Within- and Between-Visit Variability and Reproducibility of the Oral Microbiome in Young Adults with Alcohol Use Disorder.

BACKGROUND: The oral microbiome has emerged as a potential biomarker and pharmacological target in alcohol use disorder (AUD) due to its associations with alcohol use and related biological processes. However, its temporal variability and reproducibility remain poorly understood, limiting its utility. METHODS: Saliva samples were obtained from participants enrolled in a randomized controlled trial of young adults with AUD. Temporal variability and reproducibility were evaluated using within-visit samples (∼4 hours apart; before and after sesame oil placebo and standardized snack) and between visits (∼25 days; pre-treatment). Alpha diversity, beta diversity, differential abundance, and intraclass correlation coefficients (ICCs) were calculated at the genus and species levels. RESULTS: Significant within-visit differences were observed in alpha diversity, beta diversity, and the abundance of several genera and species. Snack type and time since last alcohol use explained comparable or greater variance in microbial composition than within-visit timepoint (4-6%). Although reproducibility of alpha diversity within-visit was generally low, most genera (70.3%) and species (74.6%) demonstrated at least moderate reproducibility. In contrast, no detectable systematic between-visit differences were observed in diversity or taxon abundance, and reproducibility was generally moderate for alpha diversity measures and most genera (67.6%) and species (72.2%). CONCLUSIONS: Despite group-level microbial shifts within-visit, individual-level microbial features were generally reproducible both within (∼4 hours) and between (∼25 days) visits. No systematic group-level differences were detected between-visits.These findings support the use of the oral microbiome in AUD research while emphasizing the importance of longitudinal designs and accounting for recent exposures.