The effect of daily usage of Listerine Cool Mint mouthwash on the oropharyngeal microbiome: a substudy of the PReGo trial

Listerine is a bactericidal mouthwash widely used to prevent oral health problems such as dental plaque and gingivitis. However, whether it promotes or undermines a healthy oral microbiome is unclear. We hypothesized that the daily use of Listerine Cool Mint would have a significant impact on the oropharyngeal microbiome. We aimed to assess if daily usage of Listerine Cool Mint influenced the composition of the pharyngeal microbiome. The current microbiome substudy is part of the Preventing Resistance in Gonorrhoea trial. This was a double-blind single-centre, crossover, randomized controlled trial of antibacterial versus placebo mouthwash to reduce the incidence of gonorrhoea/chlamydia/syphilis in men who have sex with men (MSM) taking HIV pre-exposure prophylaxis (PrEP). Fifty-nine MSM taking HIV PrEP were enrolled. In this crossover trial, participants received 3 months of daily Listerine followed by 3 months of placebo mouthwash or vice versa. Oropharyngeal swabs were taken at baseline and after 3 months use of each mouthwash. DNA was extracted for shotgun metagenomic sequencing (Illumina Inc.). Non-host reads were taxonomically classified with MiniKraken and Bracken. The alpha and beta diversity indices were compared between baseline and after each mouthwash use. Differentially abundant bacterial taxa were identified using ANOVA-like differential expression analysis. was the most abundant genus in most samples ( = 103, 61.7 %) with a median relative abundance of 31.5% (IQR 20.6-44.8), followed by [13.5% (IQR 4.8-22.6)] and [10.0% (IQR 4.0-16.8)]. Compared to baseline, the composition of the oral microbiome at the genus level (beta diversity) was significantly different after 3 months of Listerine ( = 0.006, pseudo- = 2.29) or placebo ( = 0.003, pseudo- = 2.49, permutational multivariate analysis of variance) use. and were significantly more abundant after Listerine use compared to baseline. Listerine use was associated with an increased abundance of common oral opportunistic bacteria previously reported to be enriched in periodontal diseases, oesophageal and colorectal cancer, and systemic diseases. These findings suggest that the regular use of Listerine mouthwash should be carefully considered.