Bacterial leakage along the implant-abutment interface: culture and DNA Checkerboard hybridization analyses

Objective Bacterial species have been found harboring the internal surface of dental implants as consequence of their failed connections. The aim of the present study was to compare the detection frequency of bacterial leakage from human saliva through the implantabutment interface, under non-loading conditions, using either DNA Checkerboard or culture method. Materials and methods Thirty dental implants with hexagonal platforms were connected to pre-machined abutments according to the manufacturers specifications. The assemblies were individually incubated in human saliva under anaerobic conditions for 7 similar to days at 37 degrees C. Afterward, contents from the inner parts of the implants were collected and evaluated with either DNA Checkerboard (s similar to=similar to 15) or culture (n similar to=similar to 15). Subsequently, identification and quantitation of bacterial species from saliva and implants were carried out for the group evaluated with the DNA Checkerboard method. Results Both DNA Checkerboard and culture showed positive signals of bacterial leakage in 6 of the 15 evaluated samples. Capnocytophaga gingivalis and Streptococcus mutans were the most frequently detected species harboring the internal surface of the implants followed by Veillonella parvula. Conclusion Occurrence of bacterial leakage along the implantabutment interface is comparably detected with both DNA Checkerboard hybridization and conventional culture methods.

Leakage of Saliva Through the Implant-Abutment Interface: In Vitro Evaluation of Three Different Implant Connections Under Unloaded and Loaded Conditions

Purpose: Bacterial leakage along the implant-abutment interface, with consequent species harboring the inner parts of two-part dental implant systems, has been reported in the literature. The aim of this in vitro study was to evaluate bacterial leakage from human saliva to the internal part of the implants along the implant-abutment interface under loaded and unloaded conditions using DNA Checkerboard. Materials and Methods: Sixty denial implants-20 each of external-hexagon, internal-hexagon, and Morse cone-connection designs-and their conical abutments were used in this study. Each group was subdivided into two groups of 10 loaded and 10 unloaded implants. The assemblies were immersed in human saliva and either (1) loaded with 500,000 cycles at 120 N (experimental group) or (2) incubated in static conditions for 7 days at 35 degrees C (unloaded control group). Results: Microorganisms were found in the internal surfaces of all types of connections. The Morse cone connection presented the lowest count of microorganisms in both the unloaded and loaded groups. Loaded implants presented with higher counts of microorganisms than unloaded implants for external- and internal-hex connections. Conclusion: Bacterial species from human saliva may penetrate along the implant-abutment interface under both unloaded and loaded conditions for all connections evaluated. Morse cone-connection implants showed the lowest counts of microorganisms for both conditions. External- and internal-hex implants showed a higher incidence of bacteria and higher bacterial counts after simulated loading. INT J ORAL MAXILLOFAC IMPLANTS 2012;27:551-560.

In vivo evaluation of a metronidazole-containing gel for the adjuvant treatment of chronic periodontitis: preliminary results

The present study developed an experimental metronidazole-based gel and evaluated its efficacy for the adjuvant treatment of chronic periodontitis. Sixteen patients were randomly allocated into two groups of eight subjects according to the following proposed treatments: (1) scaling and root planing (active control) or (2) scaling and root planing and direct periodontal intrapocket application of 15% metronidazole-based gel in two sites (a parts per thousand yen5 mm in depth) (experimental group). Potential changes in the subgingival microbiota were assessed using a DNA Checkerboard method at three proposed times: baseline and following 7 or 30 days of drug administration. High-performance liquid chromatography (HPLC) monitored metronidazole concentrations in the crevicular fluid during treatment. The metronidazole experimental group presented lower bacterial counts than the control group at the three evaluated times (p < 0.01 for baseline, p < 0.001 for 7 or 30 days) when the target species were analyzed as a pool of bacteria. Samples revealed significantly lower counts 7 days after drug administration compared with baseline or after 30 days (p < 0.05). HPLC analysis detected gel 1 h after application. The metronidazole-based gel significantly decreased the total bacterial count at the three evaluated times. Periodontopathogenic species were not different after gel administration.

Molecular detection of in-vivo microbial contamination of metallic orthodontic brackets by checkerboard DNA-DNA hybridization

Introduction: Knowing the microbiota that colonizes orthodontic appliances is important for planning strategies and implementing specific preventive measures during treatment. The purpose of this clinical trial was to evaluate in vivo the contamination of metallic orthodontic brackets with 40 DNA probes for different bacterial species by using the checkerboard DNA-DNA hybridization (CDDH) technique. Methods: Eighteen patients, 11 to 29 years of age having fixed orthodontic treatment, were enrolled in the study. Each subject had 2 new metallic brackets bonded to different premolars in a randomized manner. After 30 days, the brackets were removed and processed for analysis by CDDH. Data on bacterial contamination were analyzed descriptively and with the Kruskal-Wallis and Dunn post tests (alpha = 0.05). Forty microbial species (cariogenic microorganisms, bacteria of the purple, yellow, green, orange complexes, "red complex + Treponema socranskii," and the cluster of Actinomyces) were assessed. Results: Most bacterial species were present in all subjects, except for Streptococcus constellatus, Campylobacter rectus, Tannerella forsythia, T socranskii, and Lactobacillus acidophillus (94.4%), Propionibacterium acnes I and Eubacterium nodatum (88.9%), and Treponema denticola (77.8%). Among the cariogenic microorganisms, Streptococcus mutans and Streptococcus sobrinus were found in larger numbers than L acidophillus and Lactobacillus casei (P < 0.001). The periodontal pathogens of the orange complex were detected in larger numbers than those of the "red complex + T socranskii" (P < 0.0001). Among the bacteria not associated with specific pathologies, Veillonella parvula (purple complex) was the most frequently detected strain (P < 0.0001). The numbers of yellow and green complex bacteria and the cluster of Actinomyces were similar (P > 0.05). Conclusions: Metallic brackets in use for 1 month were multi-colonized by several bacterial species, including cariogenic microorganisms and periodontal pathogens, reinforcing the need for meticulous oral hygiene and additional preventive measures to maintain oral health in orthodontic patients. (Am J Orthod Dentofacial Orthop 2012;141:24-9)

Molecular detection of Aggregatibacter actinomycetemcomitans on metallic brackets by the checkerboard DNA-DNA hybridization technique

Introduction: The purpose of this randomized clinical study was to evaluate the presence of the periodontal pathogen Aggregatibacter actinomycetemcomitans on metallic brackets and the effectiveness of a 0.12% chlorhexidine digluconate mouthwash in inhibiting this microorganism. Methods: The study involved 35 patients of both sexes having orthodontic treatment with fixed appliances between the ages of 14 and 22 years, randomized into 2 groups: experimental (n = 17) and control (n = 18). Two new metallic brackets were placed on the patients' premolars, and the subjects rinsed with a solution of 0.12% chlorhexidine digluconate or a placebo solution twice a week for 30 days. After that, the brackets were removed and underwent microbiologic analysis with the checkerboard DNA-DNA hybridization technique. Data were analyzed by using the Student t, Fisher exact, and Mann-Whitney tests at the significance level of 5%. Results: The results showed that A actinomycetemcomitans was present in all brackets from the subjects in the control group vs 83% of the subjects who rinsed with chlorhexidine digluconate (P<0.0001). There were also significantly lower levels of this species in the chlorhexidine digluconate group compared with the control group (P = 0.0003). Conclusions: We concluded that 0.12% chlorhexidine digluconate rinsing, twice a week for 30 days during orthodontic treatment, is effective in reducing the presence and levels of A actinomycetemcomitans on metallic brackets. (Am J Orthod Dentofacial Orthop 2012;142:481-6)

Effect of sample storage time on detection of hybridization signals in Checkerboard DNA-DNA hybridization

Long-term sample storage can affect the intensity of the hybridization signals provided by molecular diagnostic methods that use chemiluminescent detection. The aim of this study was to evaluate the effect of different storage times on the hybridization signals of 13 bacterial species detected by the Checkerboard DNA-DNA hybridization method using whole-genomic DNA probes. Ninety-six subgingival biofilm samples were collected from 36 healthy subjects, and the intensity of hybridization signals was evaluated at 4 different time periods: (1) immediately after collecting (n = 24) and (2) after storage at -20 degrees C for 6 months (n = 24), (3) for 12 months (n = 24), and (4) for 24 months (n = 24). The intensity of hybridization signals obtained from groups 1 and 2 were significantly higher than in the other groups (p < 0.001). No differences were found between groups 1 and 2 (p > 0.05). The Checkerboard DNA-DNA hybridization method was suitable to detect hybridization signals from all groups evaluated, and the intensity of signals decreased significantly after long periods of sample storage.

Use of the checkerboard DNA-DNA hybridization technique for bacteria detection in Aedes aegypti (Diptera:Culicidae) (L.)

Abstract Background Bacteria associated with insects can have a substantial impact on the biology and life cycle of their host. The checkerboard DNA-DNA hybridization technique is a semi-quantitative technique that has been previously employed in odontology to detect and quantify a variety of bacterial species in dental samples. Here we tested the applicability of the checkerboard DNA-DNA hybridization technique to detect the presence of Aedes aegypti-associated bacterial species in larvae, pupae and adults of A. aegypti. Findings Using the checkerboard DNA-DNA hybridization technique we could detect and estimate the number of four bacterial species in total DNA samples extracted from A. aegypti single whole individuals and midguts. A. aegypti associated bacterial species were also detected in the midgut of four other insect species, Lutzomyia longipalpis, Drosophila melanogaster, Bradysia hygida and Apis mellifera. Conclusions Our results demonstrate that the checkerboard DNA-DNA hybridization technique can be employed to study the microbiota composition of mosquitoes. The method has the sensitivity to detect bacteria in single individuals, as well as in a single organ, and therefore can be employed to evaluate the differences in bacterial counts amongst individuals in a given mosquito population. We suggest that the checkerboard DNA-DNA hybridization technique is a straightforward technique that can be widely used for the characterization of the microbiota in mosquito populations.