|Year : 2014 | Volume
| Issue : 2 | Page : 91-96
Comparison of antimicrobial efficacy of chlorhexidine and combination mouth rinse in reducing the Mutans streptococcus count in plaque
Laxmi S Lakade1, Preetam Shah2, Dayanand Shirol3
1 Assistant Professor, Pediatric and Preventive Dentistry, Bharati Vidyapeeth Deemed University Dental College and Hospital, Pune, Maharashtra, India
2 Professor, Pediatric and Preventive Dentistry, Bharati Vidyapeeth Deemed University Dental College and Hospital, Pune, Maharashtra, India
3 Professor, Pediatric and Preventive Dentistry, M. A. Rangoonwala Dental College and Hospital, Pune, Maharashtra, India
|Date of Web Publication||17-Apr-2014|
Laxmi S Lakade
Patang plaza tower building phase 6, Flat no 1, Opp- Pune Institute of Computer Technology College, Katraj, Pune - 411 046, Maharashtra
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: The removal of plaque is utmost important to control dental caries. But in children, factors like lack of dexterity, individual motivation and monitoring limit the effectiveness of tooth brushing. This necessitates the use of chemotherapeutic agents for control of plaque. Aims: To compare the antimicrobial efficacy of 0.2% chlorhexidine mouth rinse and mouth rinse containing 0.03% triclosan, 0.05% sodium fluoride, and 5% xylitol in reducing the Mutans streptococcus count in plaque. Materials and Methods: Thirty healthy children aged 8-10 years with dmft (decay component) of three or four were selected. They were divided randomly into two groups: The control or chlorhexidine group and the study group or combination mouth rinse. Both the groups practiced rinsing with respective mouth wash for 1 min for 15 d twice a day. The plaque samples were collected and after incubation Mutans streptococcus count was estimated on the strips from the Dentocult SM kit and evaluated using manufacture's chart. Statistical Analysis Used: Wilcoxon matched pairs signed ranks test and Mann-Whitney U test were used to analyze the findings. Results: Statistically significant reduction in the Mutans streptococci count in the plaque was seen in the control and study group from baseline level. But when both the groups were compared, the antimicrobial effect of chlorhexidine was more.
Keywords: Dental plaque, chlorhexidine, combination mouth rinse, triclosan, xylitol, Mutans streptococci, Dentocult SM strip
|How to cite this article:|
Lakade LS, Shah P, Shirol D. Comparison of antimicrobial efficacy of chlorhexidine and combination mouth rinse in reducing the Mutans streptococcus count in plaque. J Indian Soc Pedod Prev Dent 2014;32:91-6
|How to cite this URL:|
Lakade LS, Shah P, Shirol D. Comparison of antimicrobial efficacy of chlorhexidine and combination mouth rinse in reducing the Mutans streptococcus count in plaque. J Indian Soc Pedod Prev Dent [serial online] 2014 [cited 2019 Nov 19];32:91-6. Available from: http://www.jisppd.com/text.asp?2014/32/2/91/130780
| Introduction|| |
Modern concepts consider caries as an interaction between genetic and environmental factors in which social, behavioral, psychological, and biological factors are expressed in a highly complex interactive manner.  But the important part in the understanding of the caries process is that it does not occur in the absence of dental plaque or dietary fermentable carbohydrate hence, is considered a dietobacterial disease. 
The role played by bacteria in initiation of dental caries and periodontal diseases  is well established. The removal of plaque is utmost important to control dental caries that is commonly maintained by mechanical methods. But in children, factors like lack of dexterity and individual motivation and monitoring limit the effectiveness of tooth brushing. They also experience difficulty in maintaining adequate plaque control, particularly at interproximal sites, which necessitates the use of chemotherapeutic agents for control of plaque. 
Among the chemotherapeutic agents used in mouthwashes, chlorhexidine is the "gold-standard" or positive control for comparison with other substances due to its proven efficiency. , Though effective, it has certain side effects like brown discoloration of the teeth, oral mucosal erosion, and bitter taste. Hence, there is need of an alternative mouth rinse that could negate all the side effects of chlorhexidine, but yet effective equivalent to it.
Therefore, this study was carried out to evaluate the antimicrobial efficacy of 0.2% chlorhexidine mouth rinse and mouth rinse containing 0.03% triclosan, 0.05% sodium fluoride, and 5% xylitol in reducing the Mutans streptococci count in plaque.
| Materials and Method|| |
The randomized, controlled, and double-blind study was carried out in the 30 healthy children aged 8-10 years from the Department of Pedodontics and Preventive Dentistry, Bharati Vidyapeeth Dental College, Pune. Children with high caries risk experience of dmft of three or four (decay component) were selected.
Exclusion criteria were children with physical limitations, which might preclude the normal tooth brushing and mouth rinsing, intraoral soft tissue pathology, medically compromised patients, and subjects with history of taking antibiotics three months before or during the course of study. Subjects undergoing orthodontic treatment or with extensive intraoral prosthesis, children who had previously restored/crowned teeth and children having teeth with periapical pathology were excluded from the study.
The study was explained to parents and consent was obtained. Child's personal details, details of past medical history including any recent antibiotic exposure, past dental history including recent fluoride treatment, frequency of brushing, sweets/snacks intake and consumption of sugared/energy drinks, and the brand of toothpaste (to determine fluoride content) were obtained from parents through questionnaire.
The Mutans streptococci count in plaque was determined by using the chairside method (Dentocult SM Strip mutans Orion Diagnostica, Espoo, Finland) [Figure 1]. This test is based on the principle of use of a selective culture broth, the adherence and growth of Mutans streptococcus bacteria on the test strip.
The procedure of using this test was in accordance to the manufacturer. Before collecting the plaque samples, the vials were brought down to room temperature 1 h before use and shaken gently. Using a forceps, two bacitracin discs were placed in the selective culture broth about 15 min before sampling [Figure 2].
Isolation with cotton rolls was done. Plaque samples were collected using different toothpicks from the four sites enamel buccal surface of the maxillary right molar, labial surface of the maxillary incisor, labial surface of the mandibular incisor, and lingual surface of the mandibular left molar. These samples were spread thoroughly but gently on the four sites of the rough surface of the strip [Figure 3]. The selective culture vial was gently shaken for even distribution of bacitracin, and the strip was placed in the selective culture broth. The vials were then labeled with the numbers and incubated in an upright position at 37C for 48 h with one quarter of the cap turned open to allow growth of the organisms. Following incubation, the presence of Mutans streptococcus was confirmed by dark-blue to light-blue raised colonies [Figure 4].
Inspection of growth was done sideways against light or with a magnifying glass to look for raised colonies. Colonies suspended in the culture broth were excluded from the evaluation as suggested by manufacturer of Dentocult SM Strip mutans. The results were interpreted according to the manufacture's model density chart and classified as
Class 0: < 10,000 CFU/ml*
Class 1: < 100,000 CFU/ml
Class 2: 100,000-1,000,000 CFU/ml
Class 3: > 1,000,000 CFU/ml
*CFU/ml - colony forming unit/milliliter
Inspection of the growth was done with the strip held sideways against light and magnifying glass. The presence of epithelial cells on the strip surface can be differentiated from the Streptococcus mutans colonies by passing a gloved finger along the strip: The epithelial cells are smooth, whereas the S. mutans colonies are rough.
After the collection of baseline status of plaque samples, the subjects were randomly divided into two groups, 15 children in each group. The control group was advised to rinse with 10 ml of 0.2% chlorhexidine gluconate (Hexidine, ICPA products Ltd) in the morning and in the night after brushing for l min for 15 d under parent's supervision. The study group was advised to rinse with 5 ml of combination mouth rinse containing 0.03% triclosan, 0.05% sodium fluoride, and 5% xylitol (Kidodent, Warren) in the morning and in the night after brushing for l min for 15 d under parent's supervision. After rinsing, the children were advised not to eat or drink for 30 min. During the course of the study, the children were asked to use the non-fluoridated tooth paste and the new brush (Colgate Zig Zig - Junior) that was provided to them to prevent microbial contamination. The subjects in both the groups were blinded about the division of group.
After 15 d of using mouth rinse, the plaque samples were again collected, incubated, and interpreted in similar manner as taken for baseline status to assess change in count level of Mutans streptococcus. The data obtained from the study were tabulated and analyzed statistically with Wilcoxon matched pairs signed ranks test and Mann-Whitney U tests.
| Results|| |
There was no difference in total number of Mutans streptococci at baseline level in both the groups as seen in [Table 1].
[Table 2] shows statistically significant reduction (P = 0.001) in mean Mutans streptococci count after rinsing with 0.2% chlorhexidine when compared with baseline count.
|Table 2: Comparison of mutans streptococcus score in plaque in chlorhexidine group with respect to prerinse and postrinse|
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[Table 3] shows statistical significant reduction (P = 0.002) in mean Mutans streptococci count after rinsing with combination mouth rinse when compared with baseline count.
|Table 3: Comparison of mutans streptococcus score in plaque in combination mouth rinse group with respect to prerinse and postrinse|
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[Table 4] when both the groups were compared 0.2% chlorhexidine reduced more number of Mutans streptococcus then the other group. Therefore, 0.2% chlorhexidine was more effective antimicrobial agent.
|Table 4: Comparison of postrinse mutans streptococcus score with respect to chlorhexidine group and combination mouth rinse group|
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| Discussion|| |
Many children have inadequate oral and general health because of active and uncontrolled dental caries. It is the single most common chronic childhood disease. Owing to its non-life-threatening nature and ubiquitousness has minimized its significance in overall human health. Initiation of dental caries and the microbial composition of plaque have generally involved either S. mutans or Lactobacilli. Children with high dmft have increased S. mutans count. As a result variety of anti-plaque agents has been examined for their ability to control S. mutans.
Among the chemotherapeutic agents used in mouthwashes, chlorhexidine is the "gold-standard" or positive control for comparison with other substances due to its proven efficiency. , Along with antibacterial effect, its main advantage is 'substantivity'. But due to its known disadvantages, there is need for ideal anti-plaque agent, which is not yet available. Hence, this study evaluated the efficacy of two mouth rinses to reduce S. mutans. An approach to increase the efficacy of anti-plaque agent and to reduce the adverse effects may be to combine two or more agents. Triclosan is broad spectrum antimicrobial activity and is effective against S. mutans at low concentration.  The effects of fluoride on bacterial metabolism are well-known. Fluorides inhibit several essential enzymes in oral bacteria as stated by Hamiton and Bowden in 1988.  Mellberg and Ripa  suggested that low potency-high frequency rinsing may be more beneficial. Xylitol is non-sugar sweetener permitted for use in food.  Hence, combination mouth rinse of 0.03% triclosan, 0.05% sodium fluoride, and xylitol was used.
In this study, a chairside test - the "Strip mutans" test - was used that was developed by Jensen and Bratthall.  The conventional technique ,,, involves laborious laboratory steps. Hence, it lacks widespread use in routine dental practice. The sensitivity, specificity, and accuracy of the Dentocult SM was found to be better than those of conventional methods.  In their study, the plaque test surpassed the salivary strip test in terms of sensitivity and accuracy when both were compared. Therefore, plaque collection was preferred over saliva collection in the present study.
There was no difference in total number of Mutans streptococci at baseline level in both the groups as seen in [Table 2]. Similar finding was observed by Neeraja et al.,  This can be explained by the fact that dmft (decay component) was standardized. 
A total of 0.2% chlorhexidine significantly reduced [Table 3] Mutans streptococci count. Its action is due to adsorption of chlorhexidine onto the cell wall of the microorganism, resulting in a leakage of intracellular components.  At low concentration, chlorhexidine a small molecular weight substance such as potassium and phosphorus, will leach out, exerting a bacteriostatic effect. At higher concentrations, chlorhexidine is bactericidal because of precipitation or coagulation of the cytoplasm, probably caused by protein cross-linking.  But bactericidal effect is thought to be less important than the bacteriostatic effect provided by a slow release of chlorhexidine. 
This observation adds to earlier studies carried by Happonen et al.,  Heffi and Huber  Kulkarni and Damle  Neeraja et al.,  noted immediate significant reduction in Mutans streptococci in chlorhexidine group, but after 15 d therapy there was an increase in Mutans streptococcus count. But this increase was less in chlorhexidine as compared with povidone-iodine. Whereas Kulkarni and Damle  found the significant reduction in Mutans streptococcus in chlorhexidine and triclosan group than control group after 2 weeks of rinsing. Many studies have shown that it is not possible to devoid the mouth completely of S. mutans for an extended period, no matter how rigorous or extended the application. ,, So, complete elimination of Mutans streptococcus (or Class 0) was not seen in any children at the end of 15 d in our study also.
The combination mouth rinse [Table 4] also showed significant reduction in Mutans streptococcus count due to effects of its components. Triclosan has the antimicrobial action. Owing to its hydrophobic and lipophilic nature, it adsorbs to lipid portion of the bacterial cell membrane and in low concentrations it interferes with vital transport mechanism.  Fluoride, which is another component, is a powerful inhibitor of acid formation by plaque microorganisms.  The current evidence indicates that fluoride has a multitude of direct and indirect effects on the bacterial cell, which have a significant influence on those organisms in dental plaque.  Enzyme enolase in glycolytic pathway of the carbohydrate metabolism is fluoride sensitive.  Guha-Chowdhury et al.,  found that it inhibits the streptococcal enolases also. Maguire and Rugg-Gunn  described the action of xyiltol as non-fermentability and non-cariogenicity as passive effects, whereas active caries prevention effects as bacteriostatic and cariostatic.
However, in this study 0.2% chlorhexidine showed a greater reduction of Mutans streptococcus count than combination mouth rinse [Table 4]. Similarly Sharma et al.,  found 0.2% chlorhexidine most effective than combination mouthwash containing 0.03% triclosan and 0.05% sodium fluoride.
The high efficacy of chlorhexidine could be due to its immediate bactericidal action during the time of application followed by a prolonged bacteriostatic action due to adsorption at the tooth surface.  The adsorbed (substantivity) chlorhexidine is gradually released for up to 24 h.  A total of 0.2% chlorhexidine inhibits acid production for 24 h after sucrose applied to dental plaque in vivo. 
Studies ,, showed that chlorhexidine significantly reduced the bacterial numbers upto 7 h, and effect persists for many hours.  Perhaps more importantly there was no evidence of bacterial regrowth following chlorhexidine as indicated by negative increments. Such was not the case for triclosan, the incremental values were positive from 60 min indicating recovery of bacterial counts.  Triclosan has plaque inhibitory effects mediated by an antimicrobial action, which would not be much different from that of sodium lauryl sulfate.  Therefore, triclosan itself has moderate plaque-inhibitory effect.  Limited potential for absorption and reduced substantivity of triclosan compared with chlorhexidine. , There is evidence indicating that the ingredients, vehicle, and other active substances may influence its antimicrobial activity and consequently its efficiency of triclosan.  Effects of fluoride are well established as anticaries process. But certain studies state that fluoride preparations (except SnF 2 and amine fluorides) seem to have little effect on the quantity of plaque and thereby antimicrobial effect. 
Therefore, more extensive studies with larger samples and varied time periods should be carried out to establish the efficacy of combination mouth rinse.
Summary and conclusions
Apparently, the ideal anti-plaque agent is not available. Any chemical agent that affects microbial cells may be expected to have some adverse effects against host cells, unless the target structure or metabolic pathway is unique to the microbial cell. But plaque is a complex aggregation of various bacterial species. Thus, no single agent can be effective in complete elimination of plaque. Combining two or more agents will compliment the modes of action resulting in additive or synergistic effects, with minimal adverse effects. For effective anti-plaque agents to be developed, it is crucial for the target structures to be defined, and the exact modes of action of the potential agents be known. Conclusions from this study:
- A total of 0.2% chlorhexidine gluconate was a more effective antimicrobial agent than mouth rinse containing 0.03% triclosan, 0.05% sodium fluoride, and 5% xylitol in reducing the Mutans streptococci count in plaque.
Although the chemical anti-plaque agents are effective in reducing the microbial count in plaque but are only as an adjunct to mechanical modes of plaque control and not the substitutes.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4]