The comparison of the antibacterial effect of various

mouth rinses towards common oral bacteria

PRESENTER : FARALIZA ALIASDepartment of Oral Biology,

Faculty of Dentistry, University of Malaya

INTRODUCTIONORAL MICROBES

The microorganisms that form the plaque/biofilm on the surfaces of the teeth are mainly Streptococcus mutans.

Other species :-› Streptococcus mitis› Streptococcus sanguis.

If not taken care of, via brushing or flossing and rinsing, the plaque can turn into tartar (its hardened form) and lead to gingivitis or periodontal disease.

Various streptococci in a biofilm in the oral cavity (electron microscope)

Gram staining of Streptococcus mutan

MOUTH RINSES

Mouth rinse is a product used to enhance oral hygiene.

Antibacterial agent in the mouth rinse can kill the bacterial plaque causing caries, gingivitis, and bad breath.

Active ingredients :- Chlorhexidine gluconate (CHX), Cetylpyridinium chloride (CPC), Thymol (THY).

Mouth Rinse

OBJECTIVES

To compare the antibacterial effect of the commercially available mouth rinses towards mixed three oral bacteria commonly associated with dental caries.

› Streptococcus mutans› Streptococcus mitis› Streptococcus sanguinis).

MATERIAL & METHODOLOGY

SUSCEPTIBILITY TESTS (AST)

CULTURE PREPARATIONS

MINIMAL INHIBITORY CONCENTRATION

TEST (MIC)

MINIMAL BACTERICIDAL

CONCENTRATION DETERMINATIONS

(MBC)

CULTURAL PREPARATIONS

3 Streptococcus spp. from ATTC were used in this study.

BHI media was used to revived & culture the microorganisms.

Take the bacteria using sterile cotton swab & make a suspension.

Read OD

0.144

Mixed 3 suspension of

bacteria.

Take 100ul suspension and & place on the BHI

agar

CULTURAL PREPARATIONS

Using hockey stick spread the suspension

using lawn method

Incubated 24 hrs at 37°C

Bacteria ready to use

SUSCEPTIBILITY TESTING The mouth rinses were tested for sensitivity using Kirby-

Bauer susceptibility test with principle of agar disc diffusion.

Let it dry at

RT

Repeat procedure 2 & 3 until total solution diffused in a disk 100ul

Prepare disk containing mouth rinses using whatman sterile

paper disk size 6mm

Pipette 20ul mouth rinse into the disk

Let it completely dry & ready to

use

SUSCEPTIBILITY TESTING

Place the disk on the agar using

sterile forcep

Fresh bacterial suspension

Using sterile cotton swab culture the bacteria on BHI

agar

Inubated 24hrs at 37 oC

2 1-

+3

Measure & record the results

MIC TESTING

Using 96 wells broth micro-dilution assay was used to determine the minimal inhibition concentrations (MIC).

MIC TESTING

1 2 3 4 65

Mouth rinse

(500 ul) Discard

Mixed bacteria suspension

MouthRinse(X)

• The first tube with no bacteria growth (no turbid suspension) is the dilution containing the concentration of the mouth rinse which is refered MIC .

Incubated 37ºc (24 hour)

MBC DETERMINATIONS The Minimum Bactericidal Concentration

(MBC) is the lowest concentration of antibiotic required to kill the germ.

MBC Techniques

Flow chart MIC & MBC

RESULTS & DISCUSSIONS

Disc Inhibition Zones (cm)

0.12 % CHX 2.73 cm0.05 % CPC 2.27 cm

THY Resistant0.12 % CHX - Standard(+ve

control)

2.53 cm

D.H2O (-ve controll)

Resistant

Table 1: Sizes of inhibition zones that were present after 24 hour.

The susceptibility of active compound was shown by the formation of clear zone of growth inhibition around the paper discs.

AST

RESULTS & DISCUSSIONS

It was clearly shows that CHX and CPC containing mouth rinse are able to prevent the growth of the mixed oral bacteria at the tested concentrations. THY showed less susceptibility effect when compared with blank control and positive control (fig.AST).

The greater the size of inhibition zones, the greater the antibacterial potency of the mouth rinse.

RESULTS & DISCUSSIONSMIC & MBC

HIGHER CONCENTRATION

LOWER CONCENTRATION

TURBID - HIGH BACTERIA

RESULTS & DISCUSSIONSMIC & MBC

Figure MBC: Minimal bactericidal concentration (MBC) Test. The concentration on the agar that shows no bacteria growth was recorded as the MBC of the mouth rinse – agar label as tube 3 shown NO GROWTH (MBC)

RESULTS & DISCUSSIONS

The MICs and MBCs for CHX and CPC were determined as low as 0.00375% w/v (tube 3) and 0.0125% w/w respectively .

This showed that CHX and CPC exhibited profound antibacterial activity on selected common oral bacteria as demonstrated by the very low MIC and MBC values.

THY showed its antibacterial effect only at higher concentration (>50 % v/v).

MIC & MBC

CONCLUSIONS

The present study showed that mouth rinses vary significantly in their capability to inhibits and kill oral bacteria.

The CHX containing mouth rinse demonstrated superior antibacterial activity toward selected mixed tested bacteria followed by CPC containing mouth rinse .

This implies that the incorporation of CHX and CPC in mouth rinses are better than that containing THY.

REFERENCES A.R. Fathilah and Z.H.A Rahim. J. Oral Sci., 45: 201-206 (2003). A.R. Fathilah, Y. Othman and Z.H.A Rahim. J. Oral Sci., 48 (2): 71-75 (2006). A.R. Fathilah, Z.H.A. Rahim, Y. Othman and M. Yusoff. Pak. J. Biol. Sci., 12(6):

518-521 (2009). T. Nalina and Z.H.A. Rahim. Pak. J. Biol. Sci., 9: 1470-1475 (2007). Pan P. C., Harper S., Ricci-Nittel D., Lux R. and Shi W. J. Den 38, S1 S16-S20

(2010)

ACNOWLEDGEMENT•This study was financially supported by the Department of Oral Biology, University of Malaya Grant (BM271)

•All Staff & student Oral biology Department

THANK YOU