Victoria S. Lucas

Statistical analyses of complex denaturing gradient gel electrophoresis profiles

ABSTRACT Studies using molecular techniques have demonstrated that a culture-based approach can severely underestimate the bacterial diversity in most environments. One of the molecular techniques that has been applied in microbial ecology is denaturing gradient gel electrophoresis (DGGE). The purpose of this study was to investigate differences in the microbiota of plaque, using a number of analysis techniques, from children without gingivitis (n = 30) and from those with gingivitis (n = 30). Extracted DNA from gingival margin plaque was subjected to PCR targeting the 16S rRNA gene using universal primers. DGGE profiles were analyzed in three ways. (i) Bacterial diversity was compared between cohorts by using the Shannon-Wiener index (also known as the Shannon-Weaver index). (ii) A hierarchical cluster analysis of the banding patterns was calculated and expressed as a dendrogram. (iii) Individual DGGE bands and their intensities for both cohorts were compared using a logistic regression analysis. The Shannon-Wiener indices demonstrated a greater bacterial diversity associated with no-gingivitis plaque (P = 0.009). Dendrograms demonstrated that seven clades associated with gingivitis and five clades associated with no gingivitis. The logistic regression demonstrated that one band was significantly associated with no gingivitis (P = 0.001), while two bands were significantly associated with gingivitis (P = 0.005 and P = 0.042). In conclusion, this study demonstrates that the development of gingivitis might be accompanied by a decrease in bacterial diversity. Furthermore, we have demonstrated that logistic regression is a good statistical method for analyzing and characterizing DGGE profiles.

Duration, prevalence and intensity of bacteraemia after dental extractions in children

Objective: To investigate the duration, prevalence and intensity of bacteraemia after dental extractions in children by comparing within-patient bacteraemia before and after dental extraction. Methods: Children were randomly allocated to one of 10 postprocedure time groups from 10 s to 60 min. The differences between intensity and prevalence of the bacteraemia at each time after extractions were used to estimate the duration of the bacteraemia. After attainment of general anaesthesia, pre-extraction and postextraction blood samples were processed by broth culture and lysis filtration to isolate and quantify bacteria present in the patients’ blood. Results: 500 subjects between 3 and 16 years old were recruited. The estimated duration of bacteraemia was about 11 min. Conclusions: The duration of bacteraemia after dental extractions is less than previously thought. This has implications for the interpretation of odontogenic bacteraemia studies.

Prevalence of Periodontal Pathogens in Dental Plaque of Children

ABSTRACT Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, and Tannerella forsythensis have been implicated as the main etiological agents of periodontal disease. The purpose of this work was to estimate the prevalence of these organisms in plaque from children without gingivitis (group 1; n = 65) and from those with gingivitis (group 2; n = 53). Extracted DNA from plaque was subjected to two rounds of PCR targeting the 16S rRNA gene using both universal primers and species-specific primers. The results were as follows: group 1, P. gingivalis, 49%; A. actinomycetemcomitans, 55%; and T. forsythensis, 65%; group 2, P. gingivalis, 47%; A. actinomycetemcomitans, 59%; and T. forsythensis, 45%. T. forsythensis was detected more frequently in children with no gingivitis than in those with gingivitis (P = 0.03). There was no significant difference between the two groups with respect to the presence of P. gingivalis or A. actinomycetemcomitans in either group (P > 0.05). Logistic regression analysis revealed that the odds of a patient having gingivitis were 2.3 times greater in the absence of T. forsythensis. In conclusion, the results of this study have shown that the three pathogens can be detected in the dental plaque of healthy children and of those with gingivitis and that T. forsythensis is associated with dental plaque at sites with no gingivitis.

Oro-dental health in children with chronic renal failure and after renal transplantation: a clinical review

As a consequence of chronic renal failure (CRF) and its treatment, a number of oro-dental changes occur that persist after the end-stage is reached. An early effect is enamel hypoplasia due to a defect of enamel development and mineralisation. This is usually reparable to a high aesthetic standard using dental composite filling material. Children with CRF have significantly less dental caries than healthy children due to the inhibitory effect of increased salivary urea levels. Grafted patients frequently develop gingival enlargement as a result of immunosuppression with cyclosporine A, which is further exacerbated by the additional use of antihypertensive calcium-blocking agents. Surgical reduction of gingival hyperplasia is effective and is required in approximately one third of adolescents. A very high standard of home care should be encouraged for all children with CRF in the form of thorough mechanical tooth cleaning and plaque inhibition through the use of an antibacterial mouthwash. In transplanted children presenting an increased risk of infection, antibiotic prophylaxis may be indicated for dental treatment procedures. The drug dosage should be adapted to the reduced renal function. Pediatric nephrologists should be aware that dental surgeons can make a considerable contribution to the general health and well-being of children with CRF. Thus, only oro-dental problems that are mainly encountered and treated by dental surgeons are reviewed.

Comparison of Lysis Filtration and an Automated Blood Culture System (BACTEC) for Detection, Quantification, and Identification of Odontogenic Bacteremia in Children

ABSTRACT Lysis filtration (LyF) was compared with BACTEC PAEDS PLUS in estimating the prevalence of, and sensitivity for, detection of odontogenic bacteremia. Both real bacteremia and simulated bacteremia (seeded blood or saline samples) were assessed to determine the validity of LyF in estimating bacteremia. The simulated bacteremia was also used to assess the reliability of LyF to estimate intensity of bacteremia in CFU per milliliter of blood. Reference organisms were used to assess the abilities of LyF and BACTEC to isolate known oral streptococci. There was no difference in the number of CFU per milliliter of seeded saline, seeded blood, and drop cultures of the organisms plated directly onto agar. Blood cell volume had a negligible effect on the yield of organisms for simulated bacteremia. When LyF and BACTEC were compared, the time to detection of bacteremia was always significantly shorter for BACTEC. For aerobic cultures, these times were 43.7 and 9.6 h, respectively (P < 0.01). For anaerobic cultures, these times were 45.1 and 9.9 h, respectively (P < 0.01). These differences occurred as well for bacteremia following the extraction of a single tooth, with LyF and BACTEC aerobic cultures taking 78 and 30.5 h, respectively (P < 0.0001). For anaerobic cultures, the times were 90.8 and 45 h, respectively (P < 0.0004). A preextraction bacteremia was detected on 2.1% of occasions with BACTEC compared to 31% of occasions with LyF (P < 0.05) The use of LyF was an effective and reliable means of estimating the intensity of pre- and postextraction bacteremia. The values were 3.6 and 5.9 CFU/ml, respectively (P < 0.4729), and the difference was not statistically significant. In summary, BACTEC is quicker than LyF, but less sensitive. LyF provides additional important information in estimating the intensity of bacteremia.

Dental health indices and caries associated microflora in children with unilateral cleft lip and palate.

OBJECTIVE To investigate the dental health and caries related microflora of children with unilateral cleft lip and palate. STUDY GROUP Sixty children with unilateral cleft lip and palate and matched controls. OUTCOME MEASURES The decayed, missing, and filled teeth and surfaces in both the deciduous and permanent dentitions. The presence of developmental defects and plaque and gingivitis scores were also recorded. Plaque was collected from 25 of the children and their matched controls from three different sites, which were (1) the first approximal site distal to the cleft, (2) a contralateral anterior site, and (3) a remote site. It was cultured for Streptococcus mutans and lactobacilli. Plaque was collected from two sites in the matched controls. RESULTS There was no significant difference in the caries, plaque, and gingivitis scores between the children with cleft palate and the controls. A greater number of enamel opacities were recorded in the control group, and there was a higher prevalence of enamel discoloration in the children with cleft lip and palate. There was no significant difference in the proportion of S. mutans or lactobacilli at the cleft site, compared with the unaffected site in the study group, although there was an anterior-posterior gradient in the proportion of S. mutans. There was no significant association between the stagnation area at the cleft site and the bacteria associated with dental caries.

Prevalence, intensity and nature of bacteraemia after toothbrushing

OBJECTIVE To estimate the prevalence, intensity and microbial identity of bacteraemia associated with toothbrushing. METHODS A total of 141 children and adolescents, aged between 3 and 17 years, having dental treatment under general anaesthesia at the Eastman Dental Hospital were recruited. Six millilitre of blood was taken before toothbrushing (baseline) with (1) Oral B 30 toothbrush or (2) Braun or (3) Sonicare electric toothbrush or (4) dental handpiece and rubber cup. A second 6-ml sample was taken 30s after toothbrushing. All blood samples were processed using lysis filtration and bacteria were identified to species level. RESULTS There was a significantly greater prevalence of bacteraemia following the dental handpiece only (p=0.02). There was a significantly greater aerobic and anaerobic intensity of bacteraemia following brushing with both the Sonicare (p=0.03 and p=0.05) and the dental handpiece (p=0.001 and p=0.005). CONCLUSIONS Toothbrushing causes a bacteraemia that is often statistically significantly greater than baseline. Toothbrushing is an important contributory factor in cumulative dental bacteraemia.

Dental age assessment (DAA): Reference data for British caucasians at the 16 year threshold

UNLABELLED The purpose of this study was to determine reference data for dental age assessment (DAA) for the 16 year threshold in British caucasians. PATIENTS, MATERIALS AND METHODS One thousand seven hundred and twenty-two Dental Panoramic Radiographs of individuals aged between 4 and 24 years were re-used to establish reference intervals using the tooth development stages (TDSs) previously described [A. Demirjian, H. Goldstein, J.M. Tanner, A new system of dental age, assessment, Human Biology 45 (1973) 221-227]. All teeth present in the left maxilla and mandible, and the third permanent molars were assessed. In addition, to test the accuracy of DAA, the radiographs of a study sample of 50 children of known chronological age (CA) were assessed in the same manner as the reference population. These were aged between 15 and 17 years and separate from the database. For each subject in the study sample a new method of mathematical manipulation based on meta-analysis was applied to all teeth that were still developing [G.J. Roberts, S. Parekh, A. Petrie, V.S. Lucas, Dental age assessment (DAA): a simple method for children and emerging adults, British Dental Journal 204(4) (2008) 192-193]. The estimated calculated average of all the teeth present on the radiograph of each individual generated by the meta-analysis was assigned to each individual as the dental age. For each test subject this was then compared to the gold standard of chronological age. RESULTS The average difference between dental age and chronological age for individuals in the test sample was 0.27 years (3.24 months) in females and 0.23 years (2.76 months) in males. CONCLUSION Dental age assessment obtained by calculation of tooth development stages using meta-analysis provides estimates of age around the 16 year threshold.

Dental health indices and caries-related microflora in children with severe haemophilia

The purpose of this work was to investigate the prevalence of dental caries, bacterial dental plaque, gingivitis, enamel defects and caries‐ related microflora in children with severe haemophilia. Thirty‐eight children with severe haemophilia (factor VIII and IX < 2 U dL–1) were recruited from Great Ormond Street Hospital for Children and matched for age, gender and ethnicity with healthy control children from the Eastman Dental Institute. Indices were recorded for decayed, missing, and filled teeth and surfaces in both the deciduous dentition (dmfs/dmft) and the permanent dentition (DMFS/DMFT). The plaque and gingivitis scores and developmental enamel defects were also recorded. The caries‐related microflora was sampled and cultured for Streptococcus mutans, and Lactobacilli and Candida species. A significantly greater proportion of children with severe haemophilia were caries‐free compared with the controls (36.7% vs. 13.3%; P=0.04). Both the DMFS and DMFT were significantly greater in the controls (3.6 and 2.8, respectively) compared with the haemophilia group, (0.8 and 0.7; P=0.007 and P=0.04). The plaque score for the permanent dentition only was significantly greater for the control children (24.2) compared with the haemophilia group, (10.2; P=0.04). The mean number of colony forming units of S. mutans was significantly greater in the control group compared with the haemophilia group (P=0.05). We conclude that children with severe haemophilia have a significantly lower prevalence of dental caries compared with matched, healthy controls.

Prevalence, intensity and identity of bacteraemia following conservative dental procedures in children

INTRODUCTION The aim of this study was to investigate the prevalence, intensity and microbial identity of bacteraemia following conservative dental procedures. The procedures were placement of rubber dam, use of the fast drill, use of the slow drill and placement of a matrix band and wedge. METHOD Two hundred and five children and adolescents undergoing general anaesthesia for dental treatment at the Eastman Dental Hospital were recruited. Each subject was randomly allocated to one of the procedure groups. A baseline blood sample was taken before any dental treatment was carried out. A second blood sample was taken 30 s after a single conservative procedure. The blood samples were processed using lysis filtration. All bacterial isolates were identified using comparative 16 S ribosomal RNA gene sequencing. Oral Streptococcus spp. and coagulase-negative Staphylococcus spp. were further identified by comparative sodA gene sequencing. RESULTS The prevalence of bacteraemia was significantly greater following placement of rubber dam (P = 0.01) and placement of matrix band and wedge, compared with baseline. The intensity of bacteraemia was significantly greater following placement of rubber dam (P = 0.001) and placement of matrix band and wedge (P = 0.0001). The most frequently isolated bacteria were Streptococcus spp. (56%), Actinomyces spp. (15%) and coagulase-negative Staphylococcus spp. (15%). CONCLUSION Conservative dental procedures are a significant cause of bacteraemia.