to a diet low in carbohydrates, rich in Omega-3 fatty acids, and rich in vitamins C and D, antioxidants and fiber for four weeks. Participants of the control group (n = 5) did not change their dietary behavior. Plaque index, gingival bleeding, probing depths, and bleeding upon probing were assessed by a dentist with a pressure-sensitive periodontal probe. Measurements were performed after one and two weeks without a dietary change (baseline), followed by a two week transitional period, and finally performed weekly for four weeks. RESULTS: Despite constant plaque values in both groups, all inflammatory parameters decreased in the experimental group to approximately half that of the baseline values (GI: 1.10 ± 0.51 to 0.54 ± 0.30; BOP: 53.57 to 24.17 %; PISA: 638 mm(2) to 284 mm(2)). This reduction was significantly different compared to that of the control group. CONCLUSION: A diet low in carbohydrates, rich in Omega-3 fatty acids, rich in vitamins C and D, and rich in fibers can significantly reduce gingival and periodontal inflammation. TRIAL REGISTRATION: German Clinical Trials Register; https://www.germanctr.de (DRKS00006301). Registered on 2015-02-21.

Listerine® Products: An Update on the Efficacy and Safety.

UNLABELLED: In the 19th century, the mouthwash Listerine® was formulated from four essential oils. Later, the oils were replaced by their marker substances. To keep them in solution, 24-27% ethanol was added as a vehicle. This is an update of our previous review on the efficacy and safety of Listerine®. METHOD: PubMed was searched for clinical studies on the therapeutic benefits and safety of Listerine® from the end of 2011 to the end of October 2015. RESULTS: Sixteen studies were found and extracted. Three of the four 6-month studies were of sound confirmatory design. Two of these investigated Listerine® and one Listerine Zero®. The evidence of effectiveness for Listerine®, based on the bulk of three confirmatory studies and numerous exploratory studies carried out so far, is strong, but only moderate for Listerine® Zero and poor for Listerine® Cool Blue. In the three safety studies identified, we found methodological flaws that biased the results. CONCLUSIONS: Evidence is accumulating that Listerine® is effective in improving oral health, but the absence of systematic toxicological studies means that an accurate safety assessment cannot be made.

Prevalence of Candida albicans and Candida dubliniensis in caries-free and caries-active children in relation to the oral microbiota-a clinical study.

OBJECTIVES: The correlation between caries and the oral prevalence of Candida spp. in children is contradictory in literature. Thereby, authors focused on Candida albicans as the most isolated Candida species from the oral cavity. Therefore, the aim of the present study was to compare caries-free and caries-bearing children regarding their oral carriage of Candida spp. MATERIAL AND METHODS: Twenty-six caries-free (CF group) and 26 caries-active children (CA group) were included into this study. Three different types of specimens were assessed, saliva and plaque, and in the case of caries, infected dentine samples were microbiologically analyzed for aerobic and anaerobic microorganisms and their counts. Special attention was given to the differentiation between C. albicans and Candida dubliniensis. Additionally, different biochemical tests, VITEK 2 (VITEK®2, bioMérieux, Marcy-l'Etoile, France) and 16S and 18S ribosomal DNA (rDNA) sequencing, were applied for identification. RESULTS: The detection of C. albicans did not differ between the CF and CA groups. C. dubliniensis was never detected in any specimen of the CF group, but occurred in one quarter of the CA group (27 % in plaque, 23 % in saliva), thus leading to a statistically significant difference between the two groups (p < 0.05). In six of these cases, C. dubliniensis was detected concomitantly in saliva and plaque and once only in plaque. CA group harbored statistically more Streptococcus mutans than the control group revealing a correlation between S. mutans and C. dubliniensis regarding the caries group. CONCLUSIONS: This is the first study reporting a frequent detection of C. dubliniensis in caries-active children, which could have been underestimated so far due to difficulties in differentiation between this yeast species and C. albicans. CLINICAL RELEVANCE: Microbiological diagnostic-especially of oral Candida species-is an important determinant for identifying etiological factors of dental caries in children.

Antimicrobial Photoinactivation Using Visible Light Plus Water-Filtered Infrared-A (VIS + wIRA) Alters In Situ Oral Biofilms.

Recently, growing attention has been paid to antimicrobial photodynamic therapy (aPDT) in dentistry. Changing the microbial composition of initial and mature oral biofilm by aPDT using visible light plus water-filtered infrared-A wavelengths (VIS + wIRA) has not yet been investigated. Moreover, most aPDT studies have been conducted on planktonic bacterial cultures. Therefore, in the present clinical study we cultivated initial and mature oral biofilms in six healthy volunteers for 2 hours or 3 days, respectively. The biofilms were treated with aPDT using VIS+wIRA (200 mW cm(-2)), toluidine blue (TB) and chlorine e6 (Ce6) for 5 minutes. Chlorhexidine treated biofilm samples served as positive controls, while untreated biofilms served as negative controls. After aPDT treatment the colony forming units (CFU) of the biofilm samples were quantified, and the surviving bacteria were isolated in pure cultures and identified using MALDI-TOF, biochemical tests and 16S rDNA-sequencing. aPDT killed more than 99.9% of the initial viable bacterial count and 95% of the mature oral biofilm in situ, independent of the photosensitizer. The number of surviving bacterial species was highly reduced to 6 (TB) and 4 (Ce6) in the treated initial oral biofilm compared to the 20 different species of the untreated biofilm. The proportions of surviving bacterial species were also changed after TB- and Ce6-mediated aPDT of the mature oral biofilm, resulting in a shift in the microbial composition of the treated biofilm compared to that of the control biofilm. In conclusion, aPDT using VIS + wIRA showed a remarkable potential to eradicate both initial and mature oral biofilms, and also to markedly alter the remaining biofilm. This encourages the clinical use of aPDT with VIS + wIRA for the treatment of periimplantitis and periodontitis.

A Preliminary Investigation on the Antimicrobial Activity of Listerine®, Its Components, and of Mixtures Thereof.

Listerine® is one of the most popular mouthwashes worldwide and claims to combat harmful bacteria. In the past century, its recipe was changed from an essential oil mouthwash to a five-component mixture (thymol, menthol, eucalyptol, and methyl salicylate dissolved in 27% ethanol). The aim of this study was to get preliminary information about the antimicrobial activities of individual Listerine® components and their mixtures. We tested the bacterial strains Streptococcus mutans, Enterococcus faecalis, and Eikenella corrodens and the yeast Candida albicans. The established minimum inhibitory concentration (MIC) assay and the minimum bactericidal/fungicidal concentration (MBC/MFC) assay were applied. None of the combinations of two phenols at the concentrations contained within Listerine® were associated with either an additive or synergistic effect. Thymol had lower MIC and MBC/MFC values than the other Listerine® components and Listerine® against E. corrodens and C. albicans. The mixtures consisting of eucalyptol, methyl salicylate, and thymol were the most effective against S. mutans and E. faecalis and more effective than Listerine®. Our results demonstrate that the phenols and their concentrations as contained within Listerine® could be further optimized in terms of selecting those which increase their general effectiveness, at concentrations that do not induce harm.

Ultrasonic activation and chemical modification of photosensitizers enhances the effects of photodynamic therapy against Enterococcus faecalis root-canal isolates.

BACKGROUND: The aim of this study was to evaluate the effect of photodynamic therapy (PDT) on Enterococcus faecalis biofilms in artificially infected root canals using modified photosensitizers and passive ultrasonic activation. METHODS: Two hundred and seventy extracted human teeth with one root canal were instrumented utilizing ProTaper files, autoclaved, infected with E. faecalis T9 for 72 h and divided into different groups: irrigation with 3% sodium hypochlorite (NaOCl), 20% ethylenediaminetetraacetic acid (EDTA), or 20% citric acid, PDT without irrigation, PDT accompanied by irrigation with NaOCl, EDTA, or citric acid, PDT using an EDTA-based photosensitizer or a citric-acid-based photosensitizer and PDT with ultrasonic activation of the photosensitizer. A 15 mg/ml toluidine blue served as the photosensitizer, activated by a 100 mW LED light source. Sterile paper points were used for sampling the root canals and dentin chips were collected to assess the remaining contamination after treatment. Samples were cultured on blood agar plates and colony forming units were quantified. RESULTS: PDT alone achieved a reduction in E. faecalis counts by 92.7%, NaOCl irrigation alone and combined with PDT by 99.9%. The antibacterial effects increased by the combination of irrigation using EDTA or citric acid and PDT compared to irrigation alone. More than 99% of E. faecalis were killed using PDT with the modified photosensitizers and ultrasonic activation. CONCLUSIONS: NaOCl based disinfection achieved the highest antimicrobial effect. Using PDT with an EDTA-based or citric-acid-based phozosensitizer or activating the photosensitizer with ultrasound resulted in a significantly higher reduction in E. faecalis counts compared to conventional PDT.

Supplementary sampling of obturation materials enhances microbial analysis of endodontic treatment failures: a proof of principle study.

OBJECTIVES: Root canal treatment failures often correlate with persistent biomaterial-associated endodontic infections. The aim of the present study was to assess the impact of endodontic obturation material sampling from root canals with posttreatment apical periodontitis on improving standard study protocols. MATERIALS AND METHODS: Samples from previously filled root canals and their corresponding endodontic filling materials were obtained from five root-filled teeth with posttreatment periradicular lesions. After cultivation, the isolated microorganisms were quantified and biochemically identified. Moreover, clone libraries were constructed after the amplification of bacterial 16S ribosomal DNA (rDNA) from the same samples. DNA from selected clones was sequenced to identify microbial species. Transmission electron microscopy (TEM) aided visualization of the detected bacteria. RESULTS: Overall, 22 taxa of the phyla Firmicutes, Actinobacteria, and Bacteroidetes were detected in both obturation and root canal samples by culture-dependent and culture-independent methods. Root canal fillings sheltered 17 species (3.30-7.50 × 10(3) CFU/ml). Of these, nine were detected solely in the retrieved obturation materials. The reinfected root canals harbored 13 taxa (3.48-7.36 × 10(3) CFU/ml). Obligate and facultative anaerobic bacteria prevailed. The number of different species ranged from 1 to 5 within a single sample. Fungi were not detected. CONCLUSIONS: Bacteria can colonize both root canals and endodontic fillings in vivo. CLINICAL RELEVANCE: Integrating the sampling of obturation materials with standard root canal sample collection offers a clearer insight into the actual microbial flora of reinfected root canals and improves the study protocols of secondary/persistent endodontic infections.

Novel Broad-Spectrum Antimicrobial Photoinactivation of In Situ Oral Biofilms by Visible Light plus Water-Filtered Infrared A.

Antimicrobial photodynamic therapy (APDT) has gained increased attention as an alternative treatment approach in various medical fields. However, the effect of APDT using visible light plus water-filtered infrared A (VIS + wIRA) on oral biofilms remains unexplored. For this purpose, initial and mature oral biofilms were obtained in situ; six healthy subjects wore individual upper jaw acrylic devices with bovine enamel slabs attached to their proximal sites for 2 h or 3 days. The biofilms were incubated with 100 µg ml(-1) toluidine blue O (TB) or chlorin e6 (Ce6) and irradiated with VIS + wIRA with an energy density of 200 mW cm(-2) for 5 min. After cultivation, the CFU of half of the treated biofilm samples were quantified, whereas following live/dead staining, the other half of the samples were monitored by confocal laser scanning microscopy (CLSM). TB- and Ce6-mediated APDT yielded a significant decrease of up to 3.8 and 5.7 log10 CFU for initial and mature oral biofilms, respectively. Quantification of the stained photoinactivated microorganisms confirmed these results. Overall, CLSM revealed the diffusion of the tested photosensitizers into the deepest biofilm layers after exposure to APDT. In particular, Ce6-aided APDT presented elevated permeability and higher effectiveness in eradicating 89.62% of biofilm bacteria compared to TB-aided APDT (82.25%) after 3 days. In conclusion, antimicrobial photoinactivation using VIS + wIRA proved highly potent in eradicating oral biofilms. Since APDT excludes the development of microbial resistance, it could supplement the pharmaceutical treatment of periodontitis or peri-implantitis.

In vivo study of the initial bacterial adhesion on different implant materials.

OBJECTIVE: Biofilm formation on implant materials plays a major role in the aetiology of periimplantitis. The aim of this study was to examine in vivo the initial bacterial adhesion on six different implant materials. METHODS: The implant materials Ti-m, TiUnite®, ZiUnite®, ATZ-m, ATZ-s, TZP-A-m were tested using bovine enamel slabs as controls. All materials, fixed on splint systems, were examined after 30 min and 120 min of oral exposure. DAPI staining was used for quantitative analysis of the initially adherent microorganisms. Initial adherent microorganisms were visualised by fluorescence In situ-hybridisation (FISH) and quantified by confocal laser scanning microscopy (CLSM). The targets of the oligonucleotide probes were Eubacteria, Veillonella spp., Fusobacterium nucleatum, Actinomyces naeslundii and Streptococcus spp. RESULTS: DAPI analysis showed that increasing the time of oral exposure resulted in an increasing amount of initial adherent bacteria. The highest level of colonisation was on ZiUnite®, with the lowest occurring on the bovine enamel, followed by Ti-m. This early colonisation correlated significantly with the surface roughnesses of the materials. FISH and CLSM showed no significant differences relating to total bacterial composition. However, Streptococcus spp. was shown to be the main colonisers on each of the investigated materials. CONCLUSION: it could be shown that within an oral exposure time of 30 min and 120 min, despite the salivary acquired pellicle initial biofilm formation is mainly influenced directly or indirect by the material surface topography. Highly polished surfaces should minimise the risk of biofilm formation, plaque accumulation and possibly periimplantitis.

Influence of a mouthwash containing hydroxyapatite microclusters on bacterial adherence in situ.

OBJECTIVE: The aim of the present study was to investigate the efficacy of a new preparation in dental prophylaxis containing zinc-carbonate hydroxyapatite microclusters (Biorepair) for oral biofilm management. METHODS AND MATERIALS: Initial biofilm formation was carried out in situ with bovine enamel slabs fixed to individual upper jaw splints worn by six subjects. Rinses with the customary preparation as well as with subfractions (hydroxyapatite microclusters in saline solution; liquid phase without particles) were adopted for 1 min in situ after 1 min of pellicle formation, and the bacterial colonization was recorded after 6 h and 12 h, respectively. Rinses with chlorhexidine served as a reference. The adherent microorganisms were quantified and visualized using DAPI staining and live-dead staining (BacLight). Furthermore, the effects on Streptococcus mutans bacteria were tested in vitro (BacLight). RESULTS: Application of the customary preparation and of the separate components distinctly reduced the initial bacterial colonization of the enamel surface in situ as visualized and quantified with all techniques. After 12 h, 1.3 × 10(7) ± 2.0 × 10(7) bacteria/cm² were detected on unrinsed control samples with DAPI staining; 2.4 × 10(6) ± 3.3 × 10(6) after application of Biorepair (12 h after CHX-rinse; 1.3 × 10(5) ± 9.2 × 10(4)). Also, pure hydroxyapatite microclusters in saline solution (2.1 × 10(6) ± 3.0 × 10(6)) as well as the liquid phase without particles (5.1 × 10(5) ± 3.3 × 10(5)) reduced the amount of adherent bacteria. Furthermore, antimicrobial effects on S. mutans were observed in vitro. CONCLUSION: The preparation is an effective compound for biofilm management in the oral cavity due to antiadherent and antibacterial effects. CLINICAL RELEVANCE: The tested mouthrinse seems to be a reasonable amendment for dental prophylaxis.

Antimicrobial photodynamic therapy using visible light plus water-filtered infrared-A (wIRA).

The aim of this study was to investigate the effectiveness of antimicrobial photodynamic therapy (APDT) using visible light together with water-filtered infrared-A (VIS+wIRA) to eradicate single species of planktonic bacteria and micro-organisms during initial oral bacterial colonization in situ. A broadband VIS+wIRA radiator with a water-filtered spectrum in the range 580-1400 nm was used for irradiation. Toluidine blue (TB) was utilized as a photosensitizer at concentrations of 5, 10, 25 and 50 µg ml(-1). The unweighted (absolute) irradiance was 200 mW cm(-2) and it was applied for 1 min. Planktonic cultures of Streptococcus mutans and Enterococcus faecalis were treated with APDT. Salivary bacteria harvested by centrifugation of native human saliva were also tested. In addition, initial bacterial colonization of bovine enamel slabs carried in the mouths of six healthy volunteers was treated in the same way. Up to 2 log(10) of S. mutans and E. faecalis were killed by APDT. Salivary bacteria were eliminated to a higher extent of 3.7-5 log(10). All TB concentrations tested proved to be highly effective. The killing rate of bacteria in the initial oral bacterial colonization was significant (P=0.004) at all tested TB concentrations, despite the interindividual variations found among study participants. This study has shown that APDT in combination with TB and VIS+wIRA is a promising method for killing bacteria during initial oral colonization. Taking the healing effects of wIRA on human tissue into consideration, this technique could be helpful in the treatment of peri-implantitis and periodontitis.

Comparison of different live/dead stainings for detection and quantification of adherent microorganisms in the initial oral biofilm.

OBJECTIVES: The aim of the present study was to investigate different fluorescence-based, two-color viability assays for visualization and quantification of initial bacterial adherence and to establish reliable alternatives to the ethidium bromide staining procedure. MATERIALS AND METHODS: Bacterial colonization was attained in situ on bovine enamel slabs (n = 6 subjects). Five different live/dead assays were investigated (fluorescein diacetate (FDA)/propidium iodide (PI), Syto 9/PI (BacLight®), FDA/Sytox red, Calcein acetoxymethyl (AM)/Sytox red, and carboxyfluorescein diacetate (CFDA)/Sytox red). After 120 min of oral exposure, analysis was performed with an epifluorescence microscope. Validation was carried out, using the colony-forming units for quantification and the transmission electron microscopy for visualization after staining. RESULTS: The average number of bacteria amounted to 2.9 ± 0.8 × 10(4) cm(-2). Quantification with Syto 9/PI and Calcein AM/Sytox red yielded an almost equal distribution of cells (Syto 9/PI 45% viable, 55% avital; Calcein AM/Sytox red 52% viable, 48% avital). The live/dead ratio of CFDA/Sytox red and FDA/Sytox red was 3:2. An aberrant dispersal was recorded with FDA/PI (viable 34%, avital 66%). The TEM analysis indicated that all staining procedures affect the structural integrity of the bacterial cells considerably. CONCLUSION: The following live/dead assays are reliable techniques for differentiation of viable and avital adherent bacteria: BacLight, FDA/Sytox red, Calcein AM/Sytox red, and CFDA/Sytox red. These fluorescence-based techniques are applicable alternatives to toxic and instable conventional assays, such as the staining procedure based on ethidium bromide. CLINICAL RELEVANCE: Differentiation of viable and avital adherent bacteria offers the possibility for reliable evaluation of different mouth rinses, oral medication, and disinfections.

Microscope-based imaging platform for large-scale analysis of oral biofilms.

A microscopic method for noninvasively monitoring oral biofilms at the macroscale was developed to describe the spatial distribution of biofilms of different bacterial composition on bovine enamel surfaces (BES). For this purpose, oral biofilm was grown in situ on BES that were fixed at approximal sites of individual upper jaw acrylic devices worn by a volunteer for 3 or 5 days. Eubacteria, Streptococcus spp., and Fusobacterium nucleatum were stained using specific fluorescence in situ hybridization (FISH) probes. The resulting fluorescence signals were subsequently tested by confocal laser scanning microscopy (CLSM) and monitored by an automated wide-field microscope-based imaging platform (Scan∧R). Automated image processing and data analysis were conducted by microscope-associated software and followed by statistical evaluation of the results. The full segmentation of biofilm images revealed a random distribution of bacteria across the entire area of the enamel surfaces examined. Significant differences in the composition of the microflora were recorded across individual as well as between different enamel surfaces varying from sparsely colonized (47.26%) after 3 days to almost full surface coverage (84.45%) after 5 days. The enamel plates that were positioned at the back or in the middle of the oral cavity were found to be more suitable for the examination of biofilms up to 3 days old. In conclusion, automated microscopy combined with the use of FISH can enable the efficient visualization and meaningful quantification of bacterial composition over the entire sample surface. Due to the possibility of automation, Scan∧R overcomes the technical limitations of conventional CLSM.

Is Helicobacter pylori resident or transient in the human oral cavity?

Helicobacter pylori colonizes the stomachs of at least half of the world's human population. The role of the oral cavity in this colonization is not clear and there are, to date, no comprehensive data that clearly demonstrate the isolation of this bacterium from the oral cavity. The aim of this study was to evaluate the prevalence of H. pylori in the oral cavity of 15 patients who tested positive for H. pylori. A comprehensive dental examination of all patients was conducted. Samples were taken from supragingival and subgingival plaque, saliva, periapical exudates and tongue swabs. All samples were taken before the application of antibiotics. A total of 163 oral samples were investigated by PCR using two different H. pylori-specific primer pairs. A PCR inhibition control using a modified plasmid was always included for the most specific primer pair. In addition, a culture technique was used to confirm PCR results. Despite a PCR detection limit of 10(2) bacteria ml(-1), out of 14 patients, H. pylori could not be detected in any of the samples taken. In one patient, H. pylori-positive PCR signals were obtained in two samples using only one primer pair. H. pylori could not be cultivated from these two PCR-positive samples; therefore, no correlation to oral colonization status could be established. This study challenges the misleading preconception that H. pylori resides in the human oral cavity and suggests that this bacterium should be considered transient and independent of the oral status. To date, positive PCR results for H. pylori in the oral cavity have been overestimated and not critically interpreted in literature.

Airborne microbes in different dental environments in comparison to a public area.

OBJECTIVE: The aim of this study was to investigate quantitatively and qualitatively the airborne microbial load in a multi-chair dental clinic, a normal dental practice and a non-dental public area over a time period of four days and at different time points to estimate the risk of infections during dental surgery. METHODS: A multi-chair and a single chair treatment room each were examined in comparison to a non-medical public area over a period of four days. The colony forming units m(-3) (CFUs) were determined and isolated bacteria were characterised by morphological and biochemical analysis, gas chromatography and by 16S rRNA-gene sequencing. In the analyses enterococci were selectively searched for. RESULTS: The CFUs in the multi-chair treatment room were between 20 and 1050 CFU m(-3). During treatment the maxima reached were below 800 CFU m(-3). The values in the dental practice were between 200 and 600 CFU m(-3) and remain slightly but not significantly below the levels of the clinic (p > 0.05). In the common area, the CFUs were between 200 and 800 CFU m(-3). The proportion of micrococci was 56.8% in the clinic, 56.07% in the practice and 69.67% in the public area Coagulase-negative staphylococci constituted 35% at the dental clinic, 25% at the bank and 38% at the dental practice. No significant differences amongst the units were detected in the microbial composition of their dental aerosols (p > 0.05). CONCLUSION: Although, the bacterial counts in dental room were not significantly higher than the bacterial counts in a public area, the risk from dental clinic might be higher than a public area due to the type of micro-organisms, host susceptibility and the exposure time.