Metagenomic Comparison of Two Popular Waterpipe Tobacco Brands Uncovers Differences in Relative Abundance of Pathogens and Antimicrobial Resistance Genes

Pathogenic microbes associated with waterpipe tobacco could contribute to adverse health effects among smokers. Waterpipe (hookah) tobacco may provide for an environment that encourages microbial growth and the mechanism of smoking waterpipe tobacco may not kill microbes that enter the smoke stream. Herein, we characterized the metagenome, humectant (propylene glycol and glycerol), and flavor content in two globally popular waterpipe tobacco brands: Moassel Salloum (MS) (unflavored) and Nakhla Double Apple (NDA) (flavored). The humectant content and flavor complexity of NDA were greater than those of MS, and the concentration of vanillin, which is antimicrobial, was significantly higher in NDA. The ratio of plant to microbial DNA showed that the microbial content of MS (20.3 +- 16.5 percent plant and 78.8 +- 16.5 percent microbial DNA) was significantly higher than NDA (78.3 +- 13.3 percent plant and 19.5 +- 13.5 percent microbial DNA, p = 0.0015). While MS harbored more bacteria, most were nonpathogenic with only three opportunistic pathogens strongly associated. In contrast, NDA, was dominated by microbiome associated genera and had five pathogens and twelve opportunistic pathogens significantly associated. This study is the first to compare differences in the metagenomes of flavored and unflavored waterpipe tobacco. Our results show that the microbial content is specific to the tobacco type, with the flavored NDA having higher proportions of human pathogens, than unflavored MS. However, MS harbored more antimicrobial resistant genes than NDA. Future research is needed to confirm that the difference between flavored and unflavored tobacco microbial communities is due to waterpipe tobacco ingredients.