Scanning electron microscopy of biofilm formation in denitrifying, fluidised bed reactors

Changes in bacterial colonisation on initially clean sand grains in an anaerobic fluidised bed reactor (FBR) were studied by scanning electron microscopy (SEM). The FBR was part of a laboratory-scale treatment system for nitrate removal from fish culture water. Denitrifying bacteria in the FBR were supplied with anaerobically digested fish feed as carbon and energy source and with nitrate as electron acceptor. Bacterial biofilm formation started rapidly with single rods settling in the fissures and cavities on the sand grainʹs surface. Rods (Pseudomonas-like and Zoogloea-like) divided and accumulated and after about 10 days were integrated in a biofilm which almost covered the entire grains. Apart from single rods, colonies of bacteria resembling Zoogloea developed from the second day onwards. After approximately 10 days, the zoogloeal colonies were overgrown by bacteria resembling Pseudomonas. A steady increase in these cells was observed, eventually leading to a situation in which the majority of grains were totally covered with a compact biofilm comprised of these bacteria. The biofilm increased in thickness with time and around day 20, sloughing was observed at parts of the biofilm surface. These sloughed areas were colonised once more with bacteria according to a similar succession pattern observed on the initially clean sand grains. The colonisation pattern on sand in the laboratory-scale FBR was compared to the colonisation on sand grains in an FBR, operating in a pilot-plant of a closed intensive fish culture system. The bacterial biofilm structure was similar in both reactors. In the pilot-plant FBR all different stages of colonisation, observed in the laboratory-scale FBR, occurred simultaneously. Also in the pilot-plant FBR, bacteria resembling Zoogloea and Pseudomonas were most abundant.