Abstract
Biomaterials-associated infections are primarily initiated by the adhesion of microorganisms on the biomaterial surfaces and subsequent biofilm formation. Understanding the fundamental microbial adhesion mechanisms and biofilm development is crucial for developing strategies to prevent such infections. Suitable in vitro systems for biofilm cultivation and bacterial adhesion at controllable, constant and reproducible conditions are indispensable. This study aimed (i) to modify the previously described constant-depth film fermenter for the reproducible cultivation of biofilms at non-depth-restricted, constant and low shear conditions and (ii) to use this system to elucidate bacterial adhesion kinetics on different biomaterials, focusing on biomaterials surface nanoroughness and hydrophobicity. Chemostat-grown Escherichia coli were used for biofilm cultivation on titanium oxide and investigating bacterial adhesion over time on titanium oxide, poly(styrene), poly(tetrafluoroethylene) and glass. Using chemostat-grown microbial cells (single-species continuous culture) minimized variations between the biofilms cultivated during different experimental runs. Bacterial adhesion on biomaterials comprised an initial lag-phase I followed by a fast adhesion phase II and a phase of saturation III. With increasing biomaterials surface nanoroughness and increasing hydrophobicity, adhesion rates increased during phases I and II. The influence of materials surface hydrophobicity seemed to exceed that of nanoroughness during the lag-phase I, whereas it was vice versa during adhesion phase II. This study introduces the non-constant-depth film fermenter in combination with a chemostat culture to allow for a controlled approach to reproducibly cultivate biofilms and to investigate bacterial adhesion kinetics at constant and low shear conditions. The findings will support developing and adequate testing of biomaterials surface modifications eventually preventing biomaterial-associated infections.
Highlights
Modern medicine increasingly uses biomaterials for implant purposes, e.g., to restore human body functions
Cultivation and Investigation of Microbial Adhesion In this study, the constant-depth film fermenter (CDFF) was modified for the cultivation of nonz-restricted biofilms (Fig. 2) to meet the requirements for investigating biofilm formation at low shear conditions, with non-unidirectional fluid flow and with constant conditions throughout the biofilm reactor
This study introduced the Non-constant-depth Film Fermenter (nCDFF) for cultivating non-zrestricted biofilms under constant and low shear conditions
Summary
Modern medicine increasingly uses biomaterials for implant purposes, e.g., to restore human body functions Such implants can lead to infections by supporting adhesion of microorganisms and subsequent biofilm formation on the biomaterials surfaces [1,2]. The most frequently used microtiter well plate systems allow for fast and high sample throughput and do not require specialized laboratory equipment [4]. Many researchers use microbial cells that were washed, centrifuged and resuspended in buffer solutions to prevent growth during adhesion studies. These methods, lead to weakening or damaging the cells, which significantly affects bacterial adhesion [6,7,8], and altogether might preclude meaningful in vitro findings
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