Studying the interaction mechanism of Staphylococcus aureus with human skin cells using atomic force microscopy

Modern health care systems have to deal with an expanding number of nosocomial infections due to increased antimicrobial resistance, especially linked to meticillin-resistant Staphylococcus aureus based biofilms. Therefore, understanding the interaction mechanisms occurring between bacterial cells and their host during the infection process is a major challenge in microbiology. In particular, the bacterial cell wall of bacteria is composed of structurally and functionally diverse molecules, called adhesins, partially involved in the adhesion processes. The aim of this master thesis is to gain novel insights into the nanoscale interaction forces between a versatile pathogen implicated in a wide variety of human diseases, S.aureus and human skin. Specifically, we wish to better understand the role of bacterial adhesins and skin ligands in the bacterial-skin adhesion, taking advantage of Atomic Force Microscopy (AFM) techniques and of a close collaboration with Profs. T. Foster and J. Geoghegan (Trinity College, Dublin) who provided most of the biological material. Single Cell Force Spectroscopy (SCFS), where a single S.aureus cell is immobilized on the AFM cantilever was used as a probe to quantify and characterize the molecular forces driving the adhesion of S.aureus to corneocytes. In addition, we developed a new method combining nanoscale multiparametric imaging with the use of single bacterial probes in order to simultaneously map the topography and bacterial-binding properties of corneocytes at high spatiotemporal resolution. The interactions between S.aureus Newman cells and healthy human corneocytes resulted in strong adhesion forces of about ~500 pN. Control experiments using a S.aureus mutant strain lacking the sortase A, an enzyme responsible for the anchorage of adhesins on the cell’s surface, showed no adhesion, thus confirming the involvement of specific adhesin-ligand complexes in the adhesion process. Using smooth protein-coated surfaces, we also showed that S.aureus Newman cells, unlike mutant cells, strongly bind (~1400 pN) to two proteins expressed on the skin’s surface, keratin and loricrin. In addition to validate SCFS and multiparametric analyses regarding the high corneocyte’s roughness, we therefore suggest that keratin and loricrin binding may contribute to skin colonization and infection by S.aureus. Identifying specific adhesin/ligand complexes and potential inhibitors will constitute the next step in the development of anti-adhesive strategies. Applicable to a wide variety of microbes and skins, our novel methodology offers exciting prospects in nanomedicine for understanding the molecular details of skin colonization and disorders, such as atopic dermatitis.