Oral biofilms exposure to chlorhexidine results in altered microbial composition and metabolic profile

Abstract

Oral diseases (e.g., dental caries, periodontitis) are developed when the healthy oral microbiome is imbalanced allowing the increase of pathobiont strains. Common practice to prevent or treat such diseases is the use of antiseptics, like chlorhexidine. However, the impact of these antiseptics on the composition and metabolic activity of the oral microbiome is poorly addressed. Using two types of oral biofilms-a 14-species community (more controllable) and human tongue microbiota (more representative)-the impact of short-term chlorhexidine exposure was explored in-depth. In both models, oral biofilms treated with chlorhexidine exhibited a pattern of inactivation (>3 log units) and fast regrowth to the initial bacterial concentrations. Moreover, the chlorhexidine treatment induced profound shifts in microbiota composition and metabolic activity. In some cases, disease associated traits were increased (such as higher abundance of pathobiont strains or shift in high lactate production). Our results highlight the need for alternative treatments that selectively target the disease-associated bacteria in the biofilm without targeting the commensal microorganisms.

Fig. 1. Biofilms exhibited a pattern of kill and regrowth after treatment with 0.12% CHX.

Concentrations of live, damaged, and dead cells for a. 14-species biofilms. Points are the average of four replicates and error bars represent the standard deviation. b For tongue deriving microbiota biofilm from four individuals (donors). The red line is the concentration of CHX treated biofilms, purple/blue for the non-treated control biofilms. The vertical red lines represent the points of treatment.

Fig. 2. The bacterial composition of the 14-strains biofilms over the course of treatment.

Relative abundance of the individual strains in 14-strain biofilms every 24 h and before the next CHX treatment. The percentages are the average of four replicates.

Fig. 3. nMDS plot representing the β-diversity of the tongue deriving microbiota biofilm samples based on Bray Curtis dissimilarity index.

Samples deriving from different donors have a different color, circles are for CHX treated samples, while triangles symbolize non-treated control samples. The number indicates the time of the sampling in hours.

Fig. 4. The bacterial composition of oral biofilms over the course of treatment.

Relative abundance of the 20 most abundant OTUs for the tongue derived microbiota biofilms every 24 h and before the next CHX treatment.

Fig. 5. The effect of CHX treatment on the metabolic activity of the in vitro oral biofilms.

The organic acid production or consumption by the 14-strains (average of four replicates) and tongue-deriving microbiota biofilms in between the daily treatments with 0.12% CHX.