Manipulation of epilithic biofilm and metabolism measurements

The aim of this project was to study the role of the different constituents of epilithic biofilm (photosynthetic organisms vs bacteria only) in the metabolism of temperate rocky shores. We sampled rock chips from a mid-intertidal sandstone area, measured the surface area covered in biofilm and placed them in a cold room (8-10 °C) under constant irradiance (PAR: 400 - 700 nm) at 290 μmolphotons.m-2.h-1 in glass containers with filtered seawater. After a 2h acclimatisation phase, we took fluorescence measurements following a 15 min dark adaptation (Fv/Fm) and then applied the following treatments (see Figure for layout): 10 mg.L-1 Simazine in 1.0 μm filtered seawater (herbicide), 25 mg.L-1 Streptomycin and 50 mg.L-1 ampicillin in 1.0 μm filtered seawater (antibiotic), 1.0 μm filtered seawater (control). 5 rockchips were placed in separate compartments in of treatment and left to incubate for 4 hours udner constant irradiance. Rock chips were then emersed again and Fv/Fm measurements taken as described above. Samples were then placed back in filtered seawater in darkness for overnight recovery and the Fv/Fm measurements taken again to see test for any longer-term effect of the treatments. The same protocol was repeated several times to ensure repeatability of the effect of the treatments. This protocol did result in consistent signficant negative effect of the Simazine on the photosynthetic capacity of the biofilm. The effect of the antibiotic mixture, however was not so consistent and could not be used reliably to significantl affect the bacteria of the biofilm. Also, it was not possible to establish by an alternative method the proportion of live/dead bacteria: it was hoped that flow cytometry should be able to provide this information but lack of available expertise did not allow to take this through. Finally, the aim was to measure the metabolism of the rock chips after treatment in the lab using a small incubating chamber and an oxygen probe. A glass chamber was design with a flat lid with a 'chimeney' in its center where the probe could be inserted. The lid was sealled to the chamber with silicon grease and the probe surrounded in parafilm to make the chamber airtight (see Figure). a magntic stirrer was placed in the chamber to avoid the formation of boundary layers. Metabolism measurements were then performed by measuring the change in oxygen concentration with time under light (for net primary productivity) and under darkness (for respiration). The setup, however, did allow allow for appropriate measurements: even though the measurements were performed in the cold room, the water temperature did not remain constant or was much higher than environmental conditions. There were also problems of water saturation that could not be resolved. Clearly, the effect of the treatments could be further explored but a more complex setup is needed to be able to perform appropriate laboratory measurements of oxygen metabolism.