Biomedical subjects

J de Graaff

Publications and source records attributed to J de Graaff.

At least 37 records · Page 2Linked to original sources

PubMed1996

Discrepancy between culture and DNA probe analysis for the detection of periodontal bacteria.

The purpose of this study was to compare a commercially available DNA probe technique with conventional cultural techniques for the detection of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Prevotella intermedia in subgingival plaque samples. Samples from 20 patients with moderate to severe periodontitis were evaluated at baseline and during a 15 months period of periodontal treatment. Paperpoints from 4 periodontal pockets per patient were forwarded to Omnigene for DNA probe analysis, and simultaneously inserted paperpoints from the same pockets were analyzed by standard culture techniques. In addition, mixed bacterial samples were constructed harbouring known proportions of 25 strains of A. actinomycetemcomitans, P. gingivalis and P. intermedia each. A relatively low concordance was found between both methods. At baseline a higher detection frequency was found for A. actinomycetemcomitans and P. gingivalis for the DNA probe technique; for P. intermedia the detection frequency by culture was higher. For A. actinomycetemcomitans, 21% of the culture positive samples was positive with the DNA probe. Testing the constructed bacterial samples with the DNA probe method resulted in about 16% false positive results for the 3 species tested. Furthermore, 40% of P. gingivalis strains were not detected by the DNA probe. The present data suggest that at least part of the discrepancies found between the DNA probe technique used and cultural methods are caused by false positive and false negative DNA probe results. Therefore, the value of this DNA probe method for the detection of periodontal pathogens is questionable.

Adult↗

PubMed1996

Evaluation of the long-term efficacy and safety of locally-applied minocycline in adult periodontitis patients.

The objectives of the present study were to establish in a long-term investigation the safety as well as the clinical and microbiological efficacy of scaling and rootplaning combined with local application of 2% minocycline hydrochloride-gel versus placebo-gel in patients with moderate to severe chronic adult periodontitis. This was an 18 months, randomized, double-blind, parallel, comparative study, in which 20 healthy patients with moderate to severe chronic periodontitis participated. At baseline, all patients received professional oral hygiene-instruction and supra- and subgingival scaling and root planing. The minocycline-gel was applied subgingivally baseline, 2 weeks, 1, 3, 6, 9 and 12 months. Microbiological evaluation was carried out using DMDx to identify the following bacteria: Porphyromonas gingivalis, Prevotella intermedia, Actinobacillus actinomycetemcomitans, Campylobacter rectus, Fusobacterium nucleatum and Treponema denticola. In addition standard microbiological techniques were used for the detection of P. gingivalis, P. intermedia, P. micros, A. actinomycetemcomitans, C. rectus, F. nucleatum, C. albicans and Enterobacteriaceae. Results showed a statistically significant improvement for all clinical parameters irrespective of the treatment modality. No differences were observed between test and control with regard to probing depth and attachment level. The DMDx data showed a significant reduction in both the numbers and the prevalence over the 15 months period, but no significant difference between groups. Culture data showed that at baseline two-third were positive for P. gingivalis and P. intermedia. Analysis over the 18 month period showed no significant difference between the two treatment modalities. C. albicans and Enterobacteriaceae were detected only in small proportions at each time interval in a limited number of patients. No adverse reactions were observed during the trial period. The present patient group responded favourably to scaling and rootplaning, but did not benefit from an effect of local of minocycline. Subgingival debridement in combination with oral hygiene instruction by itself has been shown to be effective. It remains to be studied whether local application of minocycline can be effective as an adjunct to mechanical therapy in sites that respond poorly to conventional treatment.

Administration, Topical↗

PubMed1996

Typing of Helicobacter pylori with monoclonal antibodies against Lewis antigens in lipopolysaccharide.

Recently, it has been shown that the lipopolysaccharide (LPS) O antigen of Helicobacter pylori contains Lewis x (Lex), Lewis y (Ley), or both Lex and Ley antigens. We applied a serotyping method for H. pylori by an enzyme-linked immunosorbent assay with monoclonal antibodies (MAbs) specific for these antigens and the related fucosylated H type 1 (H1) antigen. The selected MAbs recognized the Lex and/or Ley structures in the LPS of H. pylori. The agreement between the results of biochemical compositional analysis and the serological data validated our serotyping system. A total of 152 strains from different geographic origins (The Netherlands, Canada, Poland, Italy, and People's Republic of China) were examined for typeability based on the presence of Lewis antigens. One hundred twenty-nine (84.9%) strains were typeable, and 12 different serotyping patterns were observed; 80.9% of the strains contained Lex and/or Le(y) antigens, and 18.4% reacted with the MAb against the related H1 antigen either alone or in combination with the Lex and/or Ley antigen. Our results show that the Lex and Ley antigens are frequently encountered in the LPS of H. pylori strains from various geographic origins. This typing method is an easy-to-perform technique, which can be used for strain differentiation in epidemiological studies of H. pylori infections.

Antibodies, Bacterial↗

PubMed1996

Practical diagnosis of Helicobacter pylori infection.

When treatment of Helicobacter pylori infection is considered, a reliable diagnosis is essential. Different biopsy-based invasive diagnostic tests are available for diagnosis in the individual patient. Non-invasive tests are of value in epidemiological studies, and have a role in the follow-up of treatment. In this review the sensitivity, specificity and appropriateness of all available tests for the diagnosis of infection in the individual patient are discussed.

Breath Tests↗

PubMed1995

Genomic DNA fingerprinting by restriction fragment end labeling.

A typing method for bacteria was developed and applied to several species, including Escherichia coli and Actinobacillus actinomycetemcomitans. Total genomic DNA was digested with a restriction endonuclease, and fragments were enabled with [alpha-32P]dATP by using the Klenow fragment of DNA polymerase and separated by electrophoresis in 6% polyacrylamide/8 M urea (sequencing gel). Depending on the restriction endonuclease and the bacterium, the method produced approximately 30-50 well-separated fragments in the size range of 100-400 nucleotides. For A. actinomycetemcomitans, all strains had bands in common. Nevertheless, many polymorphisms could be observed, and the 31 strains tested could be classified into 29 distinct types. Furthermore, serotype-specific fragments could be assigned for the three serotypes investigated. The method described is very sensitive, allowing more distinct types to be distinguished than other commonly used typing methods. When the method was applied to 10 other clinically relevant bacterial species, both species-specific bands and strain-specific bands were found. Isolates from different locations of one patient showed indistinguishable patterns. Computer-assisted analysis of the DNA fingerprints allowed the determination of similarity coefficients. It is concluded that genomic fingerprinting by restriction fragment end labeling (RFEL) is a powerful and generally applicable technique to type bacterial species.

DNA Fingerprinting↗

PubMed1995

Presence of Helicobacter pylori in the oral cavity, oesophagus, stomach and faeces of patients with gastritis.

The presence of Helicobacter pylori in the oral cavity (6 sites), oesophagus, stomach and bowel of 20 dyspeptic patients was investigated. Samples were cultured on three selective media and analyzed by 16S rDNA polymerase chain reaction (PCR) and southern hybridization. Helicobacter pylori DNA was detected by PCR from oral-cavity samples of three (20%) and from faeces samples of only one (7%) of the patients whose stomach biopsies were positive for Helicobacter pylori. When culture was used, the microorganism's rate of recovery from the oral cavity and faeces was 13% and 7%, respectively. One patient had a Helicobacter pylori-like organism in samples collected from the tongue and palate. Both strains were urease, catalase and oxidase positive and grew microaerophilically but were negative on PCR analysis. This demonstrates the possibility of false identification of Helicobacter pylori by use of routine enzyme reactions. Interestingly, specimens collected from the cheeks of three patients were positive for Helicobacter pylori by PCR analysis. This is the first instance of detection of this microorganism in the cheek.

Adult↗

PubMed1995

Metronidazole susceptibility factors in Actinobacillus actinomycetemcomitans.

Actinobacillus actinomycetemcomitans is a facultative anaerobic bacterium that displays moderate susceptibility to metronidazole and this study was undertaken to identify the factors involved. A. actinomycetemcomitans appeared two to four times less susceptible to metronidazole when grown in air supplemented with 5% CO2 than under anaerobic conditions. Ferredoxin-linked pyruvate:oxidoreductase activity was absent but each strain exhibited nitroreductase activity which corresponded directly with uptake of metronidazole and susceptibility to the drug under anaerobic conditions but not in air supplemented with 5% CO2. Nitroreductase activity therefore appears responsible for the susceptibility of A. actinomycetemcomitans to metronidazole.

4-Aminobenzoic Acid↗

PubMed1995

Oral microbiota in subjects with a weak or strong response in experimental gingivitis.

The purpose of the present study was to examine the composition of the oral microbiota in subjects who had previously demonstrated to develop either a weak or strong response to experimental gingivitis. For this study, subjects were selected from a pool of 25 individuals who had participated twice in an experimental gingivitis trial. Out of these 25 panellists, 6 subjects were selected who had developed 2X a weak gingival inflammatory response and 7 subjects who had developed 2X a strong gingival inflammatory response. Approximately 9 months after the 2nd experimental gingivitis trial, we evaluated the clinical condition and the prevalence of a panel of selected oral micro-organisms in these subjects. The subjects were clinically examined for the presence of plaque, bleeding, pocket depth and loss of attachment. For the microbiological evaluation, samples were taken from the mucous membranes, subgingival sites and saliva. Samples were analyzed for the presence of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Peptostreptococcus micros, Actinomyces spp., Fusobacterium nucleatum, Campylobacter rectus, spirochaetes and motile rods. Clinical evaluation showed that most subjects had a relatively healthy periodontal condition. No clinically significant differences could be detected between the weak and strong responding groups. The microbial evaluation showed absence of A. actinomycetemcomitans, P. gingivalis and P. micros in all subjects in either group. Analysis of the microbial data for the weak and strong responding group showed no differences between the groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Actinomyces↗

PubMed1995

Putative periodontal pathogens colonizing oral mucous membranes in denture-wearing subjects with a past history of periodontitis.

The prevalence of selected putative periodontal pathogens colonizing oral mucous membranes was investigated in 26 denture-wearing subjects with a history of periodontitis. The microbiota of the oral cavity of these subjects was examined on average 9.3 years after extraction (range: 1-40 years). Samples were taken from several locations on the oral mucous membranes, from saliva and from plaque attached to the upper and lower denture. Actinobacillus actinomycetemcomitans was not detected in any of the 26 edentulous subjects. Porphyromonas gingivalis was detected in 2 subjects (proportion of cultivable flora: 0.002% and 12.5%). Prevotella intermedia was recovered from 7 subjects with a mean proportion of 0.5% (range < 0.001%-6.3%). Analysis showed that subjects harbouring P. intermedia were wearing full dentures for a longer period of time (mean: 20.2 years) than subjects in which P. intermedia could not be detected (mean: 6.6 years). This result suggests that the absence of P. intermedia shortly after extraction may reflect only a temporary event. Other Prevotella species were found in 22 subjects, constituting < 2% of the total cultivable flora (range 0.01%-10.5%). In conclusion, the low prevalence of P. gingivalis and the absence of A. actinomycetemcomitans, suggest that the oral cavity of edentulous subjects may not be regarded as a habitat for these species. The preferable habitat of the Prevotella spp. seems to be the oral mucous membranes, possibly with the exception of P. intermedia.

Adult↗

PubMed1995

Iron-repressible outer membrane proteins of Helicobacter pylori involved in heme uptake.

Helicobacter pylori is known to be a causative agent of gastritis and peptic ulcer disease in humans. The acquisition of iron from the human host may contribute greatly to the virulence of this organism. To study this, H. pylori was cultured under iron-restrictive conditions to induce synthesis of possible iron-regulated outer membrane proteins. This was achieved by the addition of 20% (vol/vol) heat-inactivated newborn calf serum, which contains iron-binding proteins like transferrin and albumin, and no free iron. The newborn calf serum was able to bind free ionic iron in brucella broth culture medium. Electrophoretic analysis of outer membrane preparations from H. pylori cultured under conditions of iron restriction showed several proteins to be present at elevated levels. These appeared to be iron-repressible outer membrane proteins (IROMPs). In addition, IROMPs with molecular sizes of 77, 50, and 48 kDa were isolated by use of hemin-agarose affinity chromatography. These three heme-binding IROMPs might be involved in the uptake of heme from the host and might therefore be important virulence factors of H. pylori.

Animals↗

PubMed1995

Effect of polyclonal and monoclonal antibodies on surface properties of Streptococcus sobrinus.

In this study, the effect of antibody adsorption on physicochemical properties of Streptococcus sobrinus was studied. Bacteria were preincubated with polyclonal antibodies or with OMVU10, a monoclonal antibody (MAb) reactive with S. sobrinus. The zeta potentials and the hydrophobicity as determined by microbial adhesion to hydrocarbons were measured in potassium phosphate buffer with a pH ranging from 2 to 9. S. sobrinus preincubated with polyclonal antibodies was positively charged at pH 2, 3, and 4 and had an isoelectric point at pH 4.8. Untreated S. sobrinus cells or cells preincubated with MAbs were negatively charged over the whole pH range. X-ray photoelectron spectroscopy showed a decrease in O/C and P/C ratios for bacteria preincubated with polyclonal antibodies. A combination of the pH-dependent zeta potential and the X-ray photoelectron spectroscopy data of the overall chemical composition of the cell surface suggests that polyclonal antibody adsorption occurs through blocking of surface phosphate. The measurement of hydrophobicity by microbial adhesion to hydrocarbons revealed that S. sobrinus preincubated with polyclonal antibodies was hydrophobic (90% of the bacteria bound to hexadecane), whereas the controls were relatively hydrophilic. S. sobrinus preincubated with OMVU10 was found to be more hydrophobic than the controls at pH 5 and 7. Hydrophobicity as measured by water contact angles showed an increase in hydrophobicity when S. sobrinus was preincubated with polyclonal antibodies. The epitopes to which the antibodies are directed were visualized by immunogold labeling and electron microscopy. The results suggested that OMVU10 is reactive with only a few epitopes of the cell surface, whereas polyclonal antibodies were found to be reactive with many epitopes. In conclusion, adsorption of polyclonal antibodies was found to influence the overall physicochemical surface properties of the organism, probably by forming a coating over the whole cell surface. Adsorption of MAbs was more localized, which could explain their lesser influence on these surface properties.

Adsorption↗

PubMed1995

Recovery of Helicobacter pylori from gastric biopsy specimens is not dependent on the transport medium used.

The survival of six clinical isolates of Helicobacter pylori at room temperature was investigated after suspension in five different media: brucella broth with 5% lysed horse blood (BBLH), phosphate-buffered saline (PBS), 20% glucose, Stuart medium, and PBS with 10% Fildes enrichment (PBS-F). Only in BBLH and PBS-F no decrease in mean bacterial numbers was observed during the 24-h study period. No H. pylori isolates could be cultured from Stuart medium after 7 h of incubation. In contrast, the recovery rates in PBS-F or Stuart medium of H. pylori isolates from gastric tissue specimens collected from 19 H. pylori-positive patients were not significantly different even after a delay of culture of up to 24 h. Our data show that the medium composition is not critical for the survival of H. pylori within gastric tissue specimens.

Biopsy↗

PubMed1995

A comparison of the microbial flora in carious dentine of clinically detectable and undetectable occlusal lesions.

It is not known whether the aetiology of occlusal hidden caries lesions (HCL) is identical to that of small visible lesions (SVL). Previous studies of the microflora of HCL suggest that relatively few species can be isolated. The aim of the present study was to compare the bacterial composition of dentine from 10 HCL and 17 SVL in a population of children aged 8-18 years. The following bacteria were identified: Actinomyces spp., mutans streptococci, Streptococcus sanguis, Streptococcus oralis, Streptococcus gordonii. Streptococcus mitis, and Lactobacillus spp. Streptococcus mutans was found more often in the HCL group (p = 0.03), while Streptococcus sobrinus was found more often in the SVL group (p = 0.05). However, proportions of both species were lower in the SVL group as compared with the HCL group (S. mutans p = 0.07; S. sobrinus p = 0.03). Lactobacilli and Actinomyces spp. were found at similar levels in both groups. A more diverse range of streptococcal species and Veillonella was found in the SVL group (p = 0.001). These results show a less complex microflora in HCL as compared with SVL, indicating that the aetiology of hidden caries might be different from that of open caries.

Actinomyces↗

PubMed1995

Treatment of Helicobacter pylori infections.

The available literature on the relationship between several diseases and Helicobacter pylori (H. pylori) is reviewed. Duodenal ulcer, gastric ulcer, complicated peptic ulcer, abdominal symptoms and gastroduodenal mucosal damage during the use of non-steroid anti-inflammatory drugs (NSAIDs), non-ulcer dyspepsia (NUD) and gastric malignancy are discussed. The case for and against eradication is critically discussed. Eradication of H. pylori should be pursued in all patients with peptic ulcer disease, whether they are using NSAIDs or not. Eradication of H. pylori in the treatment of NUD should be considered experimental. Treatment aimed at the eradication of H. pylori should be considered in all patients with low-grade malignant mucosa-associated lymphoid tissue (MALT) lymphoma and in all patients with Ménétrier's disease. Finally, this treatment should be considered in a subset of H. pylori-infected patients who possibly are at an increased risk of gastric cancer: patients with a strong family history of gastric carcinoma and patients in need of long-term treatment with a proton-pump inhibitor. In view of the importance of patient compliance, the risk of side-effects and the possibility of inducing metronidazole resistance when treatment with a metronidazole-containing regimen is used, treatment aimed at the eradication of H. pylori should be carefully implemented and monitored.

Anti-Bacterial Agents↗

PubMed1994

Amoxycillin causes an enhanced uptake of metronidazole in Actinobacillus actinomycetemcomitans: a mechanism of synergy.

We investigated the influence of sub-inhibitory concentrations amoxycillin on metronidazole uptake of the metronidazole-susceptible facultative microorganism Actinobacillus actinomycetemcomitans. The rate of metronidazole uptake by bacterial cells simultaneously incubated with amoxycillin was higher than uptake in cells incubated with metronidazole alone. This phenomenon may explain the recently reported in-vitro synergic interaction between metronidazole and amoxycillin against A. actinomycetemcomitans.

Aggregatibacter actinomycetemcomitans↗

PubMed1994

Prevalence of periodontitis and suspected periodontal pathogens in families of adult periodontitis patients.

The aim of the present study was to investigate the prevalence of periodontopathic microorganisms and periodontal destruction in the spouses and children of adult periodontitis patients. For this study, 24 families were selected on the basis of one parent with severe periodontal breakdown and the presence of Actinobacillus actinomycetemcomitans and/or Porphyromonas gingivalis and/or more than 30% Prevotella intermedia subgingivally. The clinical examination of both parents and children included pocket depth and clinical attachment loss (CAL) measurements. Samples for bacterial examination were obtained from the mucous membranes, the saliva and pockets. Pocket selection was based on the most advanced periodontitis situation found in a subject. The samples were cultured for the detection of A. actinomycetemcomitans, P. gingivalis and P. intermedia. By phase-contrast microscopy, the % of spirochetes and motile microorganisms was assessed. The number of children within each family varied between 1 and 3. In total 49 children were investigated with a range in age of 3 months to 15 years. Results showed that under the age of 5 years, none of the children had CAL, whereas in the age group of 5-15 years, 26.5% had 1-5 sites in the primary and/or permanent dentition with 1-3 mm CAL. 3 of the spouses had no interproximal CAL. 16 of the 24 spouses had a light to moderate form of periodontitis, with at least one site with 1 to 4 mm CAL and 5 spouses had severe periodontal breakdown with sites showing at least 8 mm CAL. Spirochetes, motile microorganisms and P. intermedia were frequently present in all family members. 18 out of the 24 probands were positive for P. gingivalis. This organism was found once only in a 5-year-old boy and in 11 of the spouses. A. actinomycetemcomitans was detected in 13 probands; 5 children and 5 of the spouses were also positive for this bacterium. If a child harboured one of the periodontopathogens, at least 1 of the parents was also positive for that bacterium. This phenomenon may be due to transmission of microorganisms between family members. Comparison of the clinical data reported in the present study with similar clinical parameters from epidemiological studies of the Dutch population suggest that the spouses and children of adult periodontitis patients might be at relatively high risk of developing periodontal breakdown.

Adolescent↗

PubMed1994

Microbiological and clinical effects of metronidazole and amoxicillin in Actinobacillus actinomycetemcomitans-associated periodontitis. A 2-year evaluation.

In this study, we evaluated the microbiological and clinical effects of mechanical debridement in combination with metronidazole and amoxicillin therapy in 48 patients with Actinobacillus actinomycetemcomitans-associated periodontitis, 3 months and at least 24 months after active treatment. The results of this study showed that 47 out of 48 patients were still negative for A. actinomycetemcomitans subgingivally, at the mucous membranes, the tonsillar area and in the saliva, 2 years after therapy. The clinical results showed that a reduction of probing pocket depth, probing attachment level, bleeding index and plaque index was not only seen in the time between baseline and 3 months after therapy, but further clinical improvement was observed between 3 and 24 months after active treatment. We conclude that combined mechanical debridement and metronidazole plus amoxicillin therapy is very effective in suppressing A. actinomycetemcomitans below cultivable levels over a long period of time, suggesting elimination of this organism, and that recolonization of A. actinomycetemcomitans seems to be a rare event. The elimination of A. actinomycetemcomitans is paralleled by a further improvement of the periodontal status of the patients, even up to 24 months after active treatment.

Aggregatibacter actinomycetemcomitans↗

PubMed1994

Relationship between salivary blood group antigens, microbial flora and periodontal condition in young adults.

To investigate a possible rôle for salivary blood-group antigens in the relative frequencies of selected periodontal pathogens and commensal oral micro-organisms, we studied the clinical and microbiological condition in young adults with or without blood group reactive substances in saliva (secretors or non-secretors respectively). Clinical measurements were recorded at the Ramfjord teeth in 81 1st-year dental students. In addition, presence of interproximal loss of attachment (LA) was assessed at sites with a pocket depth of > or = 4 mm. Microbiological samples were taken from one of the Ramfjord teeth (site without interproximal LA), from interproximal sites of > or = 4 mm in conjunction with LA, and from the saliva. The samples were analyzed for the presence of Actinomyces naeslundii, Actinomyces viscosus, Campylobacter rectus, Fusobacterium nucleatum, Peptostreptococcus micros, Prevotella intermedia, Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans. Clinically, no statistically significant differences were found in the periodontal status between secretors (78% of our population) and non-secretors. Furthermore, the occurrence of the monitored micro-organisms was not correlated to the secretor status. It is concluded that bacterial colonization with the micro-organisms tested in this study, apparently occurred independent of secretor status. Among the periodontal pathogens, only P. intermedia was more frequently recovered from the saliva of subjects with interproximal LA (49%) than in those without (33%; p = 0.03). This finding was irrespective of the secretor status. Therefore, P. intermedia may be an important micro-organism in relation to the onset of periodontal destruction in young adults.

ABO Blood-Group System↗