A journey into the biofilms of Cupriavidus metallidurans CH34: substrate adaptation and response to silver nanoparticles and ionizing radiation

A biofilm is a structured microbial community often featuring different species of microorganisms embedded in a self-produced extracellular matrix and adhering to different types of surfaces. Biofilms may develop in various environments and are problematic for many industrial processes. In the medical sector, every year biofilms are responsible for a large number of premature deaths. Consequently, the formation and elimination of biofilms are widely studied topics in microbiology. However, despite years of research, biofilm biology is not fully understood and treatments to remove biofilms are not yet capable to prevent their development. Biofilms can be very complex and highly adaptive, reducing the efficacy of numerous antimicrobial treatments. The main objective of this PhD work was to investigate the adaptive character and the resilience of biofilms produced by the oligotrophic and metal-resistant bacterium Cupriavidus metallidurans CH34. Such a bacterium can form biofilms in water systems and persists in highly controlled environments such as the water recycling loop in the International Space Station. In this thesis, biofilms of this species were studied in different situations, on various types of surfaces exposed to physical and/or chemical antimicrobial treatments. The manuscript is composed of six chapters. Chapter 1 is a general introduction in which general concepts about biofilm formation and resilience are reviewed. The objectives of the thesis are explained in Chapter 2, then in Chapter 3, the interactions between C. metallidurans CH34 and industrially relevant surfaces such as the TiAl6V4 and SS316 alloys, Teflon and glass were investigated. In Chapter 4, we explored the response of this bacterium to aluminium and in Chaper 5 we studied the cellular and molecular interactions of C. metallidurans CH34 to an exposure to antimicrobial silver nanoparticles (AgNPs). In Chapter 6, biofilms were exposed to a combined treatment based on AgNPs and ionizing radiation. Finally, in Chapter 7, we present a comprehensive discussion and synthesis of our findings. Our results show that growth and biofilm formation of C. metallidurans CH34 are influenced by the physico-chemical properties of the surface. Biofilms of this bacterium were highly resistant to antimicrobial silver nanoparticles compared to planktonic cells. We demonstrate that their resilience is age-dependent, with older biofilms being less sensitive than younger ones. The combination of AgNPs to ionizing radiation affected cell survival only when biofilms were young enough while more mature colonies were mostly unaffected. The proteomic analysis revealed a default response of C. metallidurans biofilms to silver nanoparticles. This response was independent of the maturation stage and involved key molecular pathways such as siderophores synthesis, energy metabolism, the activity of electron transport chains and the production of metal resistance proteins.