Ca(OH)2 application modes: in vitro alkalinity and clinical effect on bacteria.

AIM: To compare five calcium hydroxide (CH) products, CH-gutta-percha Plus points(®) (CHGP) and conventional CH paste, for their ability to maintain alkalinity and to assess the clinical antimicrobial effect of CHGP. METHODOLOGY: Calcium hydroxide products were tested in the presence of standardized dentine powder or by titrating them with 1 mol HCl, expressed in mL (±SD). In a clinical trial, 21 single-rooted teeth with primary apical periodontitis were medicated with CHGP or with conventional CH paste. Bacterial samples were taken before and after chemo-mechanical preparation, after dressing and after leaving canals empty but sealed. To compare groups, anova with Tukey's test was used in the laboratory study and Fisher's exact test in the clinical study. Significance level was set at 5%. RESULTS: Pure CH with water (8.5 ± 0.1) and Calasept (9.3 ± 0.1) maintained the highest alkalinity, followed by the gel-like products DS CaOH gel (7.3 ± 0.3) and Ultracal XS (6.8 ± 0.2) and then Biokalkki (6.3 ± 0.3) and Calxyl blue (5.1 ± 0.2). All CH paste products had higher values compared with CHGP (1.6 ± 0.1) (P < 0.05). Saturated solutions of the products were all neutralized to pH 8.6 within 24 h by dentine powder addition. Clinically, culture-negative results were obtained in 5/10 canals in the CHGP group and 7/11 with conventional CH (P > 0.05). CONCLUSIONS: Aqueous CH mixtures kept high pH better than viscous gel products or CHGP. Dentine powder had equal buffering effect on each product tested. CHGP and traditional CH paste both had an antimicrobial effect in the clinical setting, but there was no significant difference between the groups.

In vitro yeast infection of human dentin.

An in vitro model was developed for investigation of Candida albicans penetration into human dentinal tubules. The model consisted of a dentin disc mounted between two cuvettes that each had a circular opening facing the disc. The cuvettes were filled with Tryptic-Soy-Broth, and the pulpal side cuvette was inoculated with C. albicans and incubated at 37 degrees C in air until growth occurred in the uninoculated cuvette or up to 30 days. The system was also used with Enterococcus faecalis. Completely glue-covered dentin specimens served as negative controls. Brown & Brenn-stained histological preparations of the specimens were examined with light microscopy. The time needed before growth occurred in the uninoculated cuvette showed great variation with C. albicans, whereas E. faecalis penetrated within 1 to 5 days of incubation. Slight penetration both by hyphae and yeast cells was observed in specimens inoculated with C. albicans, whereas specimens inoculated with E. faecalis showed deep and effective penetration. This study demonstrates the penetration of dentin as a possible pathway of infection by C. albicans. However, dentin penetration by C. albicans was slow and limited in comparison with E. faecalis.

Inactivation of local root canal medicaments by dentine: an in vitro study.

AIMS: The aim of the study was to investigate the inactivation by dentine of the antibacterial activity of various commonly used local root canal medicaments. METHODOLOGY: The medicaments tested were saturated calcium hydroxide solution, 1% sodium hypochlorite, 0.5% and 0.05% chlorhexidine acetate, and 2/4% and 0.2/0.4% iodine potassium iodide. Dentine was sterilized by autoclaving and crushed into powder with a particle size of 0.2-20 microns. Aliquots of dentine suspension were incubated with the medicaments in sealed test tubes at 37 degrees C for 24 h or 1 h before adding the bacteria. In some experiments bacteria were added simultaneously with dentine powder and the medicament. Enterococcus faecalis A197A was used as a test organism. Samples for bacterial culturing were taken from the suspensions at 5 min, 1 h and 24 h after adding the bacteria. RESULTS: Dentine powder had an inhibitory effect on all medicaments tested. The effect was dependent on the concentration of the medicament as well as on the length of the time the medicament was preincubated with dentine powder before adding the bacteria. The effect of calcium hydroxide on E. faecalis was totally abolished by the presence of dentine powder. Similarly, 0.2/0.4% iodine potassium iodide lost its effect after preincubation for 1 h with dentine before adding the bacteria. The effect of 0.05% chlorhexidine and 1% sodium hypochlorite on E. faecalis was reduced but not totally eliminated by the presence of dentine. No inhibition could be measured when full strength solutions of chlorhexidine and iodine potassium iodide were used in killing E. faecalis. CONCLUSIONS: The dentine powder model appears to be an efficient tool for the study of interactions between local endodontic medicaments, dentine, and microbes.

Susceptibility of oral Candida species to calcium hydroxide in vitro.

AIM: The susceptibility of common oral Candida species to saturated aqueous calcium hydroxide solution was studied. METHODOLOGY: The yeast species tested were C. albicans (16 strains). C. glabrata (three strains), C. guilliermondii (three strains), C. krusei (two strains), and C. tropicalis (two strains). At least one reference strain of each species was used; the others were clinical isolates either from persistent apical periodontitis or from marginal periodontitis. The susceptibility of Enterococcus faecalis (ATCC 29212) was studied for comparative purposes. Standardized inocula of the strains were incubated in aqueous calcium hydroxide solution, pH 12.4, for time-periods ranging from 5 min to 6 h. Volumes of 0.1 mL of the test suspension were cultured directly on Brucella blood agar and incubated in air at 37C. The plates were inspected for growth at 24 and 48 h and the colonies were counted. The time required to reduce the number of colony-forming units to less than 0.1% of the initial number was determined for each strain. RESULTS: The sensitivity of the C. albicans strains was generally low, with 16 h of incubation required to kill 99.9% of the colony-forming units. No differences in susceptibility between C. albicans strains isolated from root-canal infections and from periodontitis were found. Both strains of C. tropicalis were killed between 3 and 6 h of incubation, whilst strains of C. guilliermondii were killed after only 1020 min of incubation. All strains of C. glabrata survived 20 min, but not 1 h, of incubation, whilst 13 h were required to kill C. krusei. Compared with E. faecalis, all Candida spp. showed either equally high or higher resistance to aqueous calcium hydroxide. CONCLUSIONS: This study indicates that Candida spp. are resistant to calcium hydroxide in vitro, which may explain the isolation of yeasts from cases of persistent apical periodontitis.

In vitro susceptibility of Candida albicans to four disinfectants and their combinations.

AIM: The aim of this study was to evaluate the susceptibility of seven strains of Candida albicans to four disinfectants: iodine potassium iodide, chlorhexidine acetate, sodium hypochlorite and calcium hydroxide. In addition, all possible pairs of the disinfectants were tested in order to compare the effect of the combination and its components. METHODOLOGY: Filter paper discs were immersed in standardized yeast suspensions and then transferred to disinfectant solutions of different concentrations and incubated at 37 degrees C for 30 s, 5 min, 1 h and 24 h. After incubation the filter paper discs were transferred to vials with PBS and glass beads that were then vigorously shaken for dispersal of the yeast cells. PBS with resuspended yeasts was serially diluted 10-fold. Droplets of 25 microL from each dilution were inoculated on TSB agar plates and incubated in air at 37 degrees C for 24 h. The number of colony-forming units was then calculated from appropriate dilutions. RESULTS: C. albicans cells were highly resistant to calcium hydroxide. Sodium hypochlorite (5% and 0.5%) and iodine (2%) potassium iodide (4%) killed all yeast cells within 30 s, whilst chlorhexidine acetate (0.5%) showed complete killing after 5 min. Combinations of disinfectants were equally or less effective than the more effective component. All C. albicans strains tested showed similar susceptibility to the medicaments tested. CONCLUSIONS: This study indicates that sodium hypochlorite, iodine potassium iodide and chlorhexidine acetate are more effective than calcium hydroxide against C. albicans in vitro. However, combining calcium hydroxide with sodium hypochlorite or chlorhexidine may provide a wide-spectrum antimicrobial preparation with a long-lasting effect.

Fungi in therapy-resistant apical periodontitis.

The occurrence of yeasts in 967 microbiological endodontic samples taken from root canals in persistent endodontic infections was studied. The sampling was done by general practitioners in various parts of Finland from root canal infections which did not respond favourably to standard conservative therapy. The samples were cultivated aerobically on a non-selective enriched horse blood agar medium, on TSBV agar medium in 5% CO2 and anaerobically on horse blood agar medium. Micro-organisms were found in 692 of the samples while 275 showed no growth. Forty-eight fungi were isolated from 47 samples which is 7% of the culture-positive samples. Twenty yeast strains were identified further by their colony morphology, growth and cellular characteristics and patterns of carbohydrate assimilation. All isolates except one belonged to the genus Candida. Candida albicans was the most common species. C. glabrata was found together with C. albicans in one sample. C. guilliermondii, C. inconspicua and Geotrichum candidum were each isolated once. Yeasts were found in pure culture in six samples and together with bacteria in 41 samples. In all the samples except two, the accompanying facultative bacteria were Gram positive. The most frequent of them were alpha- and non-haemolytic Streptococcus species which were found in 31 samples. Anaerobic bacteria were isolated together with yeasts from 12 root canals. They included both Gram positive species such as Peptostreptococcus micros and Gram negative species such as Fusobacterium nucleatum. The regular isolation of yeasts, also in pure culture, indicates that yeasts may have an important role in cases of apical periodontitis persisting after conventional treatment.

Microbiological findings and clinical treatment procedures in endodontic cases selected for microbiological investigation.

The relationship between bacteriological findings and clinical treatment procedures was investigated in root canal treatment cases that were selected for bacteriological investigation by general dental practitioners in Finland. The cultures were sent to the Oral Microbiological Service Laboratory at the Institute of Dentistry in Helsinki. Two groups of teeth were selected based on the type of infection present in the root canal system. The 'enteric bacteria' group consisted of 40 sequential cases where Enterococcus faecalis and/or other facultative enteric bacteria or Pseudomonas sp. were found in the samples in pure culture (35%) or together with other types of bacteria. The group 'non-enteric bacteria' consisted of 40 sequential cases where only non-enteric bacteria were found. The dentists who had sent the bacteriological samples received a questionnaire where they were asked about the treatment protocol and procedures. A total of 70 out of 80 questionnaires were returned. If the root canals had been unsealed at some point during the treatment, enteric bacteria were found more frequently than in canals with an adequate seal between the appointments. Of cases with enteric bacteria 55% had been open during the treatment, while in the group where only non-enteric bacteria were found 30% had been open. Enteric bacteria were also more frequently isolated in cases with a high number of appointments before sampling. In the enteric bacteria group 35% of the samples were taken at the 10th visit or later, while the corresponding percentage in the non-enteric group was 3%. In addition, the number of retreatment cases was significantly higher, 12 out of 34, in the enteric bacteria group than in non-enteric bacteria group, which was five out of 36. Other clinical parameters showed no differences between the two groups. The results emphasize the importance of controlled asepsis throughout the root canal treatment.