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Bactericidal Effect of The Er:YAG Laser On Dental Implant Surfaces: An in Vitro Study
Bactericidal Effect of The Er:YAG Laser On Dental Implant Surfaces: An in Vitro Study
Volume 73 • Number 11
P
eri-implant infection results in inflam-
10556). Irradiation at pulse energies of 60 and 120 mJ and a mation of the surrounding soft tissues
frequency of 10 pps was performed on a computer-controlled and can induce a breakdown of the
XY translation stage. After laser treatment the specimens were implant-supporting bone (Figs. 1 and 2).
sonicated and the bacterial growth examined by counting colony Bacterial adherence to implant surfaces is
forming units on blood agar plates. Temperature elevations dur- a complex process not yet fully under-
ing irradiation were investigated using K-type thermocouples. stood.1,2 Several physical and biological
Laser treated implant surfaces were analyzed by means of elec- factors seem to influence the adhesion
tron microscopy. process. Surface roughness plays a major
Results: Compared to non-irradiated specimens, mean bac- role in the colonization process, sheltering
terial reductions of 99.51% (SA), 98.39% (HA), and 99.6% (TPS) the microorganisms from removal by sali-
at a pulse energy of 60 mJ and 99.92% (SA), 99.85% (HA), and vary flow and oral hygiene procedures.3-5
99.94% (TPS) at 120 mJ were calculated. At these laser param- The surface-free energy of the bacterium,
eters, no excessive temperature elevations or morphological the ionic strength of the surrounding liquid
implant surface alterations were detected. medium, and the distance of the bacterium
Conclusions: Even at low energy densities, the Er:YAG laser from the surface affect non-specific elec-
has a high bactericidal potential on common implant surfaces. trostatic interaction between the cells and
Clinical studies are justified to evaluate the applicability and effi- the colonized substratum.4,6,7 Further stud-
cacy of the Er:YAG laser in the treatment of peri-implantitis. J ies indicate that different strains have dif-
Periodontol 2002;73:1292-1298. ferent affinities to implant surfaces and that
KEY WORDS the type of organism, concentration, and
growth phase together with surface char-
Lasers/therapeutic use; dental implants/microbiology; peri-
acteristics influence the colonization
implant diseases/prevention and control.
process.8-13 Compared to biofilms on other
biomaterials, such as indwelling catheters
* Department of Oral Surgery, Johannes Gutenberg University, Mainz, Germany. and contact lenses, the communities of
† Department of Hygiene and Environmental Medicine.
‡ Institute for Applied Structure and Microanalysis. organisms on implant surfaces are very dif-
ficult to eradicate. Several treatment regi-
mens have been proposed for cleaning and
decontamination of implant surfaces. Plas-
tic curets are probably best for manual
removal of peri-implant plaque,14 as metal
curets and the application of ultrasonic
scalers induce surface alteration in implants
and are, therefore, contraindicated.15 Bac-
tericidal chemicals such as chlorhexidine
digluconate or iodine as well as local and
systemic administration of antibiotics are a
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Figures 1 and 2.
Severe peri-implant inflammation with pronounced bone loss. Note the partly exposed Ti plasma-sprayed implant surface and the massive bacterial
contamination.
useful adjunct in the treatment of peri-implantitis.16 pended in PBS. The viable bacterial colony counts
Sterilization and cleaning of implant surfaces using were determined for quantification of bacterial cell
lasers has been suggested.17,18 However, the bacteri- numbers present in suspensions of known optical den-
cidal potential of some laser systems on roughened sity (625 nm). The mean concentration was 8 × 108
surfaces requires considerable scientific investigation. cells/ml prior to specimen inoculation. The solution
Since its 2940 nm wavelength is highly absorbed in was sonicated to disperse cell clumps.
water, the Er:YAG laser is useful in various dental appli- Colonization Model
cations. It has been mainly studied for hard tissue Preliminary experiments were performed to determine
removal and for various indications in oral surgery.19,20 optimal test conditions. To ensure that only the respec-
Other studies demonstrated its applicability in the treat- tive surface on the upper side and no other parts of
ment of periodontal diseases and endodontics.21,22 It the discs were contaminated, the discs were embed-
is feasible that the sterilizing effects of the Er:YAG laser ded between 2 aluminum plates, 1 with 12 perfora-
might be useful in the treatment of ailing implants. tions for the upper side of the specimens. The space
This study is part of a research program investi- between the plates and the discs was isolated with flow
gating the possibilities and hazards of laser application impression paste. This mounting and the platelets were
in implant dentistry. The aim was the examination of incubated with the cell suspension for 1 hour at 37°C.
the bactericidal effect of Er:YAG laser irradiation on Discs with similar surfaces were incubated simultane-
common implant surfaces. ously. After incubation the 12 discs were removed from
the mounting and washed with PBS to remove non-
MATERIALS AND METHODS
adherent cells. Four specimens served as controls; 4
Titanium Discs were irradiated at 60 mJ and 4 at 120 mJ. Except for
Test platelets§ made from commercially pure (CP) the irradiation process, all specimens were treated
titanium (grade 4) with a thickness of 1.5 mm and a equally.
diameter of 10 mm with 3 different surfaces (sand-
blasted and acid etched, SA; titanium-plasma sprayed, Laser
TPS; and hydroxyapatite-coated, HA) served as sub- An Er:YAG laser# with a wavelength of 2940 nm and
strates. Surface roughness as indicated by the man- a variable pulse frequency (1 to 15 pps) and pulse
ufacturer was SA: Ra = 2.2 µm; TPS: Ra = 3.41 µm; energy (60 to 250 mJ) was used. Frequency was kept
and HA: Ra = 2.0 µm with a standard deviation of constant at 10 pps. Pulse energy was set alternatively
approximately 20%. at 60 and 120 mJ. The light was delivered by an optic
fiber and a 540 micron application tip. Irradiation was
Target Microorganisms and Culture Conditions performed without water cooling.
Streptococcus sanguis (ATCC 10556) was obtained § Friadent GmbH, Mannheim, Germany.
commercially and cultured on blood agar plates.¶ The German Collection of Microorganisms and Cell Cultures, Braunschweig,
Germany.
bacterial cells were washed 3 times with phosphate- ¶ Heipha Dr. Müller GmbH, Heidelberg, Germany.
buffered saline (PBS, pH = 7.2) and afterwards sus- # Key II, KaVo, Biberach, Germany.
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Irradiation
Subsequent to incubation, the discs were affixed on
an XY translation stage.** The movement of the
handpiece and, therefore the parameters relevant to
the irradiation of the discs (distance from the tip of
the handpiece to the specimen, irradiation time, and
working angle), were controlled by a computer pro-
gram. The tip of the handpiece was moved to the
center of each disc at a distance of 0.5 mm from the
surface. The optic fiber was positioned perpendicu-
larly to the specimen. After laser activation, the
movement was performed in concentric circles from
the center to the periphery of the disc. The radius of
the circles was successively enhanced by 540 µm,
corresponding to the diameter of the fiber, to ensure
an equal exposure of the entire specimen. The over-
all irradiation time of the entire disc was 60 seconds.
Bactericidal Efficacy
After laser irradiation the discs were placed into 10 ml Figure 3.
of sterile PBS and sonicated†† for 2 × 45 seconds at Streptococcus sanguis after 48-hour incubation on blood agar plates.
75 watts. After serial 10-fold dilution, aliquots of each
dilution (100 µl) were spread on Columbia blood agar
plates¶ (Fig. 3). Colony forming units were counted
after 48 hours incubation. A total number of collected a
log kill = log
bacteria from the control and irradiated samples was b
estimated and the bactericidal efficacy of the laser with a = mean bacteria concentration in the control
treatment was calculated. group, b = mean bacteria concentration in the laser
Scanning Electron Microscopy group, and log = decimal logarithm.
Subsequent to standard specimen preparation, single
RESULTS
discs of the laser and the control groups were placed
in the vacuum chamber of a scanning electron micro- Bactericidal Efficacy
scope‡‡ and micrographed at different magnifications. Table 1 presents the bacterial counts on the SA, HA,
Secondary electrons were detected by means of a stan- and TPS specimens. The mean bacterial count in the
dard SE detector. non-irradiated specimens was 6.38 × 104 in the SA
group 2.73 × 105 in the HA group, and 6.25 × 105 in
Temperature Measurements the TPS group. The application of the Er:YAG laser
Prior to irradiation, temperature rises of the titanium resulted in a highly significant reduction of colony
discs during a 1-minute irradiation at various pulse forming units in all irradiated specimens at both
energies were measured with a thermocouple system§§ energy levels (P <0.001). The best bactericidal effect
at room temperature (24.0°C). Three 0.5 mm probes was achieved irradiating the TPS surface followed by
(K-type) were attached to the border of the specimens. the SA and HA surfaces. The mean reduction in the
The sensitive tip of the probes was covered with a ther- TPS group expressed in log steps was 2.4 at a pulse
moconductive paste. energy of 60 pps and 3.21 at 120 mJ correspond-
Statistical Analysis ing to a bacterial reduction of 99.60% and 99.94%,
The statistical analysis was performed with a spread respectively. The values were slightly lower in the SA
sheet and a statistic package.¶¶ For each implant group: 2.31 (99.51%) at 60 mJ and 3.11 (99.92%)
surface, 8 samples served as control, 8 were irradiated at 120 mJ and in the HA group: 1.79 (98.39%) at
at 60 mJ, and 8 at 120 mJ. The median, mean, and 60 mJ and 2.82 (99.85%) at 120 mJ. The differ-
standard deviation of the bacteria concentration of the ence in bacterial counts between the 2 energy lev-
control and the 2 laser groups were calculated. Dif- els was significant in all groups (P <0.001) and
ferences between the groups were analyzed by means
of the Mann Whitney-U test and differences consid- ** HBM, Langen, Germany.
†† Branson Ultrasonics Corp., Danbury, CT.
ered to be significant at P <0.05. The bacterial reduc- ‡‡ LEO 435 VP, Zeiss-Leica Corp., Oberkochen, Germany.
tion was quoted in “log kills” in accordance with the §§ Omega Engineering, Stanford, CT.
Excel 97, Microsoft, Richmond, VA.
work of Rooney et al.23 and calculated as follows: ¶¶ SPSS for Windows, Release 10.0.5, SPSS Inc., Chicago, IL.
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DISCUSSION
Beside undisturbed osseointegration and an adequate
ranged between 0.8 and 1.03 log steps. Complete prosthetic design, maintenance is crucial for long-term
bacterial reduction could not be achieved. implant prognosis. Clinical success can be jeopardized
by bacterial infection inducing mucositis or peri-implan-
Scanning Electron Microscopy titis. Elimination of pathogenic microorganisms from
Figures 4 and 5 show the microscoping appearance implant surfaces is a prerequisite for successful treat-
of Streptococcus sanguis on the SA surface after incu- ment of ailing implants. Several in vitro studies demon-
bation. A dense colonization was predominant with strated a sterilizing effect of various laser systems on
Figures 4 and 5.
Scanning electron micrograph of a sand-blasted and acid-etched implant surface after incubation with a suspension of S. sanguis (original
magnification ×2,000 and ×5,000).
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Figures 6 and 7.
Scanning electron micrograph after laser irradiation at 120 mJ (original magnification ×1,000 and ×2,000).
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