Fibrin Networks Infected by Staphylococcal Biofilms: Mechanics, Structure and Instability

Abstract

In this dissertation, we study a new model of medical device infection – that of a fibrin infected with the common blood borne pathogen Staphylococcus epidermidis, which is shown to influence fibrin mechanically and structurally on both microscopic and macroscopic scales. S. epidermidis present during clot formation produces a visibly disorganized microstructure that increases clot stiffness and triggers mechanical instability over time.

We also noticed that the capacity for S. epidermidis to modulate fibrin formation kinetics, mechanics and microstructure is a function of bacterial growth phase. We continued to find that the bacteria-fibrinogen interaction is also growth phase dependent: stationary-phase (biofilm-like) S. epidermidis has an increased and stronger adhesion to fibrinogen as compared to exponential-phase (planktonic) ones. Furthermore, the gene expression for SdrG – the protein that adheres S. epidermidis cells to fibrinogen was significantly increased in the stationary phase.

We extended our understanding on the effect of S. epidermidis on fibrin by investigating a specific protein – IcaB, a cell surface-attached protein that has a key role in staphylococcal biofilm formation and immune evasion. It deacetylates and introduces positive charge to polysaccharide intercellular adhesin (PIA) molecules; these charges facilitate adhesion of bacterial cells within a biofilm cluster. Using an icaB deletion strain, we evaluated the effects of PIA deacetylation on staphylococcal interaction with a fibrin network and the resulting structural and mechanical changes to the network. We find that the ∆icaB strain degrades fibrin networks to a higher degree than the wild-type, both structurally and mechanically. Structurally, the connected branches in a fibrin network infected with the ∆icaB strain become disconnected patches. Mechanically, a fibrin network infected with the ∆icaB strain becomes more liquid-like and deformative. Indeed, the ∆icaB produce more sspA, a protease that putatively degrades fibrin. We further demonstrate that the ∆icaB cells, which are more diffusive than the wild type cells, co-localize with the fibrin fibers. This co-localization is correlated with fibrin network degradation.

In our study of infected fibrin, we noticed some limitations in the mechanical characterization using rheometers, e.g. the challenge of delivering additives in situ. This inspired us to develop a rheometer tooling comprised of a porous hydrogel. The tooling allows additives to be dosed through a rheometer plate or cone, and the consequent effect of these additives on a material’s mechanical response measured. We demonstrated that the tooling can be used to study the kinetics of material property variation due to the diffusion of molecular additives into the soft material though the hydrogel plate.

Overall, our understanding of fibrin infected with S. epidermidis aids in understanding of medical device infections and infection-induced thromboembolism.