Transient dormancy and recovery as a strategy for antimicrobial escape: Can the escape route be blocked? – StaphEscape

Staphylococcus aureus is a major cause of hospital- and community- acquired infections largely due to the high rate of antibiotic resistance, making development of new treatments a major socio-economic objective. One target actively developed over several years is the fatty acid (FA) synthesis (FASII) pathway. Despite widespread publicity of this strategy, our studies uncovered a flaw that could deeply compromise use of this class of antibiotics: we showed that numerous pathogenic Firmicutes use exogenous FAs, as are available in the host, to constitute their membranes, making FASII nonessential for growth. This was fully demonstrated using Streptococcus agalactiae as a genetic and virulence model. However, the status of S. aureus remained controversial, based on data from laboratories and pharmaceutical companies developing antimicrobials targeting FASII. A different response to FASII inhibitors was suggested to correlate with a need for auto-synthesized branched chain FA, which are not present in host organs. This incited us to perform a deep investigation of FASII utilization in S. aureus, which constitutes the foundation for the present project. We discovered two main strategies by which S. aureus overrides FASII inhibition:
1- adaptive mutations in FASII initiation enzymes, which facilitate exogenous FA incorporation; and
2- adaptation without mutation, in which S. aureus undergoes transient "dormancy". During dormancy, cells assimilate exogenous FA but do not multiply; after several hours, growth resumes with sustained use of exogenous FA to bypass FASII inhibitors.
Mutational adaptation is stable, whereas non-mutational adaptation is reversible.
A key finding was that FASII bypass by both mechanisms occurs in hospital isolates, showing the medical relevance of S. aureus dormancy escape. Our data change current views on staphylococcal membrane requirements, by showing their resiliency when fatty acids are totally altered. Our results underline the importance of conditions used when testing drug efficacy, and indicate that anti-FASII sensitivity in vitro does not assure efficacy in the FA-rich host.
Our main project goal is to characterize this newly discovered "dormant" state in S. aureus, which involves a transient cell division arrest, expression changes, marked cell wall thickening and membrane deformation, and exclusive use of exogenous FA for membrane synthesis. We will identify signals that trigger S. aureus escape from FASII-block-induced dormancy. Approaches include mutant library screening, and assessment of transcriptional, protein, and morphological modifications during dormancy exit. We will pursue our working hypothesis, that membrane/cell-wall crosstalk generates signals for S. aureus to resume growth post-dormancy.
Recent publications show success using a combinatorial antibiotic approach to eliminate pathogens when single antibiotics were ineffective. Our second goal is to ask whether the drastic membrane changes during FASII bypass sensitize bacteria to second antimicrobials. We will exploit our results on mutant screening, and assessment of transcriptional and protein changes during dormancy to select and screen membrane- and cell-wall- targeted antibiotics that cooperate with FASII inhibitors, to cut off the dormancy exit escape route and knock out S. aureus growth. This approach should provide cognitive knowledge and could lead to a combinatorial antimicrobial treatment of S. aureus infection.
Pertinence of the project to the Call: A novel biological state in S. aureus, dormancy due to membrane synthesis arrest, will be characterized. S. aureus dormancy and escape are likely to occur in the host and might explain cases of persistent infection. The project aims to confront the current crisis in treating infections due to multi-resistant S. aureus, by screening antimicrobials for synergistic activity when combined with an anti-FASII.