Role of branched-chain fatty acids in physiology and virulence of Staphylococcus aureus

7. Project Summary/Abstract While some bacterial infections are caused by a single species, we now know that most infections are polymicrobial in origin. We know that the activities of these species are often synergistic, as interactions between species enhances virulence, persistence, and tolerance to antibiotics. The end result is patient outcomes are typically worse in co-infection compared to mono-infection. Staphylococcus aureus is the leading cause of skin and soft tissue infections. The bacterium is also the most common organism isolated from chronic wounds and is frequently found with the opportunistic pathogens Enterococcus faecalis and Pseudomonas aeruginosa. Interactions between S. aureus and E. faecalis are relatively understudied. More broadly, our knowledge of the molecular mechanisms governing these interactions in chronic wounds is incomplete. S. aureus relies heavily on the de novo synthesis of branched-chain fatty acids for membrane biogenesis that cannot be obtained from the host during infection. These membrane fatty acids are essential for avoiding phase separation, protein aggregation, and cell death. They are also required to promote the activity of the Sae two-component system, a major regulator of S. aureus virulence. The PI’s laboratory recently discovered evidence for a new pathway specific to human-associated staphylococci for salvaging and synthesizing branched-chain fatty acids from the metabolic byproducts of co-infecting bacteria found in chronic wounds. The overall goals of this application are to i) determine the steps of this novel pathway and how Enterococcus faecalis helps S. aureus construct its membrane using this pathway during infection, and ii) determine how branched-chain fatty acids promote signaling via the Sae two-component system to increase pathogenicity. To accomplish these goals we will use penetrating genetic, biochemical, molecular biological and global approaches to interrogate the functionality of the proteins we are interested in. In addition we will use cutting-edge native mass spectrometry and spatial lipidomics approaches along with in vitro and in vivo murine models to define mechanisms. Detailed understanding of these processes is the important first step toward developing novel therapeutics to combat these complex infections.