This document discusses various tests used to assess caries activity and susceptibility. It defines caries activity as referring to the speed of progression of caries lesions, while caries susceptibility refers to the inherent tendency to develop caries. Several tests are described that examine levels of cariogenic bacteria like lactobacilli or Streptococcus mutans in saliva, or that measure the ability of saliva to neutralize acids and buffer pH changes. These include bacterial culturing tests, color change assays, and tests analyzing the dissolution of enamel. Each test is outlined with the purpose, equipment, procedure, results, advantages and limitations.

1. CARIES ACTIVITY TESTS
Dr. Sucheta Prabhu
Second year MDS
29/06/2017

2. Questions asked previously
Short essays
1.Snyders test
2.Caries activity indicators
3.Caries succeptibility
4.Application of cariogenecity tests in prevention of dental caries

3. Questions asked previously
• Long Essays
• 1.Define caries activity and caries succeptibility tests.Critically
evaluate various caries activity and succeptibility tests and their
clinical implications.

4. CONTENTS
• INTRODUCTION
• DEFINITIONS
• NEED FOR CARIES ACTIVITY TESTS
• IDEAL REQUISITES
• CLASSIFICATION
• CARIES ACTIVITY TESTS
• LIMITATIONS OF CARIES ACTIVITY TESTS
• CONCLUSION
• REFERENCES

5. INTRODUCTION
Carious lesions are
detected by a clinical
examination coupled with
bite-wing radiographs .
A clinical examination
neither predicts caries
activity nor indicates a
patient’s susceptibility to
dental caries.
A simple reliable
laboratory test that could
do this would greatly
facilitate the clinical
management of patients.

6. DEFINITIONS
• Refers to the increment of active lesions (new or
recurrent lesions) over a stated period of time.
It is a measure of the speed of progression of a
caries lesion.
CARIES ACTIVITY
• Refers to the inherent tendency of the host &
target tissue, to be afflicted by the caries
process.
CARIES
SUSCEPTIBILITY

7. CARIES ACTIVITY TEST
Are defined as tests that estimate the actual state of disease
activity(progression/regression).

8. PURPOSE OF CARIES ACTIVITY TESTS
1.Identify high risk groups .
2.Determine the need for personalized preventive measures
3.Monitor the effectiveness of oral health education programs.
4. Motivate the individual.

9. IDEAL REQUISITES OF A CARIES ACTIVITY
TEST
• Snyder,1951
• Should have a sound theoretical basis.
• Simple
• Easy to perform
• Inexpensive
• Time for test and result should be small.
• Should be adaptable to the chairside.
• Results should be accurate and reproducible.
• Tests should have maximum correlation with clinical status.

10. CLASSIFICATION
MICROBIAL
TESTS EVALUATING
SALIVARY DEFENSE

11. CLASSIFICATION
MICROBIAL
TESTS
Lactobacillus
count test
Dentocult LB
Snyder test Alban’s test
Swab test
Mutans group
of streptococci
screening tests

12. CLASSIFICATION
Mutans group
of streptococci
screening tests
Tooth pick
method
Tongue
blade
method
S .mutans
dipslide
method

13. CLASSIFICATION
EVALUATING
SALIVARY
DEFENSE
Salivary
Reductase
test
Dentobuff
test
Fosdick
calcium
dissolution
test
Caries
reactivity
tests
Cario stat
test
Dewar test
Oratest

14. LACTOBACILLUS COLONY COUNT TEST
• Hadley (1933)
• BASIS
• Tomato peptone agar is the selective medium favoring
the growth of aciduric lactobacilli.
• PRINCIPLE:
• The number of acidogenic and aciduric bacteria in the
patient’s saliva are estimated by counting the number
of colonies appearing on tomato peptone agar after
inoculation with a sample of saliva.

15. LACTOBACILLUS COLONY COUNT TEST
• EQUIPMENT:
• Saliva collecting bottles
• Paraffin
• Two 9ml tubes of saline
• Two agar plates
• Two bent glass rods
• Incubator
• Quebec counter
• Pipettes.

16. PROCEDURE
Saliva obtained by chewing on a piece of paraffin
Shaken with glass rods to break up aggregates of bacteria
Saliva +buffer solution and 1ml of the dilutions 10-2 and 10-3 is mixed
with 10ml melted agar
Plates are incubated for 4 days
Lactobacilli appear as whitish dots on the medium ; counted for the number
of colonies

17. LACTOBACILLUS COLONY COUNT TEST
Simple.
Useful as a screening test
for caries activity in large
groups
ADVANTAGES Inaccurate for predicting
onset of caries.
It takes only few minutes
to do the test, but the
results are not available
for several days.
The counting of colonies is
a very tedious process.
DISADVANTAGES

18. RESULTS:
Number of lactobacilli per ml of
saliva
Caries activity
0-1000 Little or none
1000-5000 Slight
5000-10,000 Moderate
>10,000 Marked

19. Dentoccult LB (Orion Diagnostica , Finland)
Larmas (1975)
• Highly practical and greatly simplified method of
estimating lactobacilli.
• Self-contained kit with a shelf life of at least 1
year.

20. • DENTOCULT ® LB A kit which includes
• Paraffin tablets.
• Dip-slide which on each side has a selective agar for lactobacilli.
• An evaluation chart which shows numbers of lactobacilli per ml
saliva.
• A cup or tube
• A funnel
• An incubator

21. PROCEDURE

22. PROCEDURE
The person chews a piece of paraffin for at least one minute. The saliva is
then spat out in a cup or tube.
The collected saliva is poured over both sides of the slide, the excess is
let to drip off. Insert the slide into its plastic tube, and tighten it.
Aerobic incubation for 4 days at 370c
Number of lactobacilli is estimated by comparing the slides with a model
chart supplied by manufacturer.

23. RESULTS
• For assessment of the level of lactobacilli : Colony Forming Unit (CFU)
per ml of saliva.
•High risk valueMore than
10 5 CFU /ml
•Low risk valueLess than
10 4 CFU /ml

24. COLORIMETRIC SYNDER TEST
•Snyder 1951
• PRINCIPLE
• It measures the ability of salivary
microorganisms to form organic acids
from a carbohydrate medium.

25. COLORIMETRIC SYNDER TEST
• The medium contains an indicator dye,
Bromocresol green.
• This dye changes color from green to
yellow in the range of pH 5.4 to 3.8.

26. COLORIMETRIC SYNDER TEST
• EQUIPMENT:
• Saliva collecting bottles
• Paraffin
• Tube containing Snyder glucose agar+
Bromocresol green adjusted to a pH 4.7-5
• Pipettes
• Incubator

27. PROCEDURE
Saliva is collected before breakfast by chewing paraffin
A tube of Snyder glucose agar is melted and then cooled to 500c
Saliva specimen is shaken vigorously for three minutes.
0.2ml of saliva is pipetted into the tube + mixed by rotating tube
Agar is allowed to solidify in the tube and incubated at 370c
Color change of indicator observed in 24,48,72 hours .

28. RESULTS:
24 hours 48 hours 72 hours
Color Yellow Yellow Yellow
Caries Activity Marked Definite Limited
Color Green Green Green
Caries Activity Continue to
incubate
Continue to
incubate
Caries inactive

29. COLORIMETRIC SYNDER TEST
• ADVANTAGES:
1. Simple
2. Inexpensive
• DISADVANTAGES:
1. Time consuming
2. Color changes are not very clear

30. ALBAN TEST
• Simplified substitute for Snyder test.
• PRINCIPLE:
- Measures the ability of microorganisms to form acids from a medium.
- Alban’s method uses less agar in the media.
• Advantages are that it is simple , cost effective and can act as a
motivational tool for patient.
• Color change from blue to yellow is indicative of caries activity.

31. PROCEDURE
Snyder glucose agar suspension (melted) is distributed using about 5ml per tube.
Tubes autoclaved for 15 minutes, allowed to cool in a refrigerator.
Patient is asked to expectorate saliva directly into 2 tubes of Alban medium
Tubes are incubated at 98.60F for 4 days
Tubes observed daily for color change from blue-green to definite yellow with pH
decrease

32. RESULTS
Color Change Score
No color change 3/4
Beginning color change +
One half color change ++
Three fourths color change +++
Total color change to yellow ++++

33. THE SWAB TEST
• Grainger et al 1965
• PRINCIPLE:
• It measures the ability of salivary microorganisms to form
organic acids from a carbohydrate medium
• PROCEDURE:
• Swab the buccal surface of the teeth with cotton
applicator.
• And it is subsequently incubated in the medium for 48
hours.

34. THE SWAB TEST
• ADVANTAGES
• It predicts caries increments in children with
low or no previous caries experience.
• No collection of saliva is required.

35. RESULTS
pH Caries Activity
≤ 4.1 Marked caries
activity
4.2 to 4.4 Active
4.5 to 4.6 Slightly active
4.6 and over Caries inactive

36. SALIVARY REDUCTASE TEST
• PRINCIPLE:
• Rapp claims, the test measures, ‘ the activity of the
reductase enzyme ’ present in the salivary bacteria.
• The test measures the rate at which an indicator
molecule, Diazo-resorcinol , changes from blue to red to
colorless.

37. SALIVARY REDUCTASE TEST
• EQUIPMENT
• A kit is available under the trade name “ TREA TEX ” that includes:
1.Calibrated saliva collection tubes with the reagent on the inside of
tube’s cap.
2.Flavored paraffin.

38. PROCEDURE
Saliva obtained by chewing on a piece of paraffin
The saliva is then spat out in a cup or tube.
When the saliva reaches the calibration mark(5ml),reagent cap is
replaced
Sample is mixed with a fixed amount of diazoresourcinol
Change in color after 30 seconds and after 15 minutes is taken as a measure
for caries activity

39. RESULTS
Color Time Score Caries Activity
Blue 15 minutes 1 Non-conducive
Orchid 15 minutes 2 Slightly conducive
Red 15 minutes 3 Moderately
conducive
Red Immediate 4 Highly conducive
White Immediate 5 Extremely
conducive

40. SALIVARY REDUCTASE TEST
No incubation is
required
Quick results &
good correlation of
the results with
clinical caries
experience.
ADVANTAGES
Tests results vary
with time after
food intake & after
brushing.
DISADVANTAGES

41. DENTOBUFF TEST OR SALIVARY BUFFER
CAPACITY TEST
• PRINCIPLE
• The test measures the number of milliliters of
acid required to lower the pH of saliva, from pH
7.0 to 6.0.

42. DENTOBUFF TEST OR SALIVARY BUFFER
CAPACITY TEST
• EQUIPMENT
• pH meter
• Titration equipment
• 0.05N lactic acid
• 0.05N base
• Paraffin
• Sterile glass jars

43. PROCEDURE
5ml of stimulated saliva collected under oil atleast 1 hour after eating
pH meter corrected to room temperature
pH of saliva adjusted to 7.0 by addition of lactic acid and base
Lactic acid added to sample till pH of 6.0 is reached
Number of mililitres of lactic acid required to reduce pH from 7.0 to 6.0 is a
measure of buffer capacity

44. EVALUATION
• There is an inverse relationship between the buffering capacity of
saliva and caries activity.
• The saliva of individuals whose mouths contain a considerable
number of carious lesions frequently have a lower acid buffering
capacity than the saliva of those who are caries free .
• ADVANTAGE- Simple to carry out.
• DISADVANTAGE- Does not correlate adequately with caries activity.

45. DENTOBUFF STRIP TEST
A droplet of collected saliva mixture is
deposited on the test paper covering the
area for 5 min.
The color change is checked
The strip degrees are determined by
comparing it against the standard color
chart.

46. DENTOBUFF STRIP TEST

47. Scoring
• DEGREE 1, yellow - pH ≤ 4
• DEGREE 2, green- pH 4.5-5.5
• DEGREE 3, deep blue- pH ≥ 6.0
• The higher the grade the stronger the saliva buffering capacity

48. FOSDICK CALCIUM DISSOLUTION TEST
• PRINCIPLE-
• Test measures the milligrams of
powdered enamel dissolved in 4 hours
by acid formed when the patient’s
saliva is mixed with glucose and
powdered enamel.

49. FOSDICK CALCIUM DISSOLUTION TEST
• EQUIPMENT-
• Powdered human enamel
• Saliva collection bottles
• Sterile test tubes
• Equipment for determining the calcium content
of the saliva.

50. PROCEDURE
The person chews a piece of paraffin for at least one minute. The
saliva is then spat out in a cup or tube. 5% glucose is added.
Saliva is collected and analysed for calcium content
Saliva placed in an 8 inch sterile tube with 0.1 gram of
powdered human enamel
Tube sealed and shaken for 4 hours at body temperature and
again analysed for calcium content

51. FOSDICK CALCIUM DISSOLUTION TEST
The correlation
reported is good
ADVANTAGES
The time required is 4
hours.
Test is not simple,
requires complex
equipment. Is
expensive & requires
trained personnel
DISADVANTAGES

52. DEWAR TEST
• This test is similar to the Fosdick Calcium Dissolution test except that
the final pH after 4 hours is measured instead of the amount of
calcium dissolved. This procedure is not commonly used as it has not
been adequately tested for clinical correlation.

53. STREPTOCOCCUS MUTANS SCREENING TEST
1. PLAQUE/TOOTH PICK METHOD
Kristofferson and Bratthall
The test involves a simple screening of a diluted plaque sample streaked on
a selective culture media.

54. STREPTOCOCCUS MUTANS SCREENING TEST
EQUIPMENT
Sterile toothpicks
Sterile Ringer’s solution
Platinum loop
Mitis Salivarius Agar (MSA) plates containing sulphadimetine
Incubator

55. PROCEDURE
Plaque samples collected from buccal surfaces at gingival third
Placed in Ringers solution , sample shaken till homogenised
Plaque suspension streaked on MSA plates
Aerobic incubation at 37 0 C for 72 hours
Cultures examined and colonies recorded for 10 fields.

56. STREPTOCOCCUS MUTANS SCREENING TEST
2. TONGUE BLADE METHOD
• The test estimates the numbers of S.mutans
mixed in paraffin-stimulated saliva when
cultured in Mitis Salivarius Bacitracin (MSB)
agar.

57. STREPTOCOCCUS MUTANS SCREENING TEST
• EQUIPMENT
• Paraffin wax
• Sterile tongue blades (wooden spatula)
• Disposable Contact Petri Dish (RODAC)
containing MSB agar
• Incubator

58. PROCEDURE
The person chews a piece of paraffin for at least one minute.
This displaces microorganisms increasing their proportion in saliva
Sterile tongue blades rotated in patients mouth about 10 times.
Pressed on a disposable petridish containing MSB agar
Incubation done at 370C

59. EVALUATION
• Counts of more than 100 colony forming units
by this method are proportional to greater than
10 colony forming units of S.Mutans per ml of
saliva by conventional methods.
• ADVANTAGES
Simple and practical for field studies.
Requires no transport media/dilution steps.

60. STREPTOCOCCUS MUTANS SCREENING TEST
3. DIP-SLIDE METHOD
• This method is devised for S.mutans levels in saliva.
• EQUIPMENT
• Paraffin wax
• Plastic slides coated with Mitis Salivarius Agar
• Buffered diluent

61. PROCEDURE
The person chews a piece of paraffin for at least one minute.
Stimulated saliva thus collected is poured on a special plastic slide containing MSA
Agar surface is thoroughly moistened and excess saliva is allowed to drip off
Two disks containing 5µg of Bacitracin are placed on agar 20mm apart
Slide is tightly screwed into a cover tube after inserting CO2 tablet.
Incubation at 37 0 C for 48 hours in a sealed candle jar.

62. EVALUATION
• Score 1 = Low. Colonies are discrete. CFU less than 200.
• Score 2 = Medium. Colonies are less discrete. CFU more than 200.
• Score 3 = High. Colonies are tiny & uncontrollable.

63. STREPTOCOCCUS MUTANS SCREENING TEST
• 4.ADHERENCE METHOD
• Categorizes salivary samples based on ability of S.mutans to adhere to
glass surfaces when grown in sucrose-containing broth.
• EQUIPMENT
• Tube to collect saliva
• Rack to hold culture tubes
• Disposable pipettes
• Incubator
• MSB broth

64. PROCEDURE
Unstimulated saliva (0.1ml) inoculated in MSB
broth
Innoculated tubes set at 60 degree angle and
incubated aerobically at 370C for 24 hours
After growth has been observed , supernatant
medium is removed and cells adhering to the
glass surface are examined microscopically and
scored

65. CARIOSTAT TEST
• It was formulated by Prof. Tsutomu Shimono
(1974)
• PRINCIPLE
• Designed to measure the pH decrease caused
by Streptococcus mutans in the plaque sample.

66. PROCEDURE
Medium used is semi synthetic liquid indicators like
Bromocresol Green & Bromocresol purple
Plaque samples inoculated on cariostat medium &
incubated at 37 ° C for 48 hours
Enables some acid production & acid tolerating bacteria to
survive in medium.
So the ability of acid production can be measured

67. SCORING IN CARIOSTAT TEST
pH COLOR CARIES ACTIVITY
0 6.1 ± 0.3 BLUE CARIES INACTIVE
1 .54 ± 0.3 GREEN SLIGHT CARIES
2 .47 ± 0.3 YELLOW GREEN MODERATE
CARIES
3. 4 ± 0.3 YELLOW MARKED CARIES

68. CARIES RISK TEST
• Allows estimation of a number of
cariogenic bacteria in patient’s saliva
Caries risk test
bacteria
• Determines buffer capacity of salivaCaries risk test
buffer
Recently caries reactivity tests have been advocated as new, quick
& effective.
Caries reactivity tests has 2 components

69. PROCEDURE:
Is a two-in-one dip-in-slide test which identifies counts of
S.mutans or lactobacillus.
Stimulated saliva is collected & applied to both sides of dip
in slide
Incubated for 48 hrs at 37 °C.
Are available in strip form which changes color.

70. ORA TEST
• Rosenberg et al 1989
PRINCIPLE:
Is based on the rate of oxygen depletion by microorganisms in
expectorated milk.
In normal conditions , bacterial enzyme , aerobic dehydrogenase
transfers electrons and protons to oxygen.
Once oxygen gets utilized by aerobic organisms, methylene blue acts as
an electron acceptor and gets reduced to leucomethylene blue.
This reflects the metabolic activity of aerobic organisms.

71. ORA TEST
• Estimates oral microbial levels.
• EQUIPMENT:
• Sterile beakers
• Sterilised milk
• 0.1% aqueous solution methylene blue
• 10ml disposable syringes
• Pipette
• Stopwatch
• Mirror

72. PROCEDURE:
Mouth is rinsed vigorously with 10 ml sterile milk for 30 seconds and expectorate
collected
3 ml of this is transferred to a screw cap tube with the help of a disposable
syringe
0.12 ml of 0.1% methylene blue is added, thoroughly mixed and placed on a
stand in a well illuminated area
Tubes are observed every 10 minutes for any color change at the bottom
using a mirror
Time taken for color change within 6mm ring is recorded

73. ORA TEST
• EVALUATION:
• Higher the infection , lesser was time taken for color change of
expectorate reflecting higher oral microbial levels.
• ADVANTAGE:
• Less time consuming
• Economic
• Non-toxic vehicle
• DISADVANTAGE:
• Lack of speciality

74. CARISCREEN ATP METER / CARIES
SUSCEPTIBILITY TESTING METER
• It is a new device which shows the
result on the Cari Screen meter
within 1 minute .
• The test suggests risk of caries
caused by cariogenic biofilm, & can
be explained to the patient
regarding the consequences of a
biofilm infection.

75. CARISCREEN ATP METER / CARIES
SUSCEPTIBILITY TESTING METER
The test is quick and painless .
Swab sample of the plaque from patients, combined with special
bioluminescence reagents within the swab, will create a reaction which is
then measured with the meter.

76. CARISCREEN ATP METER / CARIES
SUSCEPTIBILITY TESTING METER
• SCORING
• The Cari Screen will give a score between 0
and 9,999 .
• A score under 1,500 is considering relatively
healthy , while above that shows
considerable risk for decay.

77. Oratest: A Simple Chairside Aid for Caries Risk Assessment; Arora
Ruchia , Lahiri Prathik , Masih Updeshc ; INTERNATIONAL JOURNAL OF
DENTAL CLINICS 2009:1(1): 26-30
• The study relates Plaque indices and DMFT counts as indicators of
caries activity to Oratest scores. Despite its drawbacks this test may
be used as an indicator of caries activity. This study aimed at
establishing Oratest as a reliable and convenient chair side procedure
for routine dental practice.

78. LIMITATIONS OF CARIES ACTIVITY TESTS
• NONE of these tests are highly reliable as indicators of expected
caries increments.
• Single parameters are usually read (such as acid produced or colony
counts of bacterial species) but, dental caries is a multifactorial
disease.
• They do not determine factors involved in caries resistance such as
fluoride exposure, maturation of enamel or immune protection.

79. LIMITATIONS OF CARIES ACTIVITY TESTS
• Tests for microbial activity ( lactobacillus count, S.mutans colony
count, synder test, swab test) have media adjusted to a pH of 5.2 or
less, thus selectively eliminating growth of organisms that are not
aciduric .
• Combined use of several selected tests may give a better picture of
the caries activity of an individual.

80. CONCLUSION
• Caries activity tests are a valuable adjunct for patient motivation in a
plaque control program.
• ‘To run a caries preventive program without using microbiological
methods is like running a weight control program without a scale.’
• But, there are no ideal tests in existence at the present time. Further
research is still necessary…

81. REFERENCES
• 1.Marwah N. Textbook of pediatric dentistry.3rd edn.
• 2.Tandon S. Textbook of Pedodontics. 2nd edn. Paras medical
publishers 2009
• 3.Peter S. Essentials of Community Dentistry . 4th edn.
• 4.Studervant CM, Roberson TM, Heymann HO, Studervant JR. The Art
and science of operative dentistry. 3rd ed. Mosby Co: 1995.p. 62-3.
• 5. Michiko Nishimura et al. Using a caries activity test to predict caries
risk in early childhood. JADA. 2008; 139: 63 – 71.