Polysaccharide metabolism regulates structural colour in bacterial colonies

Abstract

The brightest colours in nature often originate from the interaction of light with materials structured at the nanoscale. Different organisms produce such coloration with a wide variety of materials and architectures. In the case of bacterial colonies, structural colours stem for the periodic organization of the cells within the colony, and while considerable efforts have been spent on elucidating the mechanisms responsible for such coloration, the biochemical processes determining the development of this effect have not been explored. Here, we study the influence of nutrients on the organization of cells from the structurally coloured bacteria Flavobacterium strain IR1. By analysing the optical properties of the colonies grown with and without specific polysaccharides, we found that the highly ordered organization of the cells can be altered by the presence of fucoidans. Additionally, by comparing the organization of the wild-type strain with mutants grown in different nutrient conditions, we deduced that this regulation of cell ordering is linked to a specific region of the IR1 chromosome. This region encodes a mechanism for the uptake and metabolism of polysaccharides, including a polysaccharide utilization locus (PUL operon) that appears specific to fucoidan, providing new insight into the biochemical pathways regulating structural colour in bacteria.

Keywords: biochemical pathway; fucoidan; metabolism; polysaccharides; regulation; structurally coloured bacteria.

Figure 1.

Colour development of IR1 colonies throughout their lifespan under different nutrient conditions: an ASWB plate without any added polysaccharides, an ASWBF plate with added fucoidan and an ASWBC plate with added κ-carrageenan. The images show the centre of each colony, at the location where the initial cell suspension was deposited. The insets show the edge of each colony, at the same scale as the main images.

Figure 2.

(a) Angular resolved spectral reflection in scattering mode for IR1 grown under three different nutrient conditions on day 1 of their growth. The illumination angle is kept at −60° for all measurements. Due to a limitation of the set-up, light cannot be detected at the angle of illumination, creating the dark band observed at −60°. Reflected light intensity is normalized against a white diffuser, and represented by a heat map, with blue low intensity and yellow high intensity. The angular range inside the white dotted lines contains the specular reflection angle (+60°), where the reflectance is much stronger than at other angles. We therefore show the reflectance in this angle range divided by a factor of 300 so that it can be displayed without saturating the colour scale. The solid white lines show a fit of the grating equation to the diffraction peaks, giving us a value for the lattice constant as shown in (b). (b) Development of the lattice constant for IR1 colonies grown under different nutrient conditions. (c) Schematic showing how the tilt angle of the crystalline domains determines the angle at which light is diffracted. (d) Development of the ratio between aligned and tilted domains over time, for different nutrient conditions.

Figure 3.

(a) Map of the region of the IR1 chromosome where transposon insertions in IR1 affecting polysaccharide degradation and structural colour were mapped by DNA sequencing. Genes encoding known and predicted functionalities are colour-coded (from left to right): cluster of genes of a polymer utilization operon (PUL) susE, susD and susC, are represented with blue arrows. Transcriptional factor (TF) lacI23, in orange. Enzymes that participate in sugar and starch metabolism in green, maltose transporter protein, malY51; beta-phosphoglucomutase enzyme, bPGM; glycoside hydrolases family 65, 97 (alpha-glucosidase) and 13 (alpha-amylase) proteins (GH65, GH97, GH13). Glycerophosphoryl diester phosphodiesterase enzyme, GDPD, in black. Black lines above the genes indicate the position of transposon insertions. (b) Schematic overview of the biochemical pathway encoded by the relevant genes indicated in (a).

Figure 4.

Optical appearance of metabolic mutants, grown in the presence of fucoidan (ASWBF, day 1 of growth). The images in the top row show the centre of each colony, at the location where the initial cell suspension was deposited. The inset for M51 shows the edge of each colony at the same scale as the main images. Images are taken with a stereo microscope. The bottom row shows the angular resolved scattered light spectra corresponding to the images above, taken at an incident light angle of −60°.