advanced diagnostic aids in periodontics

DEPARTMENT OF PERIODONTOLOGY
BY- Anamika Singh
MDS 2nd Year

CONTENTS
 Limitations of conventional periodontal diagnosis
 Advances in Clinical Diagnosis
 Gingival Bleeding
 Gingival Temperature
 Periodontal Probing
 Advances in radiographic assessment
 Bacterial Culturing
 Direct Microscopy
 Immunodiagnostic Methods
 Enzymatic Methods
 Diagnostic Assays Based on Molecular Biology Techniques
 Advances in characterizing the host response
 Source of Samples
 Inflammatory Mediators and Products Host-Derived Enzymes
 Tissue Breakdown Products
 Chair-side diagnostic Aids

Periodontal diseases are conventionally diagnosed by clinical evaluation of
signs of inflammation in gingiva without periodontal tissue destruction
(gingivitis) or by presence of both inflammation and tissue destruction
(periodontitis).
• Traditional clinical diagnosis of periodontitis is made by measuring
either the loss of connective tissue attachment to root surface, i.e.
clinical attachment loss or the loss of alveolar bone, i.e. radiographic
loss.
• Disease evaluation attempts to identify and quantify the current
clinical signs of inflammation as well as historical evidence of
damage with its extent and severity .
Conventional Techniques

Limitations
 However, evaluation can not reliably identify sites with ongoing periodontal
destruction and does not provide information on cause of condition,
patient’s susceptibility to disease whether it is in progression or remission
or whether the response to therapy will be positive or negative.
 Disease susceptibility is related to the whole person rather than local site.
 However periodontal disease is considered to be site specific, it has
multifactorial origin in which periodontal pathogens, host response and
genetic systemic and behavioral risk factors play role in development of
disease.

Therefore consideration should be given to include
microbiologic, immunologic, systemic, genetic and behavioral
factors in addition to traditional clinical and radiographic
parameters when assessing patient status.

❖ Gingival Bleeding
•
•
•
•
Clinical evaluation of degree of gingival inflammation includes
assessment of redness and swelling of gingiva along with
assessment of gingival bleeding.
Gingival bleeding is related to persistent presence of plaque on
teeth and regarded as a sign of associated inflammatory
response.
Use of gingival bleeding as an indicator of inflammation has
the clinical advantage of being more objective, because color
changes requires subjective estimation.
Gingival bleeding is a good indicator of presence of
inflammatory lesion in connective tissue at the base of sulcus.
The severity of bleeding increases with an increase in size of
inflammatory infiltrate.
Advances In Clinical Diagnosis

•
• Besides an indicator of gingival inflammation, gingival bleeding
is also an indicator of disease activity however, its relationship to
disease progression is unclear.
• Limitation of the use of bleeding as an inflammatory parameter is
the possibility that healthy sites may bleed on probing, any force
greater than 0.25N may evoke bleeding in healthy sites with an
intact periodontium.
• Absence of bleeding on probing indicates periodontal stability with
high probability.
• Periodontal probe or a wooden interdental cleaner is used to elicit
gingival bleeding

•
• Tobacco smoking may mask the inflammatory signs of gingivitis
and periodontitis, particularly the propensity of gingiva to bleed on
brushing, eating or after periodontal probing .
• Mechanism by which smoking may exert a suppressive action on
the bleeding responsiveness of the gingivae are not well understood.

•
• Commercially available system periotemp probe (abiodent),
enables the calculation of temperature differential (with the
sensitivity of 0.1℃) between the probed pocket and subgingival
temperature.
Subgingival temperature at diseased sites is increased as compared to normal
healthy sites.
• Thermal probes are sensitive diagnostic devices for measuring early
inflammatory changes in gingival tissue.
❖ Gingival Temperature
consists of 5 interconnected parts, a probe
tip, an autoclavable handpiece, a console
containing the electronic components with
an illuminated display, an
electronic printer and a foot activating
pedal switch.

Posterior sites are warmer than anterior region.
•
Mandibular sites are
warmer than maxillary
sites.
A possible
explanation is an
increase in cellular
and molecular
activity caused
by increased
periodontal
inflammation
with increasing
probing depth.
• Individual temperature differences are compared with those expected for each
tooth and higher temperature pockets are signaled with a red emitting diode.

Related Studies
 The most comprehensive temperature studies to date were reported by
Haffajee et al. (1992, a, b, c), who carried out three extensive
investigations using the Periotemp device.
 In the first study, the relationship of the subgingival temperature to
baseline clinical parameters was assessed.
 A significant, albeit weak positive correlation was demonstrated between
temperature differences and the percentages of sites with plaque, sites
which were red, sites which bled on probing and sites with loss of
attachment >3mm.
Haffajee, A. D., Socransky, S. S. & Goodson, J. M. (1992a) Subgingival temperature (I). Relation to baseline clinical parameters.
Journal of Clinical Periodontologv 19, 401- 408.

 The second paper in the series (Haffajee et al. 1992,b) assessed whether
subgingival temperature could be used as a risk indicator for future
attachment loss.
 A number of temperature variables appeared to be useful in identifying
subjects and sites at risk of new attachment loss namely, higher "whole
mouth" subgingival temperatures, and a mean subgingival temperature
>35.5°C.
 Elevated subgingival site temperature was particularly related to
attachment loss in shallow pockets <4 mm.
Haffajee, A. D.. Socransky, S. S. & Goodson. J. M. (1992b) Subgingival temperature (II). Relation to future periodontal attachment
loss. Journal of Clinical Periodontology 19. 409-416.

 The final study in this series (Haffajee et al. 1992c) examined the
relationship of temperature to selected bacterial species.
 The suspected pathogens Prevotella intermedia, Peptostreptococcus micros,
Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans were
present in higher proportions of the total microbiota in sites with higher
temperatures, and the reverse was true for the "beneficial“ Capnocytophaga
species.
 Furthermore, a combination of increased proportion of Fusobacterium
nucleatum and decreased Capnocytophaga species, together with increased
temperature was found to be more effective than temperature alone in
predicting new attachment loss at a site.
Haffajee. A. D.. Socransky, S. S. & Goodson. J. M.(i992c) Subgingival temperature (III). Relation to microbial counts.
Journal of Clinical Periodontology 19, 417-422.

 Variation in subgingival and sublingual temperature
exists in smokers compared to non smokers.
 A study was done to compare subgingival temperature in a group of smokers to
that of a group of non-smokers with similar levels of periodontal disease.
 40 adult subjects, 20 cigarette smokers and 20 non-smokers with evidence of
adult periodontitis were examined.
 Subgingival temperature was measured at 6 sites around each of 4 maxillary
anterior teeth.
Dinsdale CR, Rawlinson A, Walsh TF. Subgingival temperature in smokers and non-smokers with periodontal disease.
J Clin Periodontol. 1997 Oct;24(10):761-6.

A significant difference in
subgingival site temperature
was demonstrated between
the smokers and non-
smokers, with the mean site
temperature being 0.4
degree C warmer in smokers
(p < 0.01).
When healthy or diseased sites
were compared between
smokers and non-smokers,
smokers also had warmer
mean site temperatures than
non-smokers for both healthy
and diseased sites (p < 0.01).
For healthy sites, the smokers
had a mean delta T 0.2 degree
C lower (p < 0.01) than the
non-smokers, representing
warmer sites.
In diseased sites however, delta
T was 0.3 degree C higher (p <
0.01) in smokers, representing
cooler sites.
Smokers had a warmer mean sublingual
temperature than non-smokers.

 For diseased sites, the difference between smokers and non-smokers could
be explained by an inhibition of the inflammatory response in smokers,
resulting in a lower site temperature in the smokers group.
 As the inflammatory response is for the most part, protective (Williams et
al. 1992), this in turn may lead to the higher levels of bone and attachment
loss observed in smokers (Grossi et al. 1994, 1995).

1. First generation probes:(conventional probes)
Conventional manual probes that do not control for probing force or
pressure and that are not suited for automatic data collection.
eg: Williams periodontal probe CPITN probe, UNC-15 probe,
University of Michigan 'O’ probe, Goldman Fox probe, Glickman
probe, Merritt A and B probe Nabers probe.
• The most widely used diagnostic tool for the clinical assessment of connective
tissue destruction in periodontitis is the periodontal probe.
• Gold standard in recording changes over period of time.
Periodontal Probing
Classification of periodontal probes depending on generation.

2. Second generation probe: (Constant force probe)
• Introduction of constant force or pressure sensitive probes allowed for
improved standardization of probing.
• They include the first pressure sensitive probe, Armitage et al and van der
Velden and de Vries and electronic pressure-sensitive probe.
• However, probing errors resulting from data readout, and recording, and
estimation of attachment level can be encountered.
• Moreover, the lack of tactile sensation is another limitation; the probe tip
may pass beyond the junctional epithelium in inflamed sites.
The True Pressure Sensitive Probe, a second generation periodontal probe.
The indicator lines meet at a specified force of 20 gm.

• These probes combine controlled force application, automated
measurements and computerized data capture.
• Digital readings of the periodontal pocket depth measurement can be
stored in a computer.
• Although the automated probes decrease manual probing errors,
disadvantages include deeper penetration of the periodontal probe
beyond junctional epithelium in inflamed or diseased sites, thus
overestimating the periodontal pocket depth.
e.g.: Foster Miller probe, Florida probe, Goodson and Kondon fiber-optic
probe and the Toronto automated probe.
3. Third generation probe: (constant force-automated probes)

Schematic representation of mechanism of action of Foster-Miller probe. When the
ball tip is moved across the root surface, a "catch" is detected at the
cementoenamel junction.
Schematic representation of various parts of the Foster-Miller probe, a
third-generation periodontal probe.
LVDT = linear variable differential transducer; FT = force transducer
accelerator.
The Florida Probe with tip and sleeve diameter.

 These are aimed at recording sequential probe positions along a gingival
sulcus.
 An attempt to extend linear probing in a serial manner to take account
of the continuous and Three dimensional pocket that is being examined.
4. Fourth generation probes: (Three dimensional probes)

• Basically these will add an ultrasound or another device to a fourth
generation probes.
• This probe images and maps the upper boundary of periodontal ligament
and its variation over the time as an indicator of the presence of
periodontitis.
• This technique of periodontal diagnosis by ultrasonic probes involves
projection of an arrow ultrasonic beam with high frequency to the
periodontal pockets.
• The echoes of the ultrasound wave reflected by the crest of the periodontal
ligament are recorded by a transducer located inside the probe hand piece
then transmitted to computer software for analysis.
• The ultrasonic image is constructed and the computer software translates
the data to estimate periodontal pocket depth measurements
e.g.: Ultra sonographic probe.
5. Fifth generation probe:(Noninvasive) Three dimensional
probe.

The US periodontal probe system

Elashiry M, Meghil MM, Arce RM, Cutler CW. From manual periodontal probing to digital 3-D imaging to endoscopic
capillaroscopy: Recent advances in periodontal disease diagnosis. J Periodont Res. 2018;00:1–9

• Lack of sensitivity and reproducibility.
• Readings of clinical pocket depth obtained with periodontal probe
do not normally coincide with the histologic pocket depth because
the probe normally penetrates the coronal level of the junctional
epithelium.
• force, angulations, size of probe, precision of calibration presence of
inflammation.
• All these variables contributes to large standard deviations (0.5-1.3)
in clinical probing result.
{force to probe pocket: 30g}
{force to probe periodontal osseous defect: 50g}
Probing depends on:
Problems with probing:

 Since mid 1980s different probe prototypes are developed & tested to
overcome these limitations.
 One of the problem was of improper penetration of probe related to
improper force of probing which was solved with the development of
pressure sensitive probes which have standardized, controlled
insertion pressure.
 New technology and high tech computers are nowadays becoming the
rule.
 It has provided dentists ideal potential of standardization which
makes future comparison of health & disease simpler, more
precise and cost effective.

• Dental Radiographs are traditionally used to access destruction of
alveolar bone associated with periodontitis.
• They provide information on the periodontium that cannot be
obtained by other noninvasive methods.
Advances in Radiographic Assessment

• These radiographs are 2-D images of 3-D objects.
• Variation in projection geometry
• Variation in contrast and density
• Masking by other anatomic structures (superimposition).
• They are very specific but lack sensitivity.
Conventional radiographs do not confirm bone loss in an
incipient stage as approximately 30% of bone mineral content
must be lost to be noticed on X-ray film. {Bender I. Factors influencing the
radiographic appearance of bony lesions. J Endod. 1982;8:161-170.}

• Capturing radiographic image using a sensor.
• Digital radiography allows the use of computerized images, which can
be stored, manipulated, and corrected for underexposures and
overexposures.
• Digital radiography may yield image properties almost equal to
conventional radiographs, but through digital storage and processing,
diagnostic information can be enhanced.
• Moreover, there is a one-third to half reduction in radiation dose
obtained with digital radiographs compared with conventional
radiographs.
DIGITAL RADIOGRAPHY

• Elimination of chemical processing.
• Increased efficiency and speed of viewing.
• Diagnostic information can be enhanced.
• Computerized storage of radiographs.
• Reduced exposure to the radiation.
Advantages

SUBSTRACTION RADIOGRAPHY
• Subtraction radiography, a well-established technique in medicine, has been
introduced as a technique in periodontal diagnosis.
This technique relies on the conversion of
serial radiographs into digital images.
The serially obtained digital images can
then be superimposed and the resultant
composite viewed on a video screen.
Changes in the density and volume of
bone can be detected as lighter areas
(bone gain) or dark areas (bone loss).
Quantitative changes in comparison with
the baseline images can be detected
using an algorithm for gray–scale levels.

 computer–assisted subtraction radiography.
Limitations: needs paralleling technique and accurate superimposition.

Disadvantage: Identical projection alignment during sequential
radiographs.
Diagnostic SR: uses position device for films and software that corrects
angular alignment discrepancy.
ADVANTAGES
• High correlation between alveolar bone loss and CAL changes.
• Increased detection of small osseous lesion.
• Both quantitative and qualitative visualization.
• More sensitive.

 Recently, new image subtraction methods, called diagnostic subtraction
radiography (DSR), have been introduced combining the use of a
positioning device during film exposure with specialized software
designed for digital image subtraction using conventional personal
computers in dental offices.
 This image analysis software system applies an algorithm that corrects for
the effects of angular alignment discrepancies and provides some degree
of flexibility in the imaging procedure.
 Compared with conventional subtraction radiography and conventional
intraoral radiography, DSR showed statistically significant gains in
diagnostic accuracy over conventional radiographs but no differences with
subtraction radiography.

Computer Assisted Densitometric Image Analysis.
(CADIA)
Video camera measures the light transmitted through radiograph
and the signals from the camera is converted to gray scale image.
Advantage:
• Measures quantitative changes in bone density overtime.
• Higher sensitivity, reproducibility and accuracy as compared to
DSR.

 CADIA comparison between two stage(left) and flapless (right) technique.
Gabric, D., Granic, M., Susic, M., & Katanec, D. (2011). Current Concept of Densitometry in Dental Implantology.
Implant Dentistry - The Most Promising Discipline of Dentistry.

 Based on a study, comparison of the ability of CADIA to detect surgically
induced bone loss with interpretation of digital subtraction images and
conventional radiographic interpretation revealed that CADIA was the
most sensitive of the 3 methods, followed by interpretation of digital
subtraction images which was considerably more sensitive than
conventional radiographic interpretation.
 CADIA was capable of assessing differences in alveolar bone changes due
to periodontal surgery between sites exposed to ostectomy/osteoplasty
and control sites and sites exposed to periodontal surgery without
ostectomy/osteoplasty.
 Finally, CADIA was capable of assessing differences in remodeling activity
over 4-6 weeks after periodontal surgery between 45 surgical sites and 45
control sites.
 The system appears to be the most sensitive of previously described
radiographic interpretation techniques.
Brägger U, Pasquali L, Rylander H, Carnes D, Kornman KS. Computer-assisted densitometric image analysis in
periodontal radiography. A methodological study. J Clin Periodontol. 1988 Jan;15(1):27-37.

Computed tomography is a specialized radiographic technique
that allows visualization of planes or slices of interest.
HOWS IT WORK?
COMPUTED TOMOGRAPHY

 CT scans can identify most inferior alveolar canals when multiple cross-sectional
views are performed.
 CT scans have been shown to be very accurate with the magnification effect, the
same for both the anterior and posterior area, from a range of 0% to 6% in
horizontal as well as 0-4% in the vertical dimension.
 The technique of dental CT also known as DENTASCAN was developed by
Schwartzetal.
 The dental CT can be performed with a conventional CT, a spiral CT or a multislice
CT scanner.
The newer generation
of CT scans produces
axial images
perpendicular to the
long axis of patient by
rotating a radiation
source, which emits
fan-shaped beams 360°
around.
The detectors capture
X-rays, which transmit
the subject and the
data is processed by a
computer.
It is unique in that it
provides images of a
combination of soft-
tissues, bone and blood
vessels.
Surapaneni H, Yalamanchili PS, Yalavarthy RS, Reshmarani AP. Role of computed tomography imaging in dental
implantology: An overview. J Oral Maxillofac Radiol 2013;1:43-7

INDICATIONS IN DENTISTRY
• CT can be used to image the extent of pathologic conditions as
well as help to unravel complex facial fractures.
• CT can also be used to assess the temporomandibular joints and
the paranasal sinuses and for presurgical implant treatment
planning.

1. Used when accurate information regarding the topography of osseous
structure is needed.
2. Soft tissue contour and dimension.
3. To check continuity and density of the cortical plates.
4. Vertical height of the residual alveolar ridges.
5. Density of the medullary space and basilar bone.
6. When determining how much space is available above the mandibular
canal or amount of bone below maxillary sinus to receive a dental implant,
or whether there is a space occupying lesion in the maxillofacial region.
APPLICATIONS
Surapaneni H, Yalamanchili PS, Yalavarthy RS, Reshmarani AP. Role of computed tomography imaging in dental
implantology: An overview. J Oral Maxillofac Radiol 2013;1:43-7

Advantages over conventional radiography
• Eliminates the super imposition of images of structures superficial or deep to
the area of interest.
• Because of inherent high contrast resolution, differences may be
distinguished between tissues that differ in physical density by less than 1%.
• Multiple scans of a patient may be viewed as images in the axial, coronal, or
sagittal planes depending on the diagnostic task, referred to as multiplanar
imaging.

DISADVANTAGES of Computed Tomography
• CT scanning requires specialized equipment and setting.
• Radiologists and Technicians need to be knowledgeable of the
anatomy, anatomic variants and pathology of the jaws as well as
considerations pertinent implant treatment planning.
• CT scan delivers higher radiation dose to the patient as compared to
other modalities used during implant treatment planning.
• Metallic Restorations can cause ring artifacts that impair the diagnostic
quality of the image, it is challenging to the patients having heavy
metallic restored dentition.

■ Routine use of CT in dentistry is not accepted due to its cost and
excessive radiation.
■ In recent years, a new technology of cone-beam CT (CBCT) for
acquiring 3D images of oral structures is now available to the dental
clinics and hospitals.
■ It is cheaper than CT, less bulky and generates low dosages of X-
radiations.
■ The innovative CBCT machine designed for head and neck imaging
are comparable in size with an orthopantomogram.
• It uses a single 360° rotation around the maxillofacial region and a cone
beam, in comparison, a spiral CT, which makes several rotations and uses a
fan beam.

The X-Ray Source and the detector
are dramatically positioned and
make a 360 degree rotation around
this patient head.
In CBCT, scanner generates a Cone shaped
X-Ray Beam, which images larger area.
Images are generated in 1 degree
increments, so at the end of a single
rotation 360 images of area are
generated.
The computer uses these images to generate
a Digital three dimensional map of the face.
Once map is generated, multiplanar
reconstructions as well as axial, coronal,
sagittal, or oblique images are generated.

Applications in periodontics:
• For many decades, 2D imaging was the mainstay in periodontal diagnosis,
however, their limitations led to under / over estimation of the bone loss.
• The literature has confirmed that morphometric analysis of periodontal
diseases by CBCT to be as precise as direct measurement using a
periodontal probe.
• In addition, CBCT is far better than 2D radiographs in visualization of buccal
and lingual defects due to absence of superimposition of the structures.
• CBCT offers precise measurement of intrabony defects and lets clinicians to
evaluate furcation involvement, dehiscence, fenestration defects, and
periodontal cysts and to assess postsurgical consequences of regenerative
periodontal treatment.

CBCT scan: Periodontal evaluation – Panoramic view (A) showing horizontal bone loss with
furcation involvement. Axial views (B) are useful for evaluating furcation involvements
where as crossections are particularly useful in evaluating buccal and lingual cortical plates
as well as defining endo/ perio lesions. IVR (D) showing periodontal situation, which can
used as tool for educating the patients.

CONE-BEAM COMPUTED
COMPUTED TOMOGRAPHY TOMOGRAPHY
A thin fan-beam of x-rays rotates
around the patients to generate in
one revolution a thin axial slice of
area of interest.
The X-ray source and the Detector
are diametrically positioned and
make a 360 degree rotation around
the patient head within the gantry.
In one 360 degree rotation 360
images are formed.
In one Revolution, one Axial slice
is formed.
A Fan Beam X-Rays are generated.
CBCT scanner generates a cone-
shaped X-ray beam, forms a larger
image.
CBCT can't distinguish properly.
CT Scan offers a greater contrast
resolution or the ability to
distinguish two objects with small
density differences.

COMPUTED TOMOGRAPHY
CONE-BEAM COMPUTED
TOMOGRAPHY
• CT Scan has higher capacity to
separate muscle from fat or
connective tissue.
• Most significant difference; More
amount of Radiation dose is
delivered to patients while scanning.
• It is used for Dental implants,
Neurological Diagnosis, Orthopedics
Diagnosis etc.
• CBCT has limited capacity to
separate muscle from fat or
connective tissue.
• Most significant difference; Very
limited amount of Radiation dose is
delivers to patient while scanning,
effective dose is approximately equal
to full mouth series, this is 50 to 100
times less than the radiation dose
delivers to patient than CT Scan.
• It is mainly used in Dental Implant
purpose.

Optical coherence tomography
• Optical coherence tomography (OCT) was introduced as a biomedical
imaging modality in biological systems in 1991 by Huang et al.
• This non-invasive imaging technique, based on low coherence
interferometry, utilizes coherent near infrared light.
• OCT is capable of obtaining images with a high resolution (5-15 μm) and
penetration depth of 1-2 mm. Moreover, real time 3-D tomographic
images of the tissue can be provided.

• Mota et al analyzed the structure of periodontal tissues in a pig model
using 2 OCT systems operating in the Fourier domain at 930 and 1325 nm
wavelengths. Mota CC, Fernandes LO, Cimoes R, Gomes AS. Non-invasive periodontal probing through Fourier-
domain optical coherence tomography. J Periodontol. 2015;86:1087-1094.
• They showed that it is possible to identify the free gingiva and the attached
gingiva.
• Moreover, the gingival thickness and the gingival sulcus depth were non-
invasively measured.
• The authors suggested that OCT systems operating at 1325 nm wavelength
are of higher performance than systems operating at 930 nm, due to
deeper tissue penetration.

• Fernandes et al employed OCT technology in vivo by measuring gingival sulcus
depths of anterior teeth in periodontally healthy individuals, at 3 buccal sites per
tooth (445 sites in total). Fernandes LO, Mota C, de Melo LSA, da Costa Soares MUS, da Silva Feitosa D, Gomes ASL. In
vivo assessment of periodontal structures and measurement of gingival sulcus with Optical Coherence Tomography: a pilot study.
J Biophotonics. 2017;10:862-869.
• OCT was compared to the North Carolina manual probe (UNC-15) and to the Florida
automated probe.
• Sulcus depth values obtained by OCT were significantly lower than the values
acquired by manual or automated probes.
• On the other hand, OCT, being a non-invasive technique, was superior to manual
probing in that pain and discomfort were obviated.
• The lateral resolution of the OCT image is fixed depending on the optical
characteristic of the focusing beam, whereas the axial resolution is determined by
the properties of the objects through which the light penetrates.
• Thus, periodontal pocket depth measurements by OCT could vary depending on the
position of the periodontal pocket in the image.

Kim et al reported quantitative methods to improve the accuracy of OCT pocket
depth measurement by applying several calculations to determine the
calibration factor and accurate axial resolution. Furthermore, OCT can also be
used in periodontal tissue analysis. Kim S-H, Kang S-R, Park H-J, Kim J-M, Yi W-J, Kim T-I. Improved accuracy
in periodontal pocket depth measurement using optical coherence tomography. J Periodontal Implant Sci. 2017;47:13-19.
Kakizaki et al in a study of human periodontally healthy subjects, showed that
the gingival thickness and biological width can be evaluated by OCT. However,
OCT utilizes light waves that can give much greater scattering in deeper tissue
than subsurface tissues. Thus, this optical imaging modality cannot sustain high
spatial resolution with deep tissue imaging. Kakizaki S, Aoki A, Tsubokawa M, et al. Observation and
determination of periodontal tissue profile using optical coherence tomography. J Periodontal Res. 2017;53:188-199.

Endoscopic capillaroscopy for periodontal
pocket microcirculation Imaging
• Townsend & D’Aiuto showed the feasibility of using fiber-optic probes to
visualize directly the periodontal pocket wall through its microcirculation
and measure the change in number and diameter of blood vessels associated
with periodontal disease. Townsend D, D’Aiuto F. Periodontal capillary imaging in vivo by endoscopic
capillaroscopy. J Med Biol Eng. 2010;30:119-123.

• The core of the system is composed of a fiber-optic image probe of 950 μm,
which is inserted in the gingival sulcus or periodontal pocket.
• Illumination is provided by green 520 nm wavelength light that is absorbed
by both oxygenated and deoxygenated blood.
• Thus, blood vessels containing red blood cells will appear dark against the
green background.
• They concluded that the combination of capillaroscopy and optical fiber
technology could obtain high-resolution imaging of the periodontal pocket
microcirculation.

Photoacoustic imaging
• Photoacoustic (PA) imaging is a hybrid biomedical imaging technology
that combines the high contrast of optical imaging with the high
resolution of ultrasound imaging.
• This imaging modality is based on the PA effect that was first observed
by Alexander G Bell in 1880.

In PA imaging, absorption of optical
energy from either an endogenous
chromophore such as hemoglobin,
melanin or lipids, or exogenous contrast
agents such as organic dyes, gives rise to
thermoplastic expansion and generation
of acoustic (ultrasound) waves.
These ultrasound waves can be detected
and converted to electric signals that are
then processed for imaging.

• PA imaging can provide high-resolution images (as low as 5 μm) with a
deeper imaging depth than optical tomography due to the lower scattering of
ultrasonic waves than light in tissue.
• PA imaging has improved tissue contrast more than ultrasound imaging has.
This is attributed to the rich endogenous and exogenous optical contrasts
achieved by PA while ultrasound imaging is restricted by the mechanical
properties of the tissues.
• Another advantage of PA imaging is the absence of ionizing radiation.
• Moreover, PA imaging has been shown to be faster than magnetic resonance
imaging (MRI).
Advantages

Limitations
• Its penetration is limited to approximately 5 cm in tissue due to optical
attenuation.
• The emitted ultrasound waves are strongly reflected from gas-liquid or
gas-solid interfaces and cannot pass through gas cavities efficiently.
• Moreover, thick bones like human skull have been shown to attenuate
and distort greatly the ultrasound signals.

• Lin et al showed the feasibility of periodontal pocket depth measurements utilizing PA
imaging.
• In this study, they created artificial periodontal pockets with scalpels applied parallel to
the tooth down to alveolar bone in pig jaw models. These artificial pockets were then
loaded with food-grade cuttlefish ink as a contrast medium.
• They compared the pocket depth measurements obtained by PA imaging to those
measured by gold standard periodontal probing method.
• They found that PA imaging could visualize the entire pocket with 0.01 mm precision.
• It was suggested that PA imaging could be a non-invasive diagnostic tool for periodontal
pocket imaging and depth measurements.
• However, evidence for absorption and scattering of optical energy and ultrasound waves
by bone suggests the limitations of this technology, particularly for imaging infrabony
pockets and interproximal pockets.

Magnetic resonance imaging
• MRI is a non-invasive tool for soft tissue diagnosis, without ionizing radiation.
• MRI scanners apply a magnetic field that spins the hydrogen nuclei in water
molecules in the body.
• MRI machines pulse a radiofrequency (RF) that allows nuclear spins to resonate
in the strong static magnetic field.
• The excited hydrogen atoms give off an RF signal, which is received and
measured by a receiving coil that converts the RF signals into an electrical
current signal.
• The tissue contrast depends on the rate at which excited hydrogen atoms return
to relaxation.

• It is not feasible to image teeth by conventional MRI because of their
high mineral content and the rapid decay of the water signal within
dense mineralized tissues. This will result in low MRI image intensity.
• However, conventional MRI can image intraoral soft tissue including,
gingiva, pulp, root canals and periodontal ligament area.
• Thus, MRI may be able to image the soft tissue wall of the periodontal
pocket but not its hard tissue wall.

Subgingival microenvironment has 300+
species
Only few organisms are thought to be involved with
periodontal disease.
Strong evidence for actinomycetemcomitans (Aa),
Porphyromonas gingivalis (Pg), and Tannerella
forsythia (Tf).
Other organisms that are thought to have etiologic
role are Camphylobacter rectus, Eubaterium
nodatum, Fusobacterium nucleatum,
Peptostreptococcus micros, Prevetolla intermedia
and Prevetolla nigrescens, Trepenoma Denticola.

Uses of microbiologic
analysis
Support diagnosis.
Treatment planning
Indicator for disease activity (absence
is a better indicator)

Plaque samples are cultivated under anaerobic
conditions using selective and nonselective
media.
Advantage:
Relative and Absolute count of the cultured species.
Disadvantage:
Strict sampling conditions
Difficulty in culturing most organisms
Low sensitivity : organisms lesser then 103 is difficult
to detect
Time consuming
Expensive equipment and experienced personnel

P.
intermedia
F.
Nucleatum
P.
Forsythia
P. Gingivalis

Direct Microscopy: Most of the
periodontal pathogens are non motile so
it is difficult to identify.
That’s why dark field microscopy seems an
unlikely candidate as a diagnostic test of
destructive periodontal diseases.

Immunofloresence Assay (IFA):
Direct Indirect
DirectIFA: AB conjugated with Fluorescein marker + Bacteria ( Antigen) = Immuno complex
IndirectIFA: Primary AB + Bacteria= Immune Complex+ Secondary Fl conjugated AB

Flow Cytometry
Bacterial cells+ species specic AB + Secondary FL
Conjugated AB Introduced in flowcytometer
Bacterial cells is separated into single cell
suspension- passes through the tube Cells
identified by lasers.

Latex Agglutination Test
 Latex beads coated with species
specific AB when beads come in
contact with specific species in
sample they bind and
agglutination occurs clumping of
beads is visible test positive.
 THIS method is not clinically
available.its olny for research.
 Advantages:
 Simple and Rapid testing
 Higher sensitivity and specificity.

ELISA= Enzyme Linked
Immunosorbent Assay
 Similar AB and Antigen reaction,
but the fluorescence is read
using a photometer.
 Evalusite: commercially available kit
to detect Aa,Pg and Pi.

Well with precoated antibody + Sample to be
tested= immune complex
Specific antigen bind to the antibody + Secondary
antibody added.
Immunofloresence dye bound to secondary antibody
Substrate added which changes the color of the solution
Amount of florescence checked by photometer
(450nm)

SANDWICH ELISA
Specific antibody ->antigen conjugate
(antibody to antigen)->substrate added
->colour (positive result)

. Perioscan is a popular diagnostic
kit uses BANA reaction.
Disadvantage: May be positive in clinically
healthy site
□ Cannot detect disease activity
□ Limited organisms detected
□Other pathogens may be present if it’s
negativE

Perioscan requires a plaque sample to detect
the presence of enzymes capable of
degrading N-benzoyl-DL-arginine-2-
naphthylamide (BANA) from relatively few
anaerobic periodontal pathogens.

Basic Principle: Analysis of DNA,
RNA and protein structure.
Hybridization: Pairing of complimentary
strands of DNA to produce a double stranded
DNA.
Nucleic acid probe: is a known DNA/RNA
which is synthesized artificially and labeled
with a enzyme or a radioisotope for detection
when placed in a plaque sample.

DNA Probe: uses a segment of a single stranded
DNA, labeled with a enzyme of a radio
isotope, that is able to hybridize to a
complimentary nuclei strand, and thus
detect presence of target microorganism.

Two types of DNA
probes
Whole genomic: Targets the whole DNA strand
rather then a specific sequence or gene.
High chances to cross react with non target
microorganism
Lower sensitivity and specificity.

Oligonucleotide probes: target variable region
of
16sRNA or a specific sequence in the
DNA strand.
Higher sensitivity and specificity.

Checkerboard DNA-DNA
Hybridization Technology:
Developed by Socransky et.al.
40 bacterial species can be detected using whole
genomic digoxigenin-labeled DNA probes.
Large number of samples can be tested and upto
40 oral species detected with a single test.

Advantages of DNA probes as compared to bacterial
culturing.
1. More sensitive and specific
2. Requires as less as 104 cells of each species to be
detected.
3. Multiple species detected with a single test
4. Does not require viable bacteria
5. Large number of samples can be assessed.
Disadvantage:
1. Expensive
2. Expert personnel to carry out the test
3. Not easily available

Polymerase Chain Reaction (PCR):
 Involves amplification of a region of
DNA by a primer specific to the
target species.
 If there is amplification then it
indicates the presence of the target
species in the sample.

Advantages:
1. High detection limit. As less as 5- 10cells can
be amplified and detected.
2. Less cross reactivity under optimal conditions
3. Many species can be detected simultaneously
Disadvantage:
1. Small quantity needed for reaction may not
contain the necessary target DNA
2. Plaque may contain enzymes which may
inhibit these reactions.

Asses host response by studying mediators as
a
response to specific bacteria or local release
of inflammatory mediators or enzymes as
response to infection.
Source of samples may be; GCF, Saliva, or
Blood.
GCF is most commonly used, where as saliva is
been recently been researched recently.

Most well studied, with almost 40 components in form of
host-derived enzymes, tissue breakdown products, and
inflammatory mediators.
Collected with paper strips, micro papillary tubes,
micropipettes, microsyringes, plastic strips.
Paper strips commonly used, introduced in sulcus for
30 secs and volume is measured using Periotron 6000.
Periotron measures the capacitance across the wet
paper strip, which is converted to digital reading.
Periotron reading have high correlation with clinical
gingival indices.
Quickest and easiest way to measure GCF.
GCF
:-

Perio Paper Strips
Periotron 8000

It is the next most used after GCF
easily collected
contain both local and systemic derived markers for
periodontal disease
Collected from parotid, sub-mand or sub lingual or as
‘Whole saliva’
Whole saliva contains secretions of major and minor salivary
glands, desquamated cells, and GCF.
No diagnostic test available in the market although lot of
research is in progress.
Markers to look for in saliva: proteins and enzymes from
host, phenotypic markers, host cells, hormones, bacteria,
bacterial products, volatile compounds, and ions.
Saliva:

Cytokines: are substances released
by cells of the immune system.
Cytokines in GCF are: TNF-alpha, IL-1, IL-6, and
IL-8
Have actions on immune cells and release of
enzymes, including bone resorption.
Can be used to determine the disease activity.
Esp. Prostaglandin E in increased in GCF of
periodontitis patients.
Can be used to determine disease activity

Various enzyme are released from the host
cell
during the initiation & progression
of periodontal disease.
 Matrix components may be dissolve
either by extracellular matrix
metalloproteinase dependent or
lysosomal proteinase.
 The protease & enzyme involved
in this process may have use as
diagnostic aids & thus their role

•
•
The breakdown of collagen occurs during
inflammation , remodeling & wound
healing.
Two different pathway,
Intracellular
Extracellular

Under non-pathologic condition,
phagocytosis
& intracellular digestion of collagen fibrils is
a process observed.(in gingiva & pdl)
In pathologic condition,
the balance between synthesis &
degradation is disrupted
The collagen fibrils of PDL are broken down
with supporting alveolar bone.

Different enzymes involved in both the
intracellular & extracellular pathway of
tissue destruction.
Intracellular destruction enzyme:-
❖ Aspartate aminotransferase
❖ Alkaline phosphate
❖ Beta glucoronidase
❖ Elastase
Released from
dead & dying cells
of PDL
Mostly from
PMNs
,neutrophils.

Extracellular destruction
enzyme,
❖ Matrix metalloproteinases- produced by
inflammatory epithelial & connective tissue
cells at affected sites.

AST: derived from dead cells
Elevated in GCF in periodontal disease
Periogard is a commercially available colorimetric
test.

𝗈 Collection of GCF with filterpaper strip
Placed into Tromethamine Hydrochloride buffer
Add L-aspartic & alpha-ketoglutaric acid
10min reaction
time
If AST
present
The aspartate &glutarate catalyzed
to oxalatate & glutamate
Gives red
colour

A potenttial problem with the AST test is its
inability to discriminate between site with
severe inflammation but with no
attechment loss from sites that are losing
attachment.

ALP: released from osteoblast, neutrophils,
fibroblast..
BG and Elastase: found in Neutophils.
BG may have predictive value in patient at higher
risk for losing attachment.
Cathepsins-acidic lysosomal enzyme .
all shown to be higher in diseased sites. May be
used to predict severity of disease or to predict
disease activity.

➢zinc and calcium dependent
enzymes
➢constitutively formed in the body, secreted by
fibroblast and macrophages.
➢Normally help in degrading and remodeling of
extracellular matrices.
➢In chronic periodontitis they cause the
degradation of the collagen fibrils in PDL and
Alveolar bone.
➢MMP,2,3,8 9 and 13 play important role.

➢ MMP8 level is associated with the attachment
loss
➢ In periodontitis patient increased level of MMP-
8.
➢ Level reduces in response to treatment. (Chair
side test kit)
➢ Can be used to indicate present disease status
and predictor of future disease.

One of the major feature of
periodontitis is the destruction
of collagen & extracellular
matrices.
 The connective tissue of periodontium is composed
of fibrous (collagen & elastin)&non fibrous
(glycoproteins),water etc.
 The extracellular matrix is composed of
collagen,proteoglycan & noncollagen protein.
 In periodontitis elevated level of hydroxyproline from
collagen breakdown &glycosaminoglycans from
matrix degradation.

Conclusion:
No marker available to predict the disease activity
as there is no proven correlation between these
markers and the clinical loss of attachment.
In search of tool with high predictive value,
simple, safe and cost effective.

What are diagnostic aids and write about advanced
diagnostic aid
ELISA Test
BANA Reaction
Role of Saliva
Advanced Diagnostic Techniques
Periodontal Probes
Role of Saliva in Oral Health
GCF
MMP
B-Glucoronidase
AST
Enumerate Adv Diagnostic Aids and Elaborate on
DNA Probe/ Microbial Analysis/

advanced diagnostic aids in periodontics

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