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Completed

NCT Number: NCT07570927

Salivary Microbiota and Antimicrobial Resistance in Relation to Chlorhexidine Use

This study evaluated the ecological effects of 0.06% chlorhexidine toothpaste on the salivary microbiota of healthy individuals, focusing on taxonomic composition, functional shifts, and prevalence of predicted antimicrobial resistance genes. Saliva samples were collected from eleven healthy participants at three specific time points: baseline (following a 4-week run-in period using a control toothpaste), intervention (after 12 weeks of using 0.06% chlorhexidine digluconate toothpaste), and washout (after a subsequent 12-week period using the control toothpaste). Sequencing targeted the 16S rRNA V3-V4 region.

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Key information

Conditions

Antimicrobial Resistance (AMR) Chlorhexidine Oral Microbiome Health Oral Microbiota Saliva Microbiome

Age range

25 year–44 year

Sex eligibility

All sexes

Study type

Interventional

Phase

Not applicable

Primary location

Department of Public Health, Faculty of Medicine, Masaryk University, Brno, Czechia

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Who can participate

Healthy volunteers accepted: Yes

Only the study team can determine whether someone qualifies for participation.

Inclusion criteria

  • Age: Between 25-44 years.
  • General Health: Good systemic health (absence of chronic inflammatory diseases or conditions affecting oral tissues).
  • Oral Hygiene Routine: Participants must perform toothbrushing twice a day and use interdental cleaning aids once a day.
  • Plaque Index (PI): A score of 0.0-0.2 according to Silness and Löe.
  • Gingival Index (GI): A score of 0.0-0.143 according to Silness and Löe.

Exclusion criteria

Systemic and Lifestyle Factors:

  • Presence of any chronic systemic disease (e.g., diabetes mellitus, autoimmune disorders)
  • Regular use of medications that may affect gingival status or saliva production.
  • Use of antibiotics within the last 6 months prior to the study.
  • Current pregnancy or lactation.
  • Current smokers or users of other tobacco/nicotine products.

Oral and Dental Factors:

  • Active dental disease: Presence of untreated tooth decay, gingivitis (GI > 0.143), or any form of periodontitis.
  • Prosthetics and Implants: Presence of dental implants, dentures, or significant tooth loss (excluding extracted third molars).
  • Ongoing fixed or removable orthodontic treatment.

Treatment and study plan

0.06% Chlorhexidine Digluconate Toothpaste

Drug

Participants used the toothpaste twice daily for 12 weeks.

Primary outcomes

  1. Change in Salivary Microbial Alpha Diversity (Shannon and Simpson Indices)

    Time frame: Baseline (week 0), after intervention (week 12), and after washout (week 24).

    Evaluation of microbial richness and evenness within individual saliva samples. Alpha diversity is quantified using the Shannon and Simpson indices based on 16S rRNA gene sequencing data. Differences between study phases (baseline, intervention, washout) are assessed using the Kruskal-Wallis test or Mann-Whitney U test, as appropriate, with a significance threshold of p < 0.05.

  2. Change in Salivary Microbial Beta Diversity (Community Structure)

    Time frame: Baseline (week 0), after intervention (week 12), and after washout (week 24).

    Evaluation of differences in microbial community composition between study phases. Calculated using Bray-Curtis dissimilarity on Centered Log-Ratio (CLR) normalized data and assessed via PERMANOVA to detect significant shifts (p < 0.05).

Secondary outcomes

  1. Differential Abundance of Salivary Bacterial Taxa

    Time frame: Baseline (week 0), after intervention (week 12), and after washout (week 24).

    Evaluation of shifts in the relative abundance of bacterial taxa (at phylum, family, genus, and species levels) using 16S rRNA sequencing (V3-V4 region). Statistical analysis is performed using DESeq2 and edgeR to identify significant differences between study phases, with multiple testing corrections applied using the Benjamini-Hochberg (FDR) method (FDR < 0.05).

  2. Changes in Predicted Antimicrobial Resistance Genes (ARGs)

    Time frame: Baseline (week 0), after intervention (week 12), and after washout (week 24).

    Functional prediction of the salivary microbiota conducted using PICRUSt2. Assessment of metabolic potential and antimicrobial resistance pathways with significance assessed using adjusted p-values (p-adj < 0.05).

  3. Gingival Index (GI) for Clinical Stability Assessment

    Time frame: Screening (Week -4) and after washout (Week 24).

    Assessment of gingival health using the Silness and Löe Gingival Index (scale 0-3). This measure is used to verify that participants maintain optimal oral health throughout the study. Eligibility is strictly limited to participants with a maximum GI of 0.143 at screening to ensure a healthy baseline and to monitor that no gingival inflammation occurs during the study period.

  4. Plaque Index (PI) for Clinical Stability Assessment

    Time frame: Screening (Week -4) and after washout (Week 24).

    Assessment of dental plaque accumulation using the Silness and Löe Plaque Index (scale 0-3). This measure is used to ensure consistent oral hygiene. Eligibility is strictly limited to participants with a maximum PI of 0.2 at screening. The index is monitored to confirm that participants adhere to the required oral hygiene protocol throughout the study.

Sponsors and collaborators

Lead sponsor

Masaryk University

Other

Registry information

Official study title

Study Protocol for 16S rRNA Sequencing of Healthy Oral Microbiota

Acronym: SMARCH

Important dates

Study start
2022
Primary completion
2022
Study completion
2022
First posted
May 6, 2026
Registry last updated
May 6, 2026

OpenTrials presents study information sourced from ClinicalTrials.gov. The official registry record should be consulted for the latest information.

View the official ClinicalTrials.gov record (opens in a new tab)

This listing is for discovery and informational purposes only. It is not medical advice, does not guarantee that a study is recruiting, and does not determine eligibility. Contact the study team and a qualified healthcare professional when considering participation.

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