Figure 2.

Calcite mineralization occurs in distinct areas within the biofilms

(A–D). Biofilm cells closely associated with prismatic mineral structures, as visualized by Scanning Electron Microscopy (SEM) image of 8 (A and B) and 10-day-old (C and D) B. subtilis biofilm colonies grown with calcium. (A and C) - secondary mode; (B and D) upper panel - backscattering mode. Magnification – X25000, scale bar – 1 μm. A representative field (out of n = 5 fields, from six experiments) is shown. (B and D) Lower panel - EDX analysis of boxed areas, indicating that mineral structures are rich in calcium. Ca – calcium-rich, Cnt – control.

(E) 3D reconstruction of microCT X-ray analysis revealing 3D mineral distribution in a 5-day old B. subtilis colony grown with calcium. Color indicates intensity, with green representing the densest mineral. Scale bar – 0.2 mm. A representative image (out of n = 3 experiments) is shown.

(F) FTIR spectra of bleached B. subtilis biofilm colonies, displaying vibrations characteristic of calcite (crystalline calcium carbonate).