Introduction: The microbiota-gut-brain axis is a bidirectional pathway that has recently been shown to influences brain and behavior. Moreover, gut microbiota have been implicated in several psychiatric conditions such as depression and anxiety. To date, minimal research has investigated gut microbiota in addiction including alcohol abuse. Furthermore, the effect of vaporized ethanol administration on the gut microbiota is unknown. Methods: Adult male C57BL/6J mice were exposed to 4 weeks of either chronic intermittent vaporized ethanol (Vapor EtOH, N=10) or air (Vapor Control, N=10). A previously described vapor inhalation procedure was employed for the test group (Holmes et al., 2012; Lopez & Becker, 2005). In short, test mice received IP injections of 1.5g/kg of 20% EtOH (v/v) with 71.6 mg/kg alcohol dehydrogenase (ADH) inhibitor pyrazole (Sigma, St. Louis, MO, USA) prior to vapor ethanol exposure (16 hours/day, 5 days/week). Control group received IP injections of saline with 68.1 mg/kg ADH inhibitor pyrazole and only fresh air in vapor chambers. Fecal samples were collected at the end of the experiment and 16S prokaryotic DNA was sequenced. Bioinformatic analysis of sequenced 16S Operational Taxonomic Units (OTUs) assessed bacterial composition. Results: Phylogenetic measures of alpha (p<0.05) and beta (ANOSIM, p=0.001) diversity revealed significant differences between groups, with alpha diversity decreased in the Vapor EtOH group. Genus level bacterial composition showed significant differences (ANOSIM, p=0.002) between groups with the most significant changes in Alistipes (Kruskal-Wallis, p<0.00024) and Clostridium IV and XIVb (Kruskal-Wallis, p<0.00045). Conclusion: These results demonstrate that chronic vapor alcohol exposure is capable of significantly altering the gut microbiota in mice. These findings align with previous findings of microbiota changes associated to liver disease, inflammation, and psychological distress (Hartmann et al., 2015; Leclercq et al., 2014; Mutlu et al., 2012).