Exploring susceptibility of L-form bacteria to antimicrobial peptides

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BACKGROUND Antimicrobial resistance (AMR) is considered by the World Health Organization as one of the top 10 global public health threats. L-form switching is a mechanism, that allows bacteria (including important pathogens such as Escherichia coli (E. coli), Staphylococcus aureus (S. aureus) or Pseudomonas spp.) to temporarily lose their cell wall. L-form bacteria are, thus, completely resistant to antibiotics, which target cell wall, such as penicillins , and are implicated in recurrence of many infectious diseases including urinary tract infections (UTIs), sepsis or foodborne infections. While tollerant to cell wall-targeting agents, L-forms should retain susceptibility to other types on antibiotics, for example those targeting membranes. OBJECTIVES AND NOVELTYAntimicrobial peptides (AMPs) are naturally occurring small peptides, which are part of the immune system of various organisms, including humans. These peptides most commonly target pathogen membranes, and AMPs such as nisin were shown to be effective against L-forms in vitro. In this PhD project, we will investigate the susceptibility of E. coli and S. aureus L-forms to various antimicrobial peptides in comparison to their walled counterparts. Using cell culture infection models (including cell lines and organoids) we will investigate for the first time if antimicrobial peptides can be used to eliminate L-form bacteria in the context of host tissue, therefore potentially contributing to prevention of recurrent infections. TIMELINESSAMR contributes to ~1.3 million deaths per year globally, with this number estimated to raise to 10 million by 2050, alongside a cumulative cost of $100 trillion, if no action is taken. This work will contribute to a better understanding of mechanisms of antibiotic evasion, more efficient use of the existing, and potential development of new treatments against bacterial infections. EXPERIMENTAL APPROACHEffectiveness of various AMPs (including cathelicidins, defensins, cecropin or synthetic ones) against E. coli and S. aureus L-forms will be tested using killing assays and high-end microscopy. We will systematically test if combining cell wall-targeting antibiotics with antimicrobial peptides might constitute a more effective treatment than each of the agents used alone, and precisely what proportion of each of the agents is most effective. We will use cell lines such as RAW 264.7, THP-1 macrophages and intestinal murine organoids combined with advanced fluorescence microscopy to test the effectiveness of AMPs against L-forms in a context of a tissue.The aim of this DTP is to add value by building synergies, developing new collaborations, and opening up new areas of research. How does this project add value, over and above what could be achieved with a standard studentship? (Maximum 100 words) This research project is a new collaboration between two Newcastle University based research groups. The primary supervisor Katarzyna Mickiewicz focuses on the role of L-form bacteria in disease and the secondary supervisor Henrik Strahl von Schulten on the mode of action of cell envelope targeting antibiotics. These are two complementary areas of expertise, which will accelerate the project and provide optimal training opportunities for the student to deliver on the project, which would not be possible for only one of these supervisors to achieve. 40% contribution from the senior second supervisor who will provide mentorship as well as the expertise, will be invaluable to the junior primary supervisor (60% contribution). Publications