0.025 μm filters

The supernatant of 24h old mono- and mixed species biofilms was isolated, spun down (3000 g, 10’), and filtrated using 0.025 μM filters (Millipore). The cell-free supernatant assay used the same methods for biofilm growth as described in the in the section ‘Growth conditions’, except that the S1 strain was inoculated in media containing 30% cell-free supernatant.

For assessment of uranium concentration in the soluble and particulate fractions within the replicate microcosms, 2 ml of culture at each time point were filtered through 0.025 μm filters (Millipore Corp, Billerica, MA, USA) in an anaerobic chamber (Coy Laboratory Products, Grass Lakes, MI, USA). This soluble fraction (filtrate) was acidified by addition of 0.2 ml nitric acid (70%) and stored in glass vials until analysis. The particulate fraction (filter and the uranium adhering to the walls of the culture vials) were collected by dissolving the filter in 3 ml nitric acid and heating 250°C for 3 hours, or by rinsing the empty culture vials with 3 ml of nitric acid and vortexing for 2 minutes. The uranium concentration was determined by iCAPQ ICP-MS (Thermo Fisher Scientific Inc., Waltham, MA, USA) using Indium as an internal standard. Three iterations per sample were analyzed and RSD percentages averaged 3.6%. A commercial standard (High Purity Standards, Charleston, SC, USA) was used to generate a standard curve for uranium mass based on the instrument signal (counts per second; r² = 0.9955).