An approach for long-term in vivo investigations on cyanobacterial cell surface changes at high spatial resolution by Atomic Force Microscopy (AFM) was developed in this study.
Until recently, changes of bacterial cell surfaces due to changes of the chemical environment could neither be investigated in situ nor in vivo. However, in vivo investigations give insights into kinetics of cell response to environmental changes and mineral nucleation at the cell's surface. Continuously cultured cyanobacteria of the representative freshwater strain Synechococcus leopoliensis (PCC 7942) were washed and artificially immobilized on poly-l-lysine-coated glass slides. Both immobilization and environmental conditions were optimized in order to facilitate long-term experiments (> 100 h) with living cells. AFM samples were investigated in situ in two different solutions: Culture medium was used for cultivation experiments and nutrient-free NaHCO3/CaCl2 solutions (supersaturated with respect to calcite) for long-term characterizations of the changes in cell surface topography. Cell viability under these conditions was investigated by AFM, TEM and epifluorescence microscopy, independently.
No indications for extended starvation were found within the relevant timescales. Analysing the influence of Ca2+ on the surface of S. leopoliensis, we found significant changes compared to a Ca-free solution. Few hours after CaCl2 was added to the circumfluent solution, small protuberances were observed on the cell surface.
These are promising results to environmental scientists for a wide range of applications, as cell response to environmental changes can now be monitored online and in vivo at timescales, which are relevant for natural processes. Most especially studies of biomineralization and mineral nucleation on bacterial cell surfaces will profit from this new approach.