Anticaries Effect of Xylitol,Probiotic and Chlorhexidine Mouth Rinses Among Individuals at High Risk
Condition: Cariostatic Agents; Mouthwashes
Study type: Interventional
Phase: Phase 2/Phase 3

Summary: 
      Dental caries is a localized, post eruptive pathological process involving softening of the
      hard tooth tissue and proceeding to the formation of cavity. It results from accumulation of
      plaque on the surface of the teeth and biochemical activities of complex microorganisms.
      Streptococcus mutans is one of the main pathogens of dental caries. Although brushing has the
      potential to maintain adequate levels of oral hygiene, studies demonstrate that such methods
      are not being employed sufficiently. The need for additional help in controlling bacterial
      plaque provides the rationale for patients using mouth rinses as adjuncts. Chlorhexidine
      although considered the gold standard, its side effects due to prolonged use such as staining
      of teeth, dry mouth, altered taste sensation, mouth/ throat irritation, etc indicates the
      need for alternatives.

      Probiotics are live microorganisms that, when administered in adequate amounts, confer a
      health benefit on the host. Probiotic technology represents a breakthrough approach to
      maintaining oral health by utilizing natural beneficial bacteria commonly found in healthy
      mouths to provide a natural defense against those bacteria thought to be harmful to teeth and
      gums. The advantages of using a probiotic mouth rinse are that it contains friendly microbes,
      there is no issue of antibiotic resistance, and there are no known/proven toxicities caused
      due to their use.

      Xylitol is a non-sugar sweetener permitted for use in foods. Xylitol is a naturally occurring
      non-cariogenic sugar substitute that cannot be metabolized by oral bacteria. This possesses
      various properties favorable for caries prevention.It acts by forming of loosely attached
      biofilms on the tooth surfaces that can be easily removed. Although many studies have
      evaluated the effects of xylitol chewing gum on caries prevention, there are very few
      examining the effect of xylitol mouth rinse on oral Streptococcus mutans. Hence xylitol mouth
      rinse may be introduced as a possible alternative.

      Antimicrobial efficacies of probiotics and xylitol mouth rinses have not been compared till
      date. Also their effects on the young and elderly population have not been compared. Hence
      the purpose of the study is to compare the antimicrobial efficacies of probiotic & xylitol
      mouth rinses with that of chlorhexidine in children and elderly population.
    

Detailed description: 
      Need for the study:

      World Health Organization defines dental caries as a localized, post eruptive pathological
      process of extreme origin involving softening of the hard tooth tissue and proceeding to the
      formation of cavity. The process involves bacterial interactions in plaque accumulated on the
      surface of the teeth. Streptococcus mutans in plaque is the most commonly isolated organism
      amidst all other cariogens. It ferments sucrose and the resulting acid causes
      demineralization of tooth enamel.

      While mechanical methods of plaque control can maintain adequate oral hygiene, such methods
      are not being utilized appropriately by the population. This necessitates use of adjuncts to
      mechanical plaque control methods in the form of antiplaque mouth rinses. Chlorhexidine mouth
      rinse has been considered the most effective agent in inhibiting Streptococcus mutans.
      Although considered the gold standard, its adverse effects due to prolonged use such as
      staining of teeth, xerostomia, altered taste sensation, mouth/ throat irritation,
      antimicrobial resistance, etc. indicates the need for alternatives which have been
      extensively studied of late in literature.

      Xylitol used as an artificial sweetener in foods, is a non-cariogenic sugar substitute. This
      polyalcohol cannot be metabolized by oral bacteria thereby preventing caries. The magnitude
      of adverse effects caused by xylitol as compared to that of Chlorhexidine is indistinct.
      Although studies have assessed effects of xylitol chewing gum on caries, very few have
      studied its efficacy as a mouth rinse.

      Probiotics are live microorganisms that, when administered in adequate amounts, confer a
      health benefit on the host. Probiotic products seem to have an effect on the oral health of
      individuals by prompting beneficial bacteria to defend teeth and gums against harmful ones.
      The advantages of probiotics as compared to that of chlorhexidine mouth rinse are that there
      are no issues of antibiotic resistance as it contains only commensal flora and there has been
      no proof of intoxication or allergies on consumption.

      Antimicrobial efficacies of probiotics and xylitol mouth rinses have not been compared till
      date. Their effects on the young and elderly population have not been compared. Hence the
      purpose of the present study is to compare the antimicrobial efficacies of probiotic &
      xylitol mouth rinses with the gold standard (Chlorhexidine) in children and elderly
      population.

      Aim of the study:

      To compare the antimicrobial efficacy of probiotic and xylitol mouth rinses with
      chlorhexidine (gold standard) among children and elderly population at high risk for dental
      caries.

      Objectives of the study:

        1. To assess Streptococcus mutans levels at baseline and 14 days after intervention in
           children and elderly population.

        2. To compare efficacies of the 3 mouth rinses in reducing Streptococcus mutans levels in
           children and elderly.

      Materials and methods:

      Source of data: Residential school children aged 5-12 years at high risk for caries. Elderly
      population above 60 years residing in old age homes at high risk for caries.

      Study Design: Double Blind Randomized Controlled Trial.

      Sample Size: Twelve participants will be chosen per group. With 3 products involved in each
      of the 2 population groups (children and elderly) the sample size will be 36 child
      participants and 36 elderly participants.

      Consent from participants: Permission to conduct the study in the residential school and old
      age home will be obtained from the concerned authorities. Details about the study will be
      presented to the participants in the form of a study information sheet or communicated
      verbally in both English and in the local language. Only the eligible participants who
      provide written consent will be included in the study. In case of children, written consent
      will be obtained from the guardians.

      Caries risk assessment: Individuals with high caries risk will be identified using a
      customized caries risk assessment tool. The caries risk assessment tool will comprise of
      information on socio-economic status, oral hygiene practices, fluoride exposure, caries
      experience amongst family members, symptoms of dry mouth, quantity and frequency of intake of
      sweetened food and caries experience in the past and present. Each item will be scored as 0
      or 1 and the total score will be obtained by summing up the scores of all the items. Any
      participant with total score greater than 5 will be considered to be at high risk for caries.

      Method:

      1. Preparation of mouth rinses 2. Baseline data collection and Microbial Analysis 3.
      Randomization and Group Allocation 4. Intervention 5. Post intervention data collection and
      Microbial Analysis

        1. Preparation of mouth rinses:

           Xylitol mouth rinse at 10 percentage concentration will be used. Probiotic mouth rinse
           will be prepared by using a commercially available probiotic product (Sporolac Plus
           powder- 1gm sachet containing not less than 1.5 billion cells of Lactobacillus
           acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum, Bacillus coagulans and
           Saccharomyces boulardii). Each sachet will be dissolved in 15 ml of water in a measuring
           cup and will be used as a mouth rinse. A commercially available Chlorhexidine mouth
           rinse (Hexidine- 0.2 percentage Chlorhexidine gluconate) containing 0.2 percentage
           chlorhexidine gluconate per 10 ml will be used.

        2. Baseline data collection and microbial analysis:

      Plaque samples will be collected from the buccal surface of a non-carious permanent maxillary
      first molar. Plaque collection will be done using an autoclaved scaler under daylight. The
      collected plaque will be stored in a pre-weighed sterile eppendorf tube. Tubes will be
      weighed again after plaque collection. The weight of the collected plaque (in grams) will be
      determined by subtracting the weight of the empty eppendorf tube from the weight of the tube
      with the collected plaque. The samples will be stored at - 4 degree Celsius and transported
      to a culture lab within an hour to avoid using transport media.

      1 ml saline will be added to the eppendorf tube and vortexed for even distribution of plaque.
      This mixture will be used as stock solution for serial dilutions. From the stock solution,
      100µl will be transferred to a sterile test tube containing 900µl of saline and vortexed to
      arrive at 1:10 dilution. Similar dilutions will be prepared to obtain 1/10, 1/100 and 1/1000
      dilutions. 50µl from each of the dilutions will be plated onto the selective medium MSB
      (Mitis Salivarius Bacitracin Agar) by spread plate method. After plating, the MSB (Mitis
      Salivarius Bacitracin Agar) agar plates will be placed in the anaerobic jar and incubated at
      37 degree Celsius for 72 hours.

      Colonies of Streptococcus mutans will be identified based on the following morphologic
      characteristics, a) 0.5 mm raised convex undulated colonies b) light blue colour with rough
      margins c) granular frosted glass appearance. The colonies will be confirmed by a catalase
      test (negative catalase reaction) and gram staining (gram positive cocci). Bacterial colonies
      will be counted manually. The standard formula for determining Colony Forming Units (CFU)
      will be CFU/g= [Number of colonies X Dilution factor] / [Volume plated (in ml) X Amount of
      plaque (in g)]

      3. Randomization and Group allocation: After data collection at baseline, the study
      participants will be allocated to three groups, Group A, B and C, by simple random sampling
      (lottery method). Group A will be given Chlorhexidine mouth rinse; Group B will be given
      Xylitol mouth rinse and Group C Probiotic mouth rinse. The participants and the investigator
      will be blinded from the allocation sequence.

      4. Intervention: The participants will be asked to rinse their mouth once daily (at night)
      for 2 minutes, using 15 ml of mouth rinse. The intervention will be carried out for a period
      of 14 days. Mouth rinsing will be supervised during the study period by an assistant. A
      record will be maintained to document regular usage of the mouth rinse and also to record any
      adverse effects occurring during the intervention period.

      5. Post intervention data collection: After 14 days of using the mouth rinses, the same
      procedures will be repeated and compared with baseline.

      Statistical Analysis Plan: Data will be analysed using SPSS (Statistical Package for the
      Social Sciences Version 22.0; SPSS Incorporated, Chicago, Illinois). Comparisons before and
      after interventions will be done using Paired t Test. Comparisons between Chlorhexidine,
      probiotic and xylitol groups will be done using ANOVA (Analysis of Variance) and Tukey's Post
      Hoc Tests. Comparison of antimicrobial efficacy of mouth rinses between children and elderly
      will be done using Independent sample t Test. Statistical significance will be fixed at p ≤
      0.05.
    

Intervention name(s): Chlorhexidine mouthwash; Xylitol; Probiotic
Intervention type(s): Drug; Drug; Drug
Intervention description(s): A commercially available Chlorhexidine mouth rinse (Hexidine- 0.2 percentage Chlorhexidine gluconate) containing 0.2 percentage chlorhexidine gluconate per 10 ml will be used. 7.5 ml of the concentrate diluted with equal amounts of water to make 15ml will be used for rinsing.; Xylitol mouth rinse at 10 percentage concentration will be used. The mouth rinse will be prepared by dissolving 1.5 gm of xylitol powder in 15 ml of water.; Probiotic mouth rinse will be prepared by using a commercially available probiotic product (Sporolac Plus powder- 1gm sachet containing not less than 1.5 billion cells of Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum, Bacillus coagulans, Saccharomyces boulardii). Each sachet will be dissolved in 15 ml of water in a measuring cup and used as a mouth rinse.
Other intervention name(s): Hexidine; Xylitol powder (Loba Chemie, code 06512); Sporolac Plus powder- 1gm sachet
Intervention MeSH terms: Chlorhexidine
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Double (Participant, Outcomes Assessor)
Primary purpose: Prevention
Keywords: Streptococcus mutans; Chlorhexidine; Xylitol; Probiotic