[Medicaments and oral care: Is there still a rationale for clinical periodontal microbiology?] / Serie: Medicamenten en mondzorg. Is er nog indicatie voor bacteriologisch onderzoek bij parodontitis?

Clinical microbiology now occupies an important place in periodontics and oral implant dentistry as a supplementary diagnostic tool and in planning treatment, particularly with respect to the rational use of antibiotics. This view is in line with the emphatic call by the World Health Organization and the European Union for the prudent use of antibiotics due to the global increase in resistance to antibiotics. Furthermore side effects may occur, such as the disturbance of the microbial intestinal and oral microflora, sometimes leading to serious pathological conditions. Hyposalivation following the use of antibiotics may lead to an oral environmental condition in which caries may develop faster than usual.

[A renewed focus on extra-oral manifestations of oral infections]. / Hernieuwde aandacht voor extraorale gevolgen van ontstekingen in de mond.

Dental focal infections are extraoral manifestations caused by oral pathogens. Pathological oral conditions, such as periapical inflammation and periodontitis, can cause bacteremia. Dissemination of oral pathogens to nonoral sites can subsequently cause infections in extraoral tissues and organs. Cardiovascular infections and brain abscesses are the most common of these. The course of such infections can be lethal. In order to improve patient care, a closer collaboration between dental and medical caregivers is necessary.

Conserved Citrullinating Exoenzymes in Porphyromonas Species.

Porphyromonas gingivalis is one of the major oral pathogens implicated in the widespread inflammatory disorder periodontitis. Moreover, in recent years, P. gingivalis has been associated with the autoimmune disease rheumatoid arthritis. The peptidylarginine deiminase enzyme of P. gingivalis (PPAD) is a major virulence factor that catalyzes the citrullination of both bacterial and host proteins, potentially contributing to production of anticitrullinated protein antibodies. Considering that these antibodies are very specific for rheumatoid arthritis, PPAD appears to be a link between P. gingivalis, periodontitis, and the autoimmune disorder rheumatoid arthritis. PPAD was thus far considered unique among prokaryotes, with P. gingivalis being the only bacterium known to produce and secrete it. To challenge this hypothesis, we investigated the possible secretion of PPAD by 11 previously collected Porphyromonas isolates from a dog, 2 sheep, 3 cats, 4 monkeys, and a jaguar with periodontitis. Our analyses uncovered the presence of secreted PPAD homologues in 8 isolates that were identified as Porphyromonas gulae (from a dog, monkeys, and cats) and Porphyromonas loveana (from sheep). In all 3 PPAD-producing Porphyromonas species, the dominant form of the secreted PPAD was associated with outer membrane vesicles, while a minor fraction was soluble. Our results prove for the first time that the citrullinating PPAD exoenzyme is not unique to only 1 prokaryotic species. Instead, we show that PPAD is produced by at least 2 other oral pathogens.

Microbial Characteristics of Peri-Implantitis: A Case-Control Study.

BACKGROUND: The aim of this case-control study is to compare oral microbiologic characteristics of patients with healthy peri-implant conditions and patients with peri-implantitis and to explore the influence of various patient- and implant-related factors on microbiologic characteristics. METHODS: Peri-implant submucosal microbial samples were collected from 85 patients with peri-implantitis (cases) and from 69 patients with only implants with healthy peri-implant conditions (controls). Samples were analyzed using culturing techniques. Multivariable logistic regression was used to explore the association of disease status and various patient- and implant-related factors (sex, patient age, smoking, number of remaining teeth, percentage of teeth with bone loss, implant function time, implant surface, and presence of plaque) with microbiologic characteristics. RESULTS: Peri-implant disease status was significantly associated with the submucosal presence of Porphyromonas gingivalis (Pg), Prevotella intermedia (Pi), Tannerella forsythia (Tf), and Fusobacterium nucleatum (Fn). The association with disease status was most obvious for Pi (odds ratio [OR]: 15.1; 95% confidence interval [CI]: 5.1 to 45.3) and Tf (OR: 13.3; 95% CI: 5.4 to 32.5). The prevalence of Aggregatibacter actinomycetemcomitans and Staphylococcus species was very low. CONCLUSIONS: The periodontal pathogens Pg, Pi, Tf, and Fn are associated with peri-implantitis. A. actinomycetemcomitans and Staphylococcus species do not seem to play an important role in peri-implantitis.

Bacterial colonization of the peri-implant sulcus in dentate patients: a prospective observational study.

OBJECTIVES: The aim of the present study was to compare the composition of the periodontal microflora at baseline (T0) with the submucosal microflora at least 1 year after implant placement (T1) in periodontally healthy patients. MATERIAL AND METHODS: For all 169 consecutive patients that visited our clinic during 1 year, we determined their periodontal parameters, implant mucosal index, and presence of implant calculus. At T0, self-reported smoking status was recorded and subgingival and submucosal biofilm samples were obtained and analyzed for the presence and numbers of selected periodontal pathogens. All measurements were repeated at T1. RESULTS: One hundred twenty patients completed the study. Periodontal parameters were stable or had improved at T1. The total bacterial load was lower at implant sites (P < 0.05). The prevalence of Porphyromonas gingivalis was low at baseline, but at T1, detection rate and numbers were higher at implant sites compared to dentate sites. At T1, the frequency of detection of P. gingivalis (P = 0.01), Parvimonas micra (P = 0.018), and Fusobacterium nucleatum (P = 0.035) was higher in smoking patients (n = 23) than in non-smokers (n = 97). CONCLUSIONS: Colonization of the submucosal peri-implant area is similar to the composition of subgingival microbiota. Smoking has a measurable effect on the colonization of implant-associated biofilms and may select for P. gingivalis, P. micra, and F. nucleatum. CLINICAL RELEVANCE: The colonization of implants by well-known periodontal pathogens is very similar to that in normal dentition, also in a healthy cohort. Smoking status was related with the prevalence of periodontal pathogens where smokers harbored more often periodontal pathogens such as P. gingivalis, P. micra, and F. nucleatum.

[Medicines and oral health care 2. An optimised antibiotic policy in the interest of the patient]. / Medicamenten en mondzorg 2. Een geoptimaliseerd antibioticabeleid in het belang van de patiënt.

Treatment of infectious diseases is complex and success is dependent upon a number of factors. The proper prescription and correct dosage of antimicrobials are important elements in this respect. Incorrect prescription and insufficient dosage can result not only in treatment failure but also contribute to the development of antimicrobial resistance. In order to achieve correct antibiotic dosages it is necessary to gain insight in pharmacokinetic and pharmacodynamic principles. It is also vital to differentiate among various types of infections and to adjust antibiotic prescription to the clinical situation of the individual patient accordingly. To further optimise antibiotic policy collaboration between dental and medical professions is of major importance.

The optimization and validation of the Biotyper MALDI-TOF MS database for the identification of Gram-positive anaerobic cocci.

Gram-positive anaerobic cocci (GPAC) account for 24%-31% of the anaerobic bacteria isolated from human clinical specimens. At present, GPAC are under-represented in the Biotyper MALDI-TOF MS database. Profiles of new species have yet to be added. We present the optimization of the matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) database for the identification of GPAC. Main spectral profiles (MSPs) were created for 108 clinical GPAC isolates. Identity was confirmed using 16S rRNA gene sequencing. Species identification was considered to be reliable if the sequence similarity with its closest relative was ≥98.7%. The optimized database was validated using 140 clinical isolates. The 16S rRNA sequencing identity was compared with the MALDI-TOF MS result. MSPs were added from 17 species that were not yet represented in the MALDI-TOF MS database or were under-represented (fewer than five MSPs). This resulted in an increase from 53.6% (75/140) to 82.1% (115/140) of GPAC isolates that could be identified at the species level using MALDI-TOF MS. An improved log score was obtained for 51.4% (72/140) of the strains. For strains with a sequence similarity <98.7% with their closest relative (n = 5) or with an inconclusive sequence identity (n = 4), no identification was obtained by MALDI-TOF MS or in the latter case an identity with one of its relatives. For some species the MSP of the type strain was not part of the confined cluster of the corresponding clinical isolates. Also, not all species formed a homogeneous cluster. It emphasizes the necessity of adding sufficient MSPs of human clinical isolates.

Effect of leaving chronic oral foci untreated on infectious complications during intensive chemotherapy.

BACKGROUND: Leukaemic patients receiving intensive chemotherapy and patients undergoing autologous stem-cell transplantation (ASCT) are routinely screened for oral foci of infection to reduce infectious complications that could occur during therapy. In this prospective study we assessed the effect of leaving chronic oral foci of infection untreated on the development of infectious complications in intensively treated haematological patients. METHODS: We included and prospectively evaluated all intensively treated leukaemic patients and patients undergoing ASCT who were referred to our medical centre between September 2012 and May 2014, and who matched the inclusion/exclusion criteria. Acute oral foci of infection were removed before chemotherapy or ASCT, whereas chronic oral foci were left untreated. RESULTS: In total 28 leukaemic and 35 ASCT patients were included. Acute oral foci of infection were found in 2 leukaemic (7%) and 2 ASCT patients (6%), and chronic oral foci of infection in 24 leukaemic (86%) and 22 ASCT patients (63%). Positive blood cultures with microorganisms potentially originating from the oral cavity occurred in 7 patients during treatment, but were uneventful on development of infectious complications. CONCLUSIONS: Our prospective study supports the hypothesis that chronic oral foci of infection can be left untreated as this does not increase infectious complications during intensive chemotherapy.

Anaerococcus nagyae sp. nov., isolated from human clinical specimens.

We describe a new Anaerococcus species isolated from human clinical specimens. Analyses of 16S rRNA gene sequences of three strains showed <98% similarity with its closest relative Anaerococcus octavius. Phylogenetically the isolated strains form a cluster and can be differentiated from other species of the genus Anaerococcus based on its phenotypic characteristics and its MALDI-TOF MS profile. We propose the name Anaerococcus nagyae, with A. nagyae DSM101193 (accession number KU043522) as the type strain.

[Periodontitis and systemic diseases: from science to clinical practice]. / Parodontitis en systemische ziekten - van wetenschap naar praktijk.

The evidence for an association between systemic diseases and periodontitis is strongest with diabetes mellitus type 2 and cardiovascular disease. There is a moderate association of periodontitis with adverse pregnancy outcomes and rheumatoid arthritis. Periodontal treatment has, on average, a positive effect on reducing systemic infection and improving the condition of the vascular system. For diabetes patients, periodontal treatment can also have a positive effect on metabolic regulation. There is insufficient evidence that periodontal treatment prevents adverse pregnancy outcomes and rheumatoid arthritis.

Anaerococcus degenerii sp. nov., isolated from human clinical specimens.

Four clinical isolates of gram-positive strict anaerobic cocci were isolated from four different human mixed anaerobic infections. The taxonomical status of the four strains could not be established using standard identification techniques. The biochemical features of the strains were established and their taxonomic position was determined using 16S rRNA sequencing. The four strains form a homogeneous phenotypical and genotypical cluster. A new Anaerococcus species is proposed for these isolates: Anaerococcus degenerii sp. nov. The type strain is UMCG-104(T) = DSM29674(T) (accession number AM176528).

Long-term efficacy of a 0.07% cetylpyridinium chloride mouth rinse in relation to plaque and gingivitis: a 6-month randomized, vehicle-controlled clinical trial.

OBJECTIVE: To evaluate the effectiveness of 0.07% cetylpyridinium chloride (CPC) mouth rinse for reduction of gingival inflammation and inhibition of plaque compared to a vehicle control (VC) mouth rinse over a 6-month period. MATERIALS & METHODS: Participants (n = 62) used their randomly assigned product as adjunct to toothbrushing. Bleeding, plaque and staining scores were assessed at baseline, 3 and 6 months. Plaque and saliva samples were taken at each assessment monitoring possible shifts in the composition of the microbiota. RESULTS: A significant difference (P = 0.002) in favour of the CPC mouth rinse, with respect to plaque scores, was found. Bleeding scores at 6 months were not significantly different (P = 0.089). However, when correcting for baseline values, a tendency towards a significant difference in bleeding scores at end trail was observed in favour of the CPC mouth rinse (P = 0.061). Regarding staining at 3 and 6 months, a small but significant difference (8.6% and 10.4%, respectively) (P < 0.0001) was observed with lower scores for the VC group. There was a significant reduction in total anaerobic count in the CPC group at 6 months (P < 0.05). The ratio of aerobes/anaerobes was markedly increased at 3 months, especially in the CPC group. No further differences were observed between groups at 6 months. CONCLUSIONS: The use of 0.07% CPC mouth rinse was significantly more effective in reducing plaque scores than the vehicle control. Bleeding scores were not different at 6 months. The test product was well accepted and did not cause any serious clinical side effects or negatively affected the microbiota.

Antibiotic susceptibility profiles of anaerobic pathogens in The Netherlands.

The antibiotic susceptibility profile of the Bacteroides fragilis group, Gram-positive anaerobic cocci (GPAC), Fusobacterium spp., Prevotella spp., Veillonella spp. and Bilophila wadsworthia for amoxicillin, amoxicillin-clavulanic acid, clindamycin and metronidazole was determined. Human clinical isolates were isolated between 2011 and 2013 at the Microbiological Diagnostic Laboratory of the University Medical Center Groningen, The Netherlands and subjected to MALDI-TOF MS identification and susceptibility testing using E-test for MIC determination. Differences in clindamycin susceptibility between species of the B. fragilis group and GPAC were observed, with Bacteroides ovatus and Peptoniphilus harei having the highest resistance rates. Compared to other European countries, in The Netherlands the MIC90 for clindamycin of fusobacteria is low. Metronidazole resistance was first encountered in the genus Prevotella in 2013, but not in species of GPAC as reported in Belgium and Bulgaria. The differences in clindamycin resistance between the different European countries and reports of metronidazole resistance within the genera Prevotella and GPAC warrant more extensive susceptibility studies on anaerobic pathogens.

Implant decontamination with 2% chlorhexidine during surgical peri-implantitis treatment: a randomized, double-blind, controlled trial.

OBJECTIVE: The objective of this randomized, double-blind, controlled trial was to evaluate the clinical, radiographic, and microbiological effects of implant surface decontamination with a 2% chlorhexidine (CHX) solution in comparison with a 0.12% chlorhexidine + 0.05% cetylpyridinium chloride (CPC) solution during resective surgical peri-implantitis treatment. MATERIAL AND METHODS: Forty-four patients (108 implants) with peri-implantitis were treated with resective surgical treatment consisting of bone re-contouring, surface debridement and chemical decontamination, and apically repositioned flap. Patients were randomly allocated to decontamination with a 2% CHX solution (test group) or 0.12% CHX + 0.05% CPC (control group). Clinical and radiographic parameters were recorded before treatment (baseline), and at 3, 6, and 12 months after treatment. Microbiological parameters were recorded during surgery. RESULTS: Multilevel analysis showed no significant differences in bleeding, suppuration, probing pocket depth, and radiographic bone loss between control and test group over three follow-up measurements (3, 6, and 12 months) from baseline. Both decontamination procedures resulted in significant reductions in anaerobic bacterial counts on the implant surface, but no significant difference was noted between control and test group (mean log 3.37 ± 2.34 vs. 3.65 ± 2.87, P = 0.99). CONCLUSIONS: The use of a 2% CHX solution for implant surface decontamination during resective peri-implantitis therapy does not lead to improved clinical, radiographic, or microbiological results compared with a 0.12% CHX + 0.05% CPC solution. Overall, the additional use of CHX reduces anaerobic bacterial load on the implant surface better than mechanical debridement alone, but does not seem to enhance clinical treatment outcomes (ClinicalTrials.gov number NCT01852253).

The influence of incubation time, sample preparation and exposure to oxygen on the quality of the MALDI-TOF MS spectrum of anaerobic bacteria.

With matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), bacteria can be identified quickly and reliably. This accounts especially for anaerobic bacteria. Because growth rate and oxygen sensitivity differ among anaerobic bacteria, we aimed to study the influence of incubation time, exposure to oxygen and sample preparation on the quality of the spectrum using the Bruker system. Also, reproducibility and inter-examiner variability were determined. Twenty-six anaerobic species, representing 17 genera, were selected based on gram-stain characteristics, growth rate and colony morphology. Inter-examiner variation showed that experience in the preparation of the targets can be a significant variable. The influence of incubation time was determined between 24 and 96 h of incubation. Reliable species identification was obtained after 48 h of incubation for gram-negative anaerobes and after 72 h for gram-positive anaerobes. Exposure of the cultures to oxygen did not influence the results of the MALDI-TOF MS identifications of all tested gram-positive species. Fusobacterium necrophorum and Prevotella intermedia could not be identified after >24 h and 48 h of exposure to oxygen, respectively. Other tested gram-negative bacteria could be identified after 48 h of exposure to oxygen. Most of the tested species could be identified using the direct spotting method. Bifidobacterium longum and Finegoldia magna needed on-target extraction with 70% formic acid in order to obtain reliable species identification and Peptoniphilus ivorii a full extraction. Spectrum quality was influenced by the amount of bacteria spotted on the target, the homogeneity of the smear and the experience of the examiner.

In vitro antimicrobial effects of two antihalitosis mouth rinses on oral pathogens and human tongue microbiota.

OBJECTIVES: The aim of the study was to compare the antimicrobial activity of a mouth rinse containing chlorhexidine and cetylpyridinium chloride (MR1) with a stannous fluoride-based mouth rinse (MR2) in vitro. MATERIALS AND METHODS: Samples of the tongues from 10 subjects with and 10 subjects without halitosis were inoculated on blood agar plates. The agar was perforated, and the cylindrical holes were filled either with mouth rinse MR1 or with mouth rinse MR2. After incubation, inhibition zones of the whole tongue microbiota and Fusobacterium nucleatum were measured. In addition, MR1 and MR2 were applied in a short interval killing test (SIKT) on four oral pathogens Porphyromonas gingivalis, Prevotella intermedia, F. nucleatum and Aggregatibacter actinomycetemcomitans. Total viable cell counts were made after two minutes of incubation with increasing concentrations of MR1 and MR2. RESULTS: MR1 showed a significantly higher in vitro antimicrobial activity against the whole tongue microbiota and F. nucleatum than MR2 in both groups of subjects. In the SIK test, MR1 showed a significantly greater killing capacity than MR2. The results show that a mouth rinse with low concentrations of chlorhexidine and 0.05% cetylpyridinium chloride appears to be more effective in inhibiting growth of the human tongue microbiota in vitro than a fluoride/stannous fluoride-containing mouth rinse. CONCLUSION: This in vitro observation supports the use of chlorhexidine and cetylpyridinium chloride in the treatment of oral halitosis.

Evaluation of three selective media for isolation of Aggregatibacter actinomycetemcomitans.

BACKGROUND AND OBJECTIVE: Aggregatibacter actinomycetemcomitans is a pathogen in oral and nonoral infections. Detection and quantification of this pathogen can be performed using selective culture techniques. The aim of this study was to establish the efficacy of two known selective media in their ability to select and support the growth of A. actinomycetemcomitans. MATERIAL AND METHODS: Trypticase soy bacitracin vancomycin (TSBV) medium and brain-heart infusion agar with vancomycin (Dentaid-1), as well as a modified Dentaid-1 medium (in which the brain-heart infusion agar was substituted with brain-heart infusion broth), were compared. Two-hundred and eighteen clinical samples were used to establish the recovery rate, the number of colony-forming units (CFUs) of A. actinomycetemcomitans as well as the total number of CFUs on the three different types of medium. In addition, the numbers of gram-negative aerobic rods and yeasts were determined. RESULTS: Both types of Dentaid-1 medium showed a higher recovery of A. actinomycetemcomitans compared with TSBV. However, these differences did not reach statistical significance. The total number of CFUs of A. actinomycetemcomitans recovered was significantly higher on Dentaid-1 compared with TSBV (p = 0.029). The mean number of gram-negative aerobic rods recovered was statistically higher on both types of Dentaid-1 medium in comparison with TSBV. Low numbers of yeasts were recovered occasionally on all test plates. CONCLUSION: Dentaid-1 is a low-cost effective alternative to TSBV for the isolation and growth of A. actinomycetemcomitans from clinical samples, such as dental plaque, which contain a complex microflora.