Heterogeneous lineage-specific arginine deiminase expression within dental microbiome species

Arginine catabolism by the bacterial arginine deiminase system (ADS) has anticariogenic properties through the production of ammonia, which modulates the pH of the oral environment. Given the potential protective capacity of the ADS pathway, the exploitation of ADScompetent oral microbes through pre or probiotic applications is a promising therapeutic target to prevent tooth decay. To date, most investigations of the ADS in the oral cavity and its relation to caries have focused on indirect measures of activity or on specific bacterial groups, yet the pervasiveness and rate of expression of the ADS operon in diverse mixed microbial communities in oral health and disease remain an open question. Here, we use a multivariate approach, combining ultradeep metatranscriptomic sequencing with paired metataxonomic and in vitro citrulline quantification to characterize the microbial community and ADS operon expression in healthy and latestage cavitated teeth. While ADS activity is higher in healthy teeth, we identify multiple bacterial lineages with upregulated ADS activity on cavitated teeth that are distinct from those found on healthy teeth using both referencebased mapping and de novo assembly methods. Our dual metataxonomic and metatranscriptomic approach demonstrates the importance of species abundance for gene expression data interpretation and that patterns of differential expression can be skewed by lowabundance groups. Finally, we identify several potential candidate probiotic bacterial lineages within species that may be useful therapeutic targets for the prevention of tooth decay and propose that the development of a strainspecific, mixedmicrobial probiotic may be a beneficial approach given the heterogeneity of taxa identified here across health groups. IMPORTANCE Tooth decay is the most common preventable chronic disease, affecting more than two billion people globally. The development of caries on teeth is primarily a consequence of acid production by cariogenic bacteria that inhabit the plaque microbiome. Other bacterial strains in the oral cavity may suppress or prevent tooth decay by producing ammonia as a byproduct of the arginine deiminase metabolic pathway, increasing the pH of the plaque biofilm. While the benefits of arginine metabolism on oral health have been extensively documented in specific bacterial groups, the prevalence and consistency of arginine deiminase system (ADS) activity among oral bacteria in a community context remain an open question. In the current study, we use a multiomics approach to document the pervasiveness of the expression of the ADS operon in both health and disease to better understand the conditions in which ADS activity may prevent tooth decay.