Tove Larsen

Resistance of Streptococcus sanguis biofilms to antimicrobial agents

Bacteria living in biofilms as dental plaque on tooth surfaces are generally more resistant to antimicrobial agents than bacteria in batch culture normally used for in vitro susceptibility testing. In order to compare the resistance of free‐living and surface‐grown oral bacteria, the MIC of Streptococcus sanguis 804 and ATCC 10556 to amoxicillin, doxycycline and chlorhexidine was determined by a broth dilution method. Subsequently, S. sanguis biofilms established in an in vitro flow model were perfused with the antimicrobial agents for 48 h at concentrations equal to and up to 500 times the MIC, and biofilm cell number was determined during this period. The antibiotics at the MIC did not affect the cell number of S. sanguis biofilms compared to the starting point, and only after 48 h at 500 times the MIC were the biofilm bacteria eliminated. At intermediate concentrations biofilm cell number gradually decreased. Chlorhexidine also gradually reduced biofilm cell number, but was inhibitory at concentrations closer to the MIC than was the case for the antibiotics. Thus S. sanguis in biofilms survived up to 500 times the MIC found in batch culture for up to 48 h.

Development of a flow method for susceptibility testing of oral biofilms in vitro

Bacteria in biofilms are known to be more resistant than bacteria in batch cultures to antimicrobial agents. The purpose of the present study was to develop a flow method for formation of oral biofilms permitting susceptibility testing of plaque bacteria. A brain heart infusion (BHI) Streptococcus sanguis 804 culture was pumped through a modified Robbins Device (MRD) with 25 exchangeable silicone disks at 40 ml/h. After 24–48 h disks were removed and biofilm cells dispersed by vortex mixing and low‐output ultrasonication. Colony forming units (cfu)/cm2 were determined after aerobic incubation on blood agar plates. Optimal biofilm formation was found after growth for 48 h at 37°C in BHI + 1% sucrose, using saliva‐coated silicone disks in inverted MRDs, yielding on average 4.4 × 105 cfu/cm2. Similar results were obtained for S. sanguis ATCC 10556 and five clinical isolates. Testing the susceptibility of S. sanguis to chlorhexidine gluconate showed increased resistance of biofilms compared to batch culture. Thus an appropriate biofilm model for susceptibility testing of oral microorganisms has been established.

Occurrence of doxycycline resistant bacteria in the oral cavity after local administration of doxycycline in patients with periodontal disease

Topical antimicrobial treatment is appearing as a means of therapy in patients with advanced periodontal disease. The purpose of the present study was to examine the occurrence of doxycycline resistant bacteria in subgingival plaque and oral cavity after local administration of doxycycline. Five patients with advanced marginal periodontitis were scaled, and one approximal pocket in each patient was additionally treated with locally delivered doxycycline. Microbiological samples were obtained from the test site, a contralateral control site and tongue and tonsils before treatment and 3, 13, 26 and 52 weeks after treatment. The occurrence and morphological distribution of doxycycline resistant bacteria was determined after anaerobic cultivation on enriched tryptic soy agar with and without doxycycline incorporated. At the test site and on tongue and tonsils the percentage of doxycycline resistant bacteria increased from less than 1% before treatment to 22% and 35%, respectively, immediately after treatment, but decreased again at week 13. At the control site no increase was observed. Gram-positive cocci constituted the majority of doxycycline resistant bacteria at all 3 sampling sites (73-94%). The morphological distribution of resistant bacteria was not affected by the doxycycline therapy. Thus, local doxycycline therapy resulted only in a transient increase in resistance in the oral microflora.

Effect of steam sterilization inside the turbine chambers of dental turbines

OBJECTIVE It has been demonstrated that contamination of the insides of high-speed dental turbines occurs and that bacteria as well as viruses may remain infectious when expelled from such turbines during subsequent use. Consequently, it has been widely recommended that a high-speed turbine be sterilized after each patient. The purpose of this study was to evaluate the effect of steam autoclaving on a high-speed dental turbine with a contaminated turbine chamber. STUDY DESIGN Streptococcus salivarius and endospores of Bacillus stearothermophilus were used as test organisms to determine the effectiveness of 4 different small non-vacuum autoclaves and one vacuum autoclave. RESULTS The study demonstrated different efficiencies among the small non-vacuum autoclaves, the best showing close to a 6 log reduction of the test organisms inside the turbine chamber. When cleaning and lubrication of the high-speed dental turbine was carried out before autoclaving, this level of reduction was observed for all the examined non-vacuum autoclaves. CONCLUSIONS It is concluded that cleaning before sterilization is essential for safe use of high-speed dental turbines and that small non-vacuum autoclaves should be carefully evaluated before being used for the reprocessing of hollow instruments such as high-speed turbines.

Dental biofilm infections – an update

Teeth are colonized by oral bacteria from saliva containing more than 700 different bacterial species. If removed regularly, the dental biofilm mainly comprises oral streptococci and is regarded as resident microflora. But if left undisturbed, a complex biofilm containing up to 100 bacterial species at a site will build up and may eventually cause development of disease. Depending on local ecological factors, the composition of the dental biofilm may vary considerably. With access to excess carbohydrates, the dental biofilm will be dominated by mainly gram‐positive carbohydrate‐fermenting bacteria causing demineralization of teeth, dental caries, which may further lead to inflammation and necrosis in the pulp and periapical region, i.e., pulpitis and periapical periodontitis. In supra‐ and subgingival biofilms, predominantly gram‐negative, anaerobic proteolytic bacteria will colonize and cause gingival inflammation and breakdown of supporting periodontal fibers and bone and ultimately tooth loss, i.e., gingivitis, chronic or aggressive periodontitis, and around dental implants, peri‐implantitis. Furthermore, bacteria from the dental biofilm may spread to other parts of the body by bacteremia and cause systemic disease. Basically, prevention and treatment of dental biofilm infections are achieved by regular personal and professional removal of the dental biofilm.

Evaluation of a new device for sterilizing dental high-speed handpieces

Dental high-speed turbines and handpieces can take up and expel microorganisms during operation and thus need regular sterilization. This study established a method for validating devices used to sterilize high-speed turbines and handpieces. The air and water channels and turbine chambers were contaminated with suspensions of Streptococcus salivarius or endospores of Bacillus stearothermophilus. The effect of flushing and/or autoclaving performed by a new device combining both procedures was evaluated by counting the number of viable bacteria recovered from these devices. Further, the effect on clinically used handpieces was evaluated. In an initial experiment, the device partially reduced S. salivarius, and the endospores survived. In a second experiment, a 5 to 6 log reduction of S. salivarius in air and water channels was obtained. No growth was observed in clinically used high-speed handpieces, and both S. salivarius and endospores were eliminated from the turbine chambers. Thus, the method of validation proved capable of discriminating between different levels of bacterial reduction.

Changing the curriculum and the role of the teacher and the students in the classroom – an analysis of the process of reforming a course in oral microbiology

In 2005, a new curriculum was introduced at the School of Dentistry at the University of Copenhagen. Amongst many changes, the pedagogical concept behind theoretical teaching was changed. The main emphasis was previously on lectures and teacher controlled teaching, but the emphasis has now shifted to a more dialogue-based teaching style for smaller groups where the students became much more active. Thus, the learning principle was now given the pride of place. The present article focuses on the pedagogical reorganisation of the course in oral microbiology where, based on interviews with students and teachers, an evaluation is made regarding how the changes were implemented and developed. Despite the fact that the students themselves wanted to abandon the lecture-based teaching style, there was great dissatisfaction with the new dialogue-based teaching style as the students did not think that it was possible for them to take responsibility for their learning. They would much rather listen to teachers who know the material. Initially, the teachers were very surprised by the negative attitude of the students and had to change the teaching style to more traditional teaching in groups whilst still maintaining the dialogue principle to a certain degree. In this article, the need to enter into a teaching contract between students and teachers prior to introducing new pedagogical principles is discussed.