The role of purine biosynthesis and stringent response in persistent MRSA endovascular infections

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ABSTRACT Staphylococcus aureus is the most common cause of life-threatening endovascular infection, including infective endocarditis (IE) and bacteremia. Despite the use of gold-standard antibiotics, morbidity and mortality associated with these syndromes remain unacceptably high. Emergence of methicillin-resistant S. aureus (MRSA), high rates of vancomycin (VAN) clinical failures, and rising daptomycin (DAP) resistance further emphasize this public health threat. Persistent MRSA bacteremia (PB), defined as  7 days of positive blood cultures despite appropriate antibiotic therapy, is a very worrisome sub-set of these infections. A particularly problematic metric is that PB strains are deemed “susceptible” in vitro to VAN and DAP by standard CLSI breakpoints, yet, persist in vivo despite appropriate use of these antibiotics. Therefore, PB outcomes present a unique variant of traditional antibiotic “resistance” mechanisms and significant therapeutic challenge to the medical community. Understanding the relevant molecular mechanisms of PB is essential to develop novel strategies to predict and successfully treat PB patients. Our Preliminary Data showed that the PB outcomes are likely to be multifactorial on both phenotypic and genotypic levels. Most interestingly, we have shown the impact of purine biosynthesis and stringent responses on the PB outcomes using both clinical MRSA isolates and laboratory isogenic strain sets. Based on our extensive Preliminary Data, we hypothesize that distinct regulatory cascades activated by purine biosynthesis perturbations and the stringent responses play an important role in the PB outcome. Therefore, in this proposal, we will: i) further define the impact of purine biosynthesis in the PB outcome both phenotypically and genotypically by constructing purF deletion mutations in prototypic clinical PB strains; ii) assess the role of purine biosynthesis in the stringent response as it relates to the PB outcome; iii) determine the relationship between purine biosynthesis and biofilm formation; and iv) validate the role of purine biosynthesis in the PB outcome in vivo in an experimental IE model. These studies will significantly advance our understanding on the mechanisms of persistent MRSA endovascular infection and identify unique signatures for new anti-MRSA agents or strategies to treat clinical infections featuring the PB outcome.