Transcript Identification of Bacteria in Periapical Lesions Using DNA

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Bacterial Adhesion on Integrated Abutment Crowns TM.
In Vitro Study (I)
ABSTRACT. Objectives: The goal of the present investigation was
to determine if the Diamond-crown material, used to make Bicon's
Integrated Abutment Crown (IAC), is less susceptible to
harbor/attract bacterial plaque than All Ceramic (AC) or Metal
Ceramic (MC) crowns. A secondary goal was to compare the
composition of the plaque attracted on tested surfaces.
Methods: 6 IAC, 6 AC and 6 MC crowns were equally divided in 2
test groups. The AC and MC crowns were cemented on titanium
abutments. Group I (3 IAC,3 AC, 3 MC)and group II were
incubated for 10 min in a bacterial solution containing 10 different
oral bacteria at O.D.1: Tanerella forsythensis, Prevotella
intermedia, Campylobacter rectus, Fusobacterium nucleatum,
Actinomyces odontololyticus, A. naeslundii, Streptococcus
intermedius, S. oralis, Actinobacillus actinomycetemcomitans
serotype b, Porphyromonas gingivalis. After a brief wash in PBS to
eliminate the unbound plaque, the crown samples in group I were
incubated in 500µl of Tris-EDTA buffer with 500µl of NaOH. The
samples were then hybridized with 10 whole chromosomal probes
to the above mentioned microorganisms. The microbiological
analysis was completed using the checkerboard DNA-DNA
hybridization method. The samples in group II were briefly washed
in PBS and fixed in 4% formalin for scanning electron microscopy
(SEM)
Results: All experimental crowns showed bacterial adhesion. There
was no statistical difference in the microbial compositions when
comparing crowns. The SEM showed that the AC crowns were
harboring the heaviest bacterial deposits. MC and IAC showed the
least bacterial deposits especially at the abutment/crown interface.
Conclusions: The IAC and MC crowns appear to be less prone to
bacterial colonization, in an in vitro setting then an all ceramic
crown. IAC and MC crowns harbored very few bacteria at the
abutment/crown interface
This study was supported by a research grant from Bicon, Inc.
1
DIBART ,
2
MARINCOLA ,
1
WARBINGTON ,
3
SKOBE
S.
M.
M.L.
and Z.
1 Boston University, MA, USA, 2 University of Cartagena, AISI, Italian Association of Restorative
3
Implant Dentistry, Rome, Italy, Forsyth Institute, Boston, MA
INTRODUCTION
The trauma surrounding the partial or total loss of the
natural dentition to periodontal diseases or caries has been
alleviated in the last decades by the introduction of the
concept of predictable osseointegration to the dental
profession 1,2. Successfully osseointegrated dental implants
have revolutionized the practice of late 20th century
dentistry. They provided patients with the ability to have
fixed restorations instead of removable devices, and help
avoid the mutilation of adjacent natural teeth when a 3 unit
fixed partial restoration was envisioned. As with all
prosthetic restorations in the oral cavity, they are subject to
factors impacting esthetics, function, and periodontal
health. Periodontal health or peri-implant health is the most
critical aspect of this trilogy, since compromising it could
mean potentially disastrous effects on esthetics and
function. The tissues supporting dental implants are
susceptible to disease (peri-implantitis), which in turn
could lead to bone loss and implant failure. The disease
process is initiated by microorganisms that are present in
the periodontal plaque. These bacteria and their byproducts (enzymes, toxins, metabolic products), in the
susceptible host, will start a whole cascade of events that
will lead to periodontal tissue damage. bacteria need to
adhere to a solid surface (i.e. prosthetic or natural crown,
soft tissue etc.) in their primary phase of colonization
before causing the disease. A restoration material that
would repel or cause bacteria to adhere minimally would
be a plus in preventing disease.
In June 2001, Bicon Inc. (Boston, MA) introduced the
Integrated Abutment Crown ™ (IAC). This is a new concept
where the implant abutment and the crown material are one
integral unit (Fig.1). A poly-ceramic material such as
Diamond Crown ™ (DRM Research Labs Inc., Branford,
CT) is fused onto the coronal post of a titanium alloy
abutment. The IAC is then placed directly into the well of
the implant, there is no need for cementation or screw
A
retention.
The goal of the present investigation was to determine if the
Diamond Crown™ material used to make Bicon’s IAC is less
susceptible to harbor/attract bacterial plaque than All Ceramic
(AC) or Porcelain Fused to Metal (PFM) crowns.
RESULTS
METHODS
Figure 3. Scanning electron micrograph of a
Porcelain fused to metal crown. Top of the
micrograph shows crown abutment junction,
with cement closing the gap. Lower half of
the micrograph shows the porcelain margin at
high magnification, with very few or no
bacteria present.
In the in vitro phase we tested bacterial presence and composition comparing
the Integrated Abutment Crown to All Ceramic (porcelain Noritake Cerabien,
Noritake Co., Nagoya, Japan) and Porcelain Fused to Metal (Noritake Super
Porcelain EX-3, Noritake Co., Nagoya, Japan) crowns in a laboratory setting.
The All Ceramic and the Porcelain Fused to Metal crowns were cemented on
the abutments using Fuji Plus cement (GC Corporation, Tokyo, Japan).
A total of 18 crowns were analyzed for bacterial presence/adhesion. All the
crowns were provided by the Bicon corporation for our testing. The 18 crowns
were divided in 2 groups. Group 1 which underwent microbiological analysis
and group 2 which underwent SEM analysis. These groups were further subdivided in 1A (3 IAC), 1B (3 All Ceramic Crowns), 1C (3 Porcelain Fused to
Metal Crowns) and 2A, 2B, 2C.
The 9 crowns of group 1 were incubated for 10 minutes in a 100 ml PBS
solution containing 10 oral bacteria at optical density of 1: Tannerella
forsythensis, Prevotella intermedia, Campylobacter rectus, Fusobacterium
nucleatum, Actynomyces odontolyticus, A. naeslundii, Streptococcus
intermedius, S. oralis, Actinobacillus actinomycetemcomitans serotype b,
Porphyromonas gingivalis. The crowns were immersed in a beaker containing
the bacterial solution, with a stir bar. The crowns were removed and rinsed in
sterile PBS for 30 seconds and put in a 15 ml tube containing 0.5 ml of TE (10
mM Tris-HCl, 1 mM EDTA, pH 7, 6). The bacterial DNA was denatured by
adding 0.5 ml of a 0.5M solution of NaOH. The samples were then hybridized
with 10 whole chromosomal DNA probes to these microorganisms according to
the protocol described by Socransky et al. The 9 crowns of group 2 were also
incubated for 10 minutes in the above mentioned bacterial mixture and rinsed in
sterile PBS for 30 seconds. They were then fixed in 4% formalin at room
temperature overnight, metal coated and prepared for scanning electron
microscopy.
Figure
2.
Scanning
electron
micrograph of an all ceramic crownabutment junction. Notice the gap
and lack of cement between
prosthesis (top) and implant abutment
(bottom).
Figure 1. Clinical photograph of a
Bicon Integrated Abutment Crown.
The porcelain is fused to the
abutment, there is no cementation.
Figure 4. Scanning electron micrograph of an
Integrated Abutment Crown. Top half of the
micrograph shows crown-abutment interface,
with no gap. Lower half of the micrograph
shows the crown margin at high magnification,
with very few or no bacteria present.
All experimental crowns (IAC, AC, PFM) showed bacterial
adhesion. All 10 bacterial species were found to adhere to each
individual crowns. There was no statistical difference in the
microbial composition found on the crowns investigated. This is
probably due to the small size of the sample that did not show any
statistically significant changes. The SEM analysis showed a clear
difference in terms of amount and location of the microorganisms
on the various crowns. All ceramic (AC) crowns harbored the
heaviest bacterial deposits (Fig 2) and this was true for all areas
studied, from occlusal surface to crown margins. Also more than
one type of microorganism could be differentiated morphologically.
This was confirmed by the microbiological analysis of the samples,
all of the 10 microbial species could be detected using DNA probes
(data not shown). At the abutment/crown interface a lack of cement
was noticed as well as porosity in the ceramic structures (Fig 4).
Many bacterial colonies were noticed in that area including in the
micro-gap. Bacteria were also present, albeit at a much lower
number, on porcelain fused to metal (PFM) and integrated abutment
crowns (IAC).
Bacterial colonization was also distributed
differently. A few clusters of bacteria were present on the occlusal
third of the IAC and extremely few on the PFM. Of much interest
was the observations carried out at the crown margins (PFM) and
abutment/crown interface (IAC).
There were very few
microorganisms observed at the abutment/crown interface for PFM
and IAC. These 2 entities appeared to show the least amount of
bacterial deposits (Figs. 3 and 4).
CONCLUSION
The results of this in vitro study seem to show that bacterial
presence is inevitable on any type of prosthetic restoration.
However the IAC design seems to go toward reducing this
eventuality by eliminating the gap between implant abutment and
crown, and providing a smooth cervical interface.