Bacterial adhesion assay

Sterile 2 h nanowire discs and control discs (polished Ti) were placed into a 12-well microtitre plate and incubated with 2 ml bacterial suspension for 1 h or 18 h at 37 °C under static conditions. Plates were placed into a plastic container containing moist tissues to maintain humidity. After incubation, discs were rinsed 3 times in Tris-HCl buffer to remove non-adherent bacteria, and submerged in 1 ml Live/Dead^® BacLight™ bacterial viability stain (Invitrogen, as per manufacturers’ instructions) for 15 min at room temperature. Discs were then rinsed twice in Tris-HCl buffer, and submerged in 1 ml Tris-HCl buffer for visualisation by confocal microscopy. (Multilaser CLSM Leica SP511). Five Z-stack images per surface were taken and Volocity 6.3 cellular imaging and analysis software was used to count the number of cells with intact membranes (SYTO 9, green) and the number of cells with damaged membranes (propidium iodide, red). The average percentage of damaged cells was determined by dividing the damaged cells by the total number of cells on the surface (*100). All experiments were carried out in triplicate and repeated on separate occasions.