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Effectiveness of a 2%
chlorhexidine solution mixedwith antifungal(clotrimazoleclotrimazole))
agent and calcium hydroxideagainst Candida albicans
Dr GaneshMurthi
M.Sc.D endo student
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Introduction
The presence of Candida albicans in the
infected pulp and periradicular areas has been
demonstrated through the use of light and
electron microscopy, as well as usingculture techniques.
The incidence of C. albicans in infected root
canals has been shown to vary between 7%
and 55% and occurs in approximately 11.36%of teeth with pulp lesions (7). It has also been
reported that C. albicans can use dentin as a
nutrient source.
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Historical backgroundHistorical background
Several studies have tested the effect of
intracanal irrigants and antiseptic agents on C.
albicans, including sodium hypochlorite,
chlorhexidine solution and calcium hydroxide
paste. These antiseptic agents have been
shown to be very effective against C. albicans.
Waltimo evaluated different oral Candida spp.,
including C. albicans, in the presence calcium
hydroxide in vitro. They reported that Candidaspp. were resistant to calcium hydroxide and
concluded that there is a need for
supplementary agents to effectively treat
persistent apical periodontitis.
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Chlorhexidine has been suggested to be a
useful endodontic irrigant and intracanal
antiseptic due to its low toxicity and excellent
anti-bacterial and anti-fungal activities.
A combination of chlorhexidine and calcium
hydroxide was tested against bacterial species
only, and was reported to be effective againstEnterococcus faecalis. It would therefore be of
value to determine the effective
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AimAim
The purpose of the current study is to examine
the In vitro susceptibility ofC. albicans to a
mixture of chlorhexidine, clotrimazole(anti
fungal) and calcium hydroxide.
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Materials and methodsMaterials and methods
The anti-fungal effect of 2% chlorhexidine
solution mixed with clotrimazole,
2% chlorhexidine mixed with calcium hydroxide
is evaluate against C. albicans.
Stock cultures of clinical isolates of C. albicans
were provided by the Microbiology Laboratory
of University of The Philippines, Manila andmaintained in Sabourauds dextrose agar
plates
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Materials and methodsMaterials and methodsMethods Sample size Number of colony
Control sample 20 teeth ?
2% chlorhexidinemixed with calcium
hydroxide paste
20 teeth ?
2%chlohexidine mixed
with clotrimazole
(Antifungal agent)
20 teeth ?
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Experimental procedureExperimental procedure
The experiments were performed in plastic
tissue culture. Clusters containing 24 wells,
each with an inner diameter of 16 mm. One mL
of test material was placed at the bottom ofeach culture well, to which 1 mL of a Candida
suspension was added. At the same time, 1 mL
of Sabouraud infusion broth media was mixed
with 1 mL of a Candida suspension in a culture
well to serve as a positive control.
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Experimental procedureExperimental procedure
For the negative control, 1 mL of Sabouraud
infusion broth was placed in a culture well.
Eight wells were thus used per test.The culture clusters plates were then incubated
at 37oC and evaluated after 1, 24 and 72 h
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Experimental procedureExperimental procedure
At the end of the incubation period, aliquots of
0.1 mL were withdrawn from each well and
transferred to tubes containing 5 mL of fresh
Sabouraud infusion broth
1 mL of a
Candidasuspension
1 mL of a
Candida+
2% CHX +
clotrimazole
1 mL of a
Candida+2% CHX +
CaoH
Group 1 Group 2 Group 3
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Experimental procedureExperimental procedure
Growth of the fungi was observed
daily, as indicated by the presence of turbidity in
the tubes. The presence of turbidity was
determined, and the purity of the cultures was
checked by the morphology of colonies onto
blood agar plates.
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Statistics testStatistics test
The results were statistically analyzed using the
KruskalWall this test is for which group is
best among the three groups
Mann-Whitney Test for compare the two group
and find which group is most significantly
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GeneralGeneral objectiveobjective
Effectiveness of a 2% chlorhexidine solution
mixed with clotrimazole ( anti fungal agent )
and calcium hydroxide against Candida
albicans and evaluated the effectiveness of
each group and find out which group is most
effective in reducing the fungi in root canal and
the drug interaction between the agents and
adverse effect of each agent.
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specificspecific objectiveobjective
1. Determine the number colony in thepresence of the clotrimazole and 2%
chlorhexidine.
2. Determine the number colony in the
presence of the calcium hydroxide and 2%
chlorhexidine
3. Compare the two methods.
4. Which method is most effective in reducethe number of colony.
5. Which method is least effective in reduce the
number of colony
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ResultsResultsDescriptive Statistics
ROUP N Minimum Maximum MeanSt
De iation
1.00 COLONY 20 567.00 976.00 754.8500 123.04996
Valid N
(list ise
20
2.00 COLONY 20 365.00 721.00 553.2500 99.97414
Valid N(listwise)
20
3.00 COLONY 20 123.00 611.00 300.7000 111.67674
Valid N(listwise) 20
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Results
Kruskal-Wallis TestHO X1 = X2 = X3
= 0.05
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Results
Kruskal-Wallis Test
Sample Number of
sample
Colony
median
Mean
Rank Result
Control
group
20 754.85 48.08 Chi square
test 43.588
Asymp.sig
.000***2% CHX +
CaoH
20 553.25 31.73
2% CHX +
clotrimazol
e
20 300.70 11.70
There is highly statically significantly
different between the three groups
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Mann-Whitney Test
Group Meanrank Z value Mann-Whitney
U
P value
Group 1
Group 2
30.35 -5.331 3.00 0.030**
10.65Group 1
Group 3
28.23 -4.181 45.50 0.000***
12.78
Group 2Group 3
29.15 -4.844 21.00 0.075*11.55
There is highly statically significantly
different between group 1 and group 3 the
three groups
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DiscussionDiscussion
The method used to evaluate inhibition of fungal
growth in this study was the dilution tube
susceptibility test, which is an effective method
to evaluate anti-fungal as well as anti-bacterialproperties of any filling material or solution.
This method allows direct contact in the
solution betweenC. albicans
and the agent tobe tested.
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DiscussionDiscussion
The growth ofC. albicans observed in thepositive controls confirmed the efficiency of the
methodology.
Three agents, chlorhexidine, clotrimazole and
calcium hydroxide, were chosen for this study.
Chlorhexidine is a broad-spectrum antimicrobial
agent that has been reported to be an effective
antiseptic in endodontic therapy owing to itsunique ability to bind to dentin and its
substantivity in the root canal system. It is also
effective against strains resistant to calcium
hydroxide
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DiscussionDiscussion
In the present investigation, mixing clotrimazole
with chlorhexidine was found to be very
effective against C. albicans at all experimental
times tested.
The anti-bacterial property of chlorhexidine
depends on its long-lasting alkalinity and has
been reported not to be affected by the addition
of clotrimazole
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DiscussionDiscussion
Such a combination will result in better diffusion
into the dentinal tubules of prepared root canal
systems with the potential to be used as a long-
term intracanal agent and therefore completeanti-microbial, as well as anti-fungal,
effectiveness can be improved.
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CConclusiononclusion
In conclusion, the results of the present study
show the additive benefits achieved by
combining Chlorhexidine with clotrimazole in
the treatment ofC. albicans
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Any Question ?Any Question ?