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Azerbaycan Saytlari

 »  Home  »  Endodontic Articles 3  »  An in vitro comparison of the bactericidal efficacy of lethal photosensitization or sodium hyphochlorite irrigation on Streptococcus intermedius biofilms in root canals
An in vitro comparison of the bactericidal efficacy of lethal photosensitization or sodium hyphochlorite irrigation on Streptococcus intermedius biofilms in root canals
Materials and methods.



A helium–neon gas laser with a measured power output of 35 mW (Spectra-Physics, Hemel Hempstead, UK) was used. The emitted radiation was collimated in a beam of 0.25 cm, with a wavelength of 632.8 nm. TBO photosensitizer (Sigma Ltd, Poole, UK) was prepared to various concentrations (0, 12.5, 25, 50 and 100 gmL –1 ), filter sterilized (0.2 m filter, Whatman International Ltd, Maidstone, UK) and stored in light excluded containers until required.
Thirty-five freshly extracted, intact, adult, human, single-rooted, mature teeth with a single canal were collected and stored in sterile saline. Calculus and stains were removed from the root surface using an ultrasonic scaler (Cavitron, Dentsply Ltd, Weybridge, UK). After accessing, the root canal was prepared to an apical size 25 using Flexofiles® (Maillefer Instruments SA, Ballaigues, Switzerland) with a 10% taper using Greater Taper™ hand files (Dentsply, Weybridge, UK) to a point 1 mm short of the apical foramen, which was subsequently sealed with composite restorative material (Durafill®, Heraeus Kulzer, Hanau, Germany). The root canal and outer surfaces were irrigated with 17% (v/v) Ethylenediaminetetra-acetic acid (EDTA) (Merck, Poole, UK) for 2 min, followed by tap water. The prepared tooth was mounted in the lid of a bijou bottle. The assembled tooth, lid and bottle were covered with aluminium foil and autoclaved at 121 C, for 15 min. The bijou bottle was then aseptically filled with sterile brain heart infusion broth (BHI) (Oxoid Ltd, Basingstoke, UK) so that the root was covered.
A root canal isolate identified as Streptococcus intermedius (by 16S rRNA gene sequence analysis) was used throughout the study. A frozen (–70 C, in BHI/glycerol, 10% v/v) stock ampoule was thawed and inoculated onto blood agar (BA) plates (Oxoid Ltd, Basingstoke, UK) containing 5% (v/v) defibrinated horse blood, incubated for 24 h at 37 C in a CO 2 incubator (B6420 Heraeus, Hanau, Germany). Cultures were then stored at 4 C until required and subcultured on a weekly basis. Bacterial cells were collected with a sterile swab and resuspended in 1 mL of sterile BHI broth to obtain an optical turbidity of 0.5 McFarland standard and incubated for 1 h at 37 C in the CO 2 incubator. The inoculum was syringed into the prepared root canal using a sterile endodontic irrigating syringe (Monoject™, Sherwood Medical Company, St. Louis, MO, USA) and incubated for 48 h under standard conditions as described previously.
Any residual medium in the root canals after the 48 h incubation was carefully removed with sterile paper points and the teeth were randomly selected for one of 31 treatment regimens. In order to reduce the effect of root canal anatomy as a variable, the 35 teeth were reautoclaved and reused up to a maximum of four times. After each test, the contents of the root canal and bijou bottle were emptied, the tooth irrigated with EDTA for 2 min, rinsed with water, reassembled in the bijou bottle and autoclaved before further use.

Test groups.

  • Lethal photosensitization.
    Root canal biofilms were subjected to lethal photosensitization using 20 combinations of four TBO concentrations and five laser energy doses ( n = 4 for each combination). The canals were filled to the level of the access cavity with TBO at various concentrations (12.5, 25, 50 or 100 gmL –1 ) and incubated for 30 s. Laser light was then applied for a specified time (60, 90, 120, 300 or 600 s equivalent to energy doses of 2.1, 3.2, 4.2, 10.5 or 21 J) at the orifice of the access cavity without introducing the laser optic fibres into the root canal.
  • Sodium hypochlorite irrigation.
    The canals ( n = 4) were filled with NaOCl (3% v/v) for 5 min, removed with sterile paper points and fresh NaOCl solution was added for a further 5 min.

Control groups.

  • Laser light only.
    The canals ( n = 4 for each dose) were filled with reduced transport fluid (RTF) (Syed & Loesche 1972) for 30 s followed by application of various laser light doses (60, 90, 120, 300 or 600 s).
  • TBO only.
    The canals ( n = 4 for each concentration) were filled with TBO at various concentrations (12.5, 25, 50 or 100 gmL –1 ) and incubated for 30 s.
  • No treatment.
    The canals ( n = 17 included positive controls for several experimental runs) were filled with RTF and incubated for 30 s.

Table 1. Viable bacteria recovered after treatment of S. intermedius biofilm in root canals with various combinations of laser energy and TBO concentration.

Viable bacteria recovered after treatment of S.intermedius biofilm in root canals with various combinations of laser energy and TBO concentration
Mean CFU per canal and standard deviation in parentheses; (n = 4 except as shown).
a) Test groups subjected to lethal photosensitization.
b) Control groups subjected to laser light only (TBO = 0 gmL-1).
c) Control groups subjected to TBO only (laser = 0 s)
d) Control group subjected to no treatment.

Post-treatment root canal sampling.
Following all treatments, the liquid content of the root canal was carefully absorbed with paper points without intentionally touching the canal walls to avoid interference with the biofilm. The canal was refilled with RTF, circumferentially filed with a sterile, size 25 Flexofile® to the working length for 20 s. The resulting bacterial suspension was completely absorbed with paper points and transferred to 1 mL of RTF. After vortexing for 30 s, a 7-fold (neat to 10 –7 ) serial dilution was prepared and 25 L of each was inoculated onto a blood agar plate and incubated for 24 h as described previously. Colony forming units (CFU) recovered from each treated root canal were calculated.