Advanced Periodontal Diagnostic Techniques(As an adjunct to conventional techniques)

Content of discussion

Introduction

Limitations of conventional periodontal diagnosis

Advances in Clinical diagnosis

Advances in Radiographic assessment

Advances in Microbiologic analysis

Advances in characterizing the Host response

Conclusion and future scopes

Introduction

Definition of DIAGNOSIS (Dx)* : Diagnosis is defined as; correct determination, discriminative estimation and logical appraisal of conditions found during examination as evidenced by distinctive marks, signs and characteristics of diseases

*Glossary of Periodontal terms

Recognizing a departure from health in the periodontium

and distinguishing one disease, disease categorization, or

etiology from another, Based on information obtained

from the medical and dental histories, clinical and

radiographic examination of the patient, and laboratory

findings.

Periodontal Dx*:

Purposes of Periodontal diagnostic procedures

According to Armitage GC(1996) Periodontal diagnostic procedures

can potentially serve 5 separate, but related purposes:

1. Screening

2. Diagnosis of specific periodontal disease

3. Identification of sites or subjects at an increased risk of experiencing the progression of periodontal destruction

4. Treatment planning and

5. Monitoring of therapy.

Current conventional techniques

Clinical diagnosis is made by measuring either clinical attachment loss (CAL) or radiographically by loss of alveolar bone

This kind of evaluation identify and quantify current clinical signs of inflammation ,

Provides historical evidence of damage with its extent and severity

Limitations !!!

Does not provide cause of the condition

No info. on patient’s susceptibility to the disease

Cannot identify sites with ongoing periodontal destruction or sites in remission

Cannot differentiate whether response to therapy is positive or negative

Periodontal disease is localized and multifactorial

Periodontal pathogens

Host response

behaviouralsystemic

Genetic

Advanced periodontal diagnostic techniques

Advances in Clinical diagnosis

Advances in Radiographic Assessment

Advances in Microbiologic Analysis

Advances in Characterizing the Host Response

Advances in Clinical Diagnosis

1. Gingival temperature

Kung et al (1990) claim that thermal probes are sensitive diagnostic

devices for measuring early inflammatory changes in gingival

tissue.

Subgingival temperature at diseased sites is increased as compared

to normal healthy sites

Commercially available system PerioTemp probe enables the

calculation of temperature differential (with sensitivity of 0.10C)

between the probed pocket and subgingival temperature

Possible explanation for ↑ temperature with increasing

probing depth is an increase in cellular and molecular activity

caused by increased periodontal inflammation

Haffajee et al. (1992): found that elevated subgingival site

temperature is related to attachment loss in shallow pockets

and elevated proportions of Pg, Pi, Tf, Aa.

2. Periodontal probing

Most widely used diagnostic tool

Probing depth is measured from the free

gingival margin to the depth of the

probeble crevice.

Longitudinal measurement of CAL or

probing depth is a ‘gold standard’ for

recording changes in periodontal status

Limitation of conventional probing

Lack of sensitivity and reproducibility.

Disparity between measurement depends on:

probing technique, probing force, angle of insertion of probe, size

of probe, precision of calibration, presence of inflammation.

Readings of clinical pocket depth measured with probe does not

coinside with the histologic pocket depth.

All these variable contribute to the large standard deviations (0.5-

1.3 mm) in clinical probing results

Classification of periodontal probesdepending on generation

1.First generation probes: (conventional probes)

Conventional manual probes that do not control

probing force or pressure and that are not suited for

automatic data collection.

example: Williams periodontal probe CPITN probe UNC-15 probe Goldman Fox probe

2.Second generation probes: (Constant force probes)

Study done by Tupta et. Al (1994) has shown that

force to probe pocket: 30g

force to probe osseous defect: 50g

Introduction of constant force or pressure sensitive probes allowed for improved standardization of probing.

e.g.: Pressure sensitive probe

Constant pressure probe

Limitation: data readout and storage is inaccurate.

3.Third generation probe:(Automated probes)

• Computer assisted direct data capture was an important step in reducing examiner bias and also allowed for generation of probe precision. (according to NIDCR criteria)

e.g.: Toronto probe Florida probe, Interprobe, Foster Miller probe.

Florida probe

Tip is 0.4mm

Sleeve- edge provides reference to make measurements

Coil Spring; provides constant probing force

Computer for data storage.

FP Handpiece tip as it enters the sulcus Handpiece tip with constant force in use (tip at bottom of sulcus) and sleeve properly positioned at the top of the gingival margin allowing the computer to measure the difference.

Clark and Yang (1992): trained operators and performing the ‘double pass’

method, the measurements taken with Florida probe system shows lower

standard deviation than those obtained with conventional probing.

Mean Standard Deviation for CAL of about 0.3mm, which is superior to an

average of 0.82mm reported by Haffajee et al. For conventional probing.

Limitations

Lack of tactile sensitivity

Fixed probing force

Underestimation of deep periodontal pockets.

4.Fourth generation probes: (Three dimensional probes)

• Currently under development, 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.

5.Fifth generation probe: (3D + Noninvasive)

• Basically these will add an ultrasound to a fourth generation probes.

• If the fourth generation can be made, it will aim in addition to identify the attachment level without penetrating it.

• e.g.: Ultra sonographic probe.

Advances inRadiographic Assessment

Dental Radiographs are traditional method to assess destruction of

alveolar bone.

“Conventional radiographs are very specific but lack sensitivity”

Primary criterion for bone loss is the distance from CEJ to the alveolar

crest and distance more than 2 mm is considered as the bone loss.

But variability affecting conventional radiographic technique are,

Variation in projection geometry

Variation in contrast and density

Masking by other anatomic structures.

1. Digital radiography

Capturing radiographic image using a sensor

The first direct digital imaging system, RadioVisioGraphy (RVG), was

invented by Dr. Frances Mouyens.

Advantages

1. Elimination of chemical processing

2. Increased efficiency and speed of viewing

3. Diagnostic information can be enhanced

4. Computerized storage of radiographs

5. Reduced exposure to the radiation

2. Subtraction radiography

Subtraction radiography was introduced to dentistry in 1980 by Ruttimann, Webber et & Grondahl HG

This is a technique by which images not of diagnostic value in a radiograph, are eliminated so that changes in the radiograph can be precisely detected

Serial radiographs

converted to digital images

superimposed composite image

Quantitative changes

This technique requires a paralleling technique to obtain a standardize geometry and accurate superimposable radiographs

This technique facilitates both quantitative and qualitative visualization of even minor density changes in the bone

Bone gain appears as light areas and bone loss appears as dark areas

Rethman et al.(1985): increased detectability of small osseous lesions by substraction method compared with conventional radiography

Recent image subtraction:“diagnostic subtraction radiography” (DSR)

Modification

Use of a positioning device during film exposure

Image analysis software system applies an algorithm to correct angular

alignment discrepancies.

3. Computer Assisted Densitometric Image Analysis (CADIA)

Video camera measures the light transmitted through radiograph

and the signals form the camera is converted to gray scale image.

Advantage:

Measures quantitative changes in bone density longitudinally.

Higher sensitivity, reproducibility and accuracy as compared to

DSR.

4. Computed tomography (CT)

In 1972, Godfrey Hounsfield announced the invention of a

revolutionary imaging technique, which he referred to as

“computerized axial transverse scanning”

Fan shaped X-ray source is used

The computed tomographic image is reconstructed by computer, which

mathematically manipulates data obtained from multiple projections.

Computed tomography is a specialized radiographic technique that

allows visualization of planes or slices of interest

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.

Advantages over conventional radiography

Axial CT

Coronal CT

Sagittal CT

Multiplaner imaging

Application of CT Used when accurate information regarding the topography of

osseous structure is needed

Soft tissue contour and dimension

To check continuity and density of the cortical plates

vertical height of the residual alveolar ridges

density of the medullary space and basilar bone

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.

Disadvantages of Computed Tomography

specialized equipment and setting.

Radiologists and Technicians need to be knowledgeable of the

anatomy, anatomic variants and pathology of the jaws

higher radiation

Metallic Restorations can cause ring artifacts that impair the

diagnostic quality of the image

Cone-beam Computed Tomography

Routine use of CT in dentistry is not accepted due to its cost, excessive radiation, and general practicality.

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.

Advantages

It gives complete 3D reconstruction

CBCT units reconstruct the projection data to provide interrelational images in three orthogonal planes (axial, sagittal, and coronal).

Its beam collimation enables limitation of X-radiation to the area of interest.

Patient radiation dose is five times lower than normal CT, as the exposure time is approximately 18 seconds, that is, one-seventh the amount compared with the conventional medical CT.

Reduced image artefacts

Evaluation of the jaw bones which includes the following:

Bony and soft tissue lesions

Periodontal assessment

Soft tissue CBCT for the measurement of gingival tissue and the dimensions

of the dentogingival unit

alveolar bone density measurement

Temporomandibular joint evaluation and

Implant placement and evaluation

Whenever there is need for 3D reconstructions

Indications of CBCT

Advances In Microbiologic Analysis

Uses of microbiologic analysis

1. support diagnosis of various Periodontal disease

2. Can tell about initiation & progression

3. To determine which periodontal sites are at high risk for active destruction

4. Can also be used to monitor Periodontal therapy

Advances In Microbiologic Analysis includes:

1. Immunohistodiagnostic methods

2. Enzymatic methods

3. Molecular biology techniques

Neucleic acid probes

Checkerboard DNA-DNA hybridization

PCR

Sample collection

It is a common need of all the microbiologic analysis to

collect an appropriate subgingival plaque sample

Mombelli et al. (2002) have shown that four individual

subgingival specimens, each from the deepest periodontal

pocket in each quadrant, should be pooled to be able to

detect the highest amount of pathogens.

Transport the specimen in a anaerobic environment

“Immunodiagnostic methods”

Immunological assays use fluorescent conjugated antibodies that recognize specific bacterial antigens, and the identification of these specific antigen-antibody reactions allows the detection of target microorganisms.

This reaction can be visualized using a variety of techniques and reactions:

1. Direct (DFA) and indirect (IFA) immunofluorescent assays

2. Flow cytometry

3. Enzyme-linked immunosorbent assay (ELISA)

4. Latex agglutination

1. Immunofluorescent assays

IFA is used mainly to detect A.a and P.g

Zambon et. al (1986) showed that IFA is comparable to

bacterial culture in its ability to identify these pathogens

Zambon et. al (1995) sensitivity of these assays ranges from

82%-100% for A.a. and 91%-100% for P.g

Specificity values of 88%-92% and 87%-89% respectively

2. Flow cytometry

Rapid identification

Principle is labelling bacterial cells with both species-specific

antibody and a second fluorescein-conjugated antibody

This suspension is introduced into flowcytometer, which

separates bacterial cells into an almost single cell suspension

Limitation is sophistication and cost involved with this

procedure

3. ELISA

ELISA has been used primarily to detect serum antibodies

to periodontal pathogens.

In research studies to quantify specific pathogens in

subgingival samples

A novel chair side ELISA commercially known as

“Evalusite” has been marketed in Europe and Canada for

the chair side detection of 3 periodontal pathogens. Aa,

Pg and Pi

4. Latex agglutination

Latex beads coated with species specifi c AB

when beads come in contact with specifi c species in sample they bind and aggluti nati on occurs

clumping of beads is visible, usually in 2-5 mins.

Test +ve

merits

•Quantitative estimate of target

species

•Not requiring stringent sampling and

transport methodology

•Higher sensitivity and specificity

than bacterial culturing for A.a, P.g

and T.f.

demerits

•Limited to the number of

antibodies tested

•Not amenable for antibiotic

susceptibility

•Lack the evidence of well-

controlled clinical studies

“Enzymatic Methods”

Tf, Pg, Td, and Capnocytophaga species share common enzymatic

profile- a trypsin like enzyme.

N-benzoyl-d L-arginine-2-naphthylamide

Trypsin like enzyme BANA hydrolysis

β-naphthylamide (chromophore)

PERIOSCAN uses this reaction for the identification of this bacterial

profile in plaque isolates

Loesh et al. (1986) detection of these pariodontal pathogens by BANA

reaction serves as a marker of disease activity

He also showed that shallow pockets exhibited 10% positive BANA

reaction, whereas deep pockets (7mm) exhibited 80%-90% +ve BANA

reaction

Beck et al. (1995) used BANA test as a risk indicator for periodontal

attachment loss

Disadvantage of BANA

May be positive in clinically healthy site

Can not detect sites undergoing periodontal destruction

Limited organisms detected

So that, negative results does not rule out the presence

of other important periodontal pathogens.

“Molecular Biology Techniques”

The principles of molecular biology technique reside in the

analysis of DNA, RNA and the structure and function of proteins

Diagnostic assays require specific DNA fragment that recognize

complementary-specific DNA sequences from target

microorganisms

This technology requires bacterial DNA extracted from the plaque

sample and amplification of the specific DNA sequence of the

target pathogen

1. Nucleic acid probes

A probe is a known, single stranded

nucleic acid molecule (DNA or RNA)

from a specific pathogen

synthesized and labeled with a

enzyme of a radio isotope

Hybridization: Pairing of

complimentary strands of DNA to

produce a double stranded DNA.

Probe DNA

B.DNA

Hybridization

DMDx and Omnigene are commercially available genomic probes

for the detection of Aa, Pg, Pi and Td.

Van Steenberghe et al. (1999) reported a sensitivity of 96% and

specificity of 86% for Aa., and 60% and 82% respectively for Pg in

pure lab isolates.

In clinical specimens, both sensitivity and specificity were reduced

significantly, suggestive of cross reactivity with non target bacteria

in plaque sample because of the presence of homologues

sequences between different bacterial species

2. Checkerboard DNA-DNA hybridization technology

Developed by Socransky et.al in 1994

40 bacterial species can be detected using whole genomic digoxigenin-labeled DNA probes.

Applicable for epidemiologic research and ecological studies

3. Polymerase chain reaction (PCR)

Repeated cycles of oligonucleotide (primer)–directed DNA

synthesis of “target sequences” are carried out in vitro.

The PCR method is considered the fastest and most

sensitive method available for detecting the presence of

bacterial DNA sequences

A modification of the original PCR technology, "real-time"

PCR, permits not only detection of specific

microorganisms in plaque, but also its quantification.

Advantages

1. High detection limit. As less as 5- 10 cells can be amplified and

detected.

2. Less cross reactivity under optimal conditions

3. Many species can be detected simultaneously

Disadvantage

4. Small quantity needed for reaction may not contain the necessary

target DNA

5. Plaque may contain enzymes which may inhibit these reactions.

Advances in characterizing

the Host response

Assessment of host response refers to the study of mediators by

immunologic or biochemical methods, that are recognized as a part of

individual’s response to the periodontal infection.

Mediators

1. specific Mediator

antibody to a putative pathogen

2. less specific reaction

the local release of the inflammatory mediators, host derived

enzymes and tissue breakdown products

For that...

Diagnostic tests have been developed that add measures of the

inflammatory process to conventional clinical measures.

Sources of the sample are:

GCF, gingival crevicular cells, Saliva, Blood serum, blood cells and

rarely urine.

Most efforts to date have been based on use of components of GCF

and to a lesser extent, saliva and blood

Assessment of Host response

Inflammatory mediators and products

Host derived enzymes

Tissue breakdown products

1. Inflammatory mediators and products

Cytokines present in GCF and investigated as potential diagnostic markers are: TNF α IL-1 α IL-1β IL-6 IL-8

PGE2 (product of COX pathway)

Cross sectional studies have shown Good correlation with disease status and severity but not disease progression

In cases of untreated periodontitis concentration of PGE2 was found increased (showing active Periodontal destruction)

Host derived enzymes

Breakdown of collagen occours by two different pathways:

Intracellular

1. Aspartate amino transferases

2. Alkaline phosphatase

3. β- Glucuronidase

4. Elastase

Extracellular

Matrix metalloproteinase's family (MMPs)

Tissue Breakdown Products

Analysis of GCF obtained from sites with active

periodontitis clearly shows elevated levels of

Hydroxyproline from collagen breakdown and GAGs from

matrix degradation

Osteocalcin and type-1 collagen peptides- progression of

alveolar bone loss

Conclusion

1. This discussion directly translates into improved periodontal

therapy by offering the clinician, the radiographic & laboratory

measure of periodontal infection as an adjunct to traditional

clinical indices of periodontal disease.

2. Future application of advanced diagnostic techniques will be of

value in documenting disease activity and treatment options

3. But, despite excellent progress in diagnostic

methodology,conventional efforts evaluating inflammation and

past evidence of tissue breakdown remain the standard for

disease evaluation

4. There is still a lack of a proven “gold standard” of disease

progression

5. After all these years of intensive research, we still lack a proven

diagnostic test that has demonstrated high predictive value for

disease progression, has a proven impact on disease incidence and

prevalence, and is safe, and cost-effective.

A tremendous amount of research is still required to

explore the role of advancements in diagnostic aids as

a possible medium for the future prediction and

prevention of periodontal disease.

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References:

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11. Page RC: Host response tests for diagnosing periodontal diseases. J Periodontol 1992; 63:356.

12. Papanou PN, Neiderud AM, Papadimitriou A, et al: Checkerboard assessments of Periodontal Microbiota and serum antibody responses: A case control study. J Periodontol 2000; 71:885.

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14. Tupta-Veselicky L, Famili P, Ceravolo FJ et al: A clinical study of an electronic constant force periodontal probe. J Periodontol 1994; 65:616.

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