The impact of bacterial oxidative stress responses on the ecology and pathogenicity of oral streptococci - PROJECT SUMMARY/ABSTRACT Bacterial diseases of the oral cavity (e.g. caries) are fundamentally polymicrobial, and understanding the inter- action between different species of bacteria is central to understanding how the oral microbiota is modulated. Among the most abundant bacteria in human saliva are Streptococcus species, some of which are dominant in healthy individuals and some of which are associated with or more abundant during diseased states. Hypothio- cyanous acid (HOSCN) is an antimicrobial oxidant abundant in saliva that is synthesized by mammalian heme peroxidases from H2O2 and thiocyanate (SCN-). Saliva contains the highest concentrations of SCN- in the human body (up to 3 mM), and the antimicrobial activity of the HOSCN produced by salivary lactoperoxidase (LPO) has been known for over 60 years. Early biochemical experiments showed that that commensal, health-associated oral streptococci (e.g. Streptococcus sanguinis) possess an enzymatic activity capable of degrading host-derived antimicrobial oxidants, while the caries-associated pathobiont S. mutans does not. We have recently identified a broadly-distributed bacterial HOSCN reductase (called RclA) that protects bacteria against the antimicrobial effects of HOSCN. S. sanguinis encodes an RclA homolog, while S. mutans does not. This now presents us with the opportunity to directly test the long-standing hypothesis that HOSCN reductase activity is important in the competition between health- and disease-associated oral streptococci. Doing so, how- ever, will also enable us to undertake a more careful examination and dissection of the role of LPO in modulating the oral microbiota and its potential use as an antimicrobial to modulate early dental plaque biofilm development. AIM 1. Identifying and characterizing HOSCN responses in oral streptococci Using genetically-tractable strains of S. sanguinis and S. mutans and a newly optimized defined artificial saliva medium, we will use molecular genetic, biochemical, and transcriptomic approaches to identify and test the impact of individual genes (e.g. rclA in S. sanguinis) and pathways that contribute to the ability of oral streptococci to respond to HOSCN. We will test the impact of these genes on the growth of HOSCN- stressed single and dual-species cultures both planktonically and in surface-associated biofilms. AIM 2. Characterizing the impact of LPO and its products on interactions between oral streptococci The extent to which LPO’s impact on oral bacteria is HOSCN-dependent under physiological conditions is unclear. We will characterize the impact of physiological concentrations of pro- and anti-oxidant products found in saliva on interactions between S. sanguinis and S. mutans in artificial saliva, both during planktonic growth and in surface-associated biofilms. The results of this work will be the first molecular-level understanding of how the pro- and anti-oxidant compo- nents of saliva drive growth and competition between cariogenic and non-cariogenic oral Streptococcus species.
