Effects of chlorhexidine and a polyherbal mouthwash on the oral microbiome and user satisfaction: a randomized controlled trial

Abstract

Objective: This study aimed to comparatively evaluate the effects of polyherbal, chlorhexidine (CHX), and normal saline (NCC) mouthwashes on oral microbiome composition, microbial metabolic pathways, and patient-reported outcomes, with the goal of assessing the potential of polyherbal mouthwash as a natural alternative to CHX for managing gingivitis, while aiming to minimize CHX-associated microbial dysbiosis.

Methods: A randomized, double-blind trial was conducted among 27 patients with gingivitis, randomly assigned to polyherbal, CHX, or NCC mouthwash groups. Participants used the assigned mouthwash twice daily for 7 days. Saliva samples were analyzed via 16 S rRNA sequencing to assess microbial diversity (alpha and beta) and taxonomic composition. Differential taxa and pathways were identified using FDR-corrected Wilcoxon tests and log₂ fold-change analysis. Patient-reported outcomes were evaluated using numerical rating scales (NRS).

Results: The polyherbal mouthwash demonstrated significantly higher patient acceptability than CHX and NCC, with superior scores in taste, smell, texture, moisture retention, overall satisfaction, and willingness for continued use (p < 0.05). While CHX exhibited strong antimicrobial activity, its alcohol-containing formulation markedly disrupted microbial diversity, increased potentially dysbiotic genera (Streptococcus, Porphyromonas), and altered 23 metabolic pathways associated with dysbiosis. In contrast, the polyherbal mouthwash-which also contained cetylpyridinium chloride (CPC)- selectively increased beneficial genera (Amnipila, Absconditabacteriales [SR1], Peptostreptococcus), preserved overall microbial diversity, and modulated only two pathways, notably upregulating L-isoleucine biosynthesis.

Conclusion: The polyherbal mouthwash represents a promising alternative to CHX, demonstrating selective microbial modulation, preservation of microbiome stability, and enhanced patient acceptability. These findings support its potential integration into routine oral healthcare as a microbiome-friendly and patient-acceptable solution. Further studies are warranted to evaluate CPC-free polyherbal formulations across larger and more diverse populations over extended durations.

Keywords: 16S rRNA sequencing; Chlorhexidine alternative; Gingivitis; Gingivitis-associated dysbiosis; Oral microbiome; Patient satisfaction; Polyherbal mouthwash; Prevotella.