Metabolic Commensalism and Competition in a Two-Species Microbial Consortium

SCLM micrographs showing the structural relationships between Acinetobacter strain C6 and P. putida R1 cells with low and high activity, respectively, in a mixed biofilm consortium which was supplied with 0.5 mM benzyl alcohol as the sole carbon source. At days 1 (A), 2 (B), 3 (C and E), and 6 (D and F) after inoculation, biofilms were embedded and hybridized. P. putida R1 and Acinetobacter strain C6 were hybridized with PP986 labeled with CY5 (blue) and ACN449 labeled with CY3 (red), respectively. The active P. putida R1 cells were monitored as cells emitting green fluorescence due to the rrnBP1-gfp[AGA] fusion inserted in the chromosome of P. putida R1. These cells appear as cyan due tothe combination of green (GFP) and blue (hybridization). For each panel similar images were collected from at least five independent biofilm experiments. Panels A and B are representative of the biofilm structures observed at day 1 and 2. Panels C and D are examples of the large Acinetobacter strain C6 microcolonies that developed after 3 days (C) and which later were overgrown by P. putida R1 (D). Panels E and F are examples of Acinetobacter strain C6 microcolonies (arrow) that were established in the upper part of P. putida R1 cell clusters after 2 to 3 days (E), which resulted in production of large macrostructures of associated P. putida R1 and Acinetobacter strain C6 cells (F). All x-y plots are presented as extended-focus images. Shown to the right and above the x-y plots are vertical sections through the biofilm collected at the positions indicated by the white triangles. The arrow indicates the direction of flow. Bars, 20 μm. Note that the green fluorescent intensities on the images have been amplified to give the best differentiation between P. putida R1 cells with low and high activity in each image. Thus, the intensity levels in the different images cannot be directly compared.